HIV TESTING MANUAL : LABORATORY DIAGNOSIS, BIOSAFETY AND

QUALITY CONTROL
NATIONAL INSTITUTE OF COMMUNICABLE DISEASES
Government of India, Directorate General of Healt Service!,
"", Samnat Mar#, Deli$%%&&'()
And
NATIONAL AIDS CONT*OL O*GANISATION
Mini!tr+ of Healt , Famil+ -elfare, Government of India,
Nirman Ba.an, Ne. Deli $ %%&&%%
1
/)0)*) 1*ASADA *AO
Addl. Secy. and Project Director
National AIDS Control
Organisation
Tel : 3017706
a! : 301"#$#
%O&'(N)'NT O INDIA
)INIST(* O +'A,T+ - A)I,* .',A('
3"" /A01 NI()AN 2+A&AN1 N'. D',+I 3
110001.
MESSAGE
O44ort5nistic in6ection are a signi6icant contri75tors o6 8or7idity
a8ongst +I& in6ected indi9id5als. )any o6 t:ese in6ections are 4re9enta7le
and s5ccess65lly treata7le 4ro9ided an acc5rate diagnosis co5ld 7e 8ade.
+I&;AIDS 4atients <it: o44ort5nistic and ot:er ende8ic in6ections 8ay not
:a9e t:e ty4ical sy84to8s 4at:ogno8ic o6 a 4artic5lar in6ection. So1
la7oratory diagnosis 7eco8es i84ortant in 8anage8ent o6 t:ese 4atients.
Acc5rate treat8ent <ill go a long <ay in alle9iating t:e 4ain and s566erings
o6 +I& in6ected <it: o44ort5nistic in6ections.
NICD :as ta=en t:e initiati9e to 4re4are a 8an5al on >,a7oratory
Diagnosis o6 Co88on O44ort5nistic In6ections?. T:e 8an5al <ill :el4 t:e
la7oratory s4ecialists and tec:nologists in diagnosis o6 in6ections associated
<it: +I&;AIDS.
)any o6 t:e s4ecialists <or=ing <it: +I&;AIDS 4atients 8ay not 7e
con9ersant <it: so8e o6 t:ese in6ections as t:ey did not a66lict :58an 7eings
7e6ore t:e era o6 +I&.
@/) 0) *) 1ra!ada *aoA
FO*E-O*D
T:e AcB5ired I885no De6iciency Syndro8e @AIDSA <as 6irst
recognised in t:e Cnited States in 1DE1. T:e etiological agent1 +58an
I885no de6iciency &ir5s @+I&A1 <as isolated in 1DE" and la7oratory
test 6or +I& 7eca8e co88ercially a9aila7le1 t:erea6ter. T:is ena7led
:ealt: care <or=ers to diagnose +I& in6ection1 indi9id5als to =no<
<:et:er or not t:ey <ere in6ected1 and 8ost i84ortantly 8edical
2
science to 5nderstand +I&;AIDS 7etter. T:e a9aila7ility o6 t:ese tests
:as also led to 8isconce4tions regarding t:e role o6 +I& testing.
Co84ared to any ot:er ty4e o6 disease1 t:e iss5es related to t:e
diagnosis o6 +I& in6ection are 6ar 8ore co84le!. AIDS is in9aria7ly
6atal and in6ection is li6elong. No dr5gs are a9aila7le to c5re AIDS or to
render an +I& in6ected 4erson1 non3in6ectio5s. Since +I& is s4read
8ainly t:ro5g: se!5al contact1 indi9id5al =no<n to 7e in6ected <it: +I&
are 5n6ort5nately o6ten stig8atised and discri8inated against. S5c: a
sit5ation res5lts 6ro8 t:e lac= o6 4ro4er 5nder3standing regarding t:e
8ode o6 trans8ission. )any indi9id5als1 4artic5larly :ealt: care
<or=ers1 see8 to 7elie9e t:at t:e s4read o6 +I& can 7e controlled 7y
identi6ying 4eo4le <it: +I& in6ection1 and testing o6 :os4ital 4atients is
o6ten ad9ocated. T:is is 8ainly d5e to ignorance a7o5t t:e o7jecti9es o6
+I& testing. A 8an5al on +I& testing <ill 7e :el465l in i84arting
adeB5ate training to 8edical 6raternity and 4ara 8edical <or=ers.
T:e la7oratory 4lays an i84ortant role in t:e National AIDS
Control Progra88e. It can assist 4rogra88e 8anagers in st5dying t:e
trend o6 e4ide8ic in t:e co885nityF ens5res sa6e 7lood1 6ree 6ro8 +I&
in6ection1 and 8ay assist 4:ysicians to assess t:e 4rognosis o6 t:e
in6ection. A4art 6ro8 t:is1 it can assist t:e indi9id5al to 8odi6y :is
7e:a9io5r or li6e style to 4re9ent t:e s4read o6 +I& in t:e co885nity
and to i84ro9e t:e B5ality o6 :is li6e.
To 6acilitate t:e training o6 la7oratory sta66 in +I& Testing
Proced5res1 a re6erence 8an5al <as 9ery 85c: needed to i84ro9e t:e
s=ill o6 la7oratory sta66 and ot:er 65nctionaries in9ol9ed in +I&;AIDS
4re9ention and control 4rogra88e.
It is e!4ected t:at t:e 4resent 8an5al <:ic: <as 4re4ared 5nder
t:e s54er9ision o6 t:e National Instit5te o6 Co885nica7le Diseases <ill
65l6il t:is need.
@/) 0) *) 1ra!ada *aoA
D*) S)1) AGGA*-AL
Director %eneral o6 +ealt:
Ser9ices
Tel : 301E"3E
a! :
%O&'(N)'NT O INDIA
)INIST(* O +'A,T+ - A)I,* .',A('
NI()AN 2+A&AN1 N'. D',+I 3 110001.
D*AFT FO* A11*O0AL
FO*E-O*D
+I&;AIDS are a 8ajor concern o6 :ealt: care 4ro6essionals all o9er t:e <orld.
,a7oratory diagnosis is t:e only 8et:od o6 de6ining;esta7lis:ing +I& stat5s o6 an
indi9id5al. India :as <ell de9elo4ed la7oratory a44roac:es 6or : ens5ring 7lood
sa6ety and donation @tiss5e1 organsA sa6ety F s5r9eillance o6 :ig: ris= gro54s F
sentinel s5r9eillance F diagnosis and researc:.
3
)ost o6 +I& anti7ody screening and s544le8ental @So8e ,a7s. onlyA testing is
4er6or8ed at t:e local le9el @G2TCs1 S5r9eillance Centres 5nder NACOA1 <:ile
centralised @re6erralA la7s. 4ro9ide re6erence diagnostic ser9ices 6or 4ro7le8atic
sera and 4er6or8 so4:isticated tec:niB5e li=e ..2.1 PC(1 9iral load assay and 9ir5s
c5lt5re etc.
(egardless o6 t:e le9el o6 t:e la7oratory in9ol9ed in +I& testing1 t:e training and
e!4ertise o6 sta66 8e87ers1 t:e ty4e o6 reagents and in6rastr5ct5ral 6acilities
a9aila7le are 8ajor deter8inants o6 t:e B5ality o6 test res5lts 4rod5ced. ,a7oratory
4ersonnel1 one and all1 85st stri9e 6or total B5ality ass5rance 4rogra88e1 seeing to
it t:at e9ery ste4 o6 t:e testing 4rocess o9er <:ic: t:ey :a9e control is 8onitored
and 9eri6ied 6or acc5racy and 4recision. .it:in t:e la7oratory1 t:e 4ersonnel 85st
6eel con6ident t:at t:ey are 4rotected 6ro8 inj5ry so t:at t:ey can do t:eir 7est <or=
sa6ely1 <it:o5t 6ear and distraction.
T:is 8an5al 4ro9ides in6or8ation t:at goes 7eyond 4ractical as4ects o6 la7oratory
analysis o6 +I&1 incl5ding t:e 7iology o6 t:e +I& 9ir5s1 t:e 4at:o7iology o6 +I&
in6ection in 8an1 t:e e4ide8iology o6 +I& in6ection in India and B5ality assess8ent
o6 test syste8s and la7oratories. T:e 8an5al t:ere6ore 4ro9ides a co84re:ensi9e
doc58ent not only 6or 5se at t:e 7enc: 75t also 6or teac:ing and training o6 +I&
@Dr) S)1) A##ar.alA
4
1*EFACE
+I& in6ection is s4reading ra4idly in India. T:e in6ection can only 7e
detected 7y la7oratory tests1 as t:ere is a long asy84to8atic 4eriod
<:en t:e indi9id5al is in6ectio5s and can s4read disease 75t :as no
s4eci6ic sy84to8s or signs o6 disease. T:e role o6 t:e la7oratory is 9ery
i84ortant and it is essential t:at t:e :ig:est standards are not only
8aintained in eac: la7oratory 75t are also reg5larly 8onitored.
T:ere are a n587er o6 diagnostic 4roced5res and 9ario5s ty4es o6
co88ercial diagnostic =its a9aila7le. 8oreo9er t:e res5lts o6 t:e
la7oratory tests can 7e in6l5enced 7y 8any 6actors. It is essential t:at
eac: la7oratory :as a Standard O4erati9e Proced5re @SOPA <:ic:
details eac: ste4 o6 t:e test incl5ding 7io3sa6ety 4reca5tions. T:e
la7oratories are also e!4ected to 6ollo< t:e national g5idelines
circ5lated 7y NACO on et:ical iss5es related to +I& testing.
T:e +I& Testing )an5al :as 7een 4re4ared as a 4ractical g5ide 6or
la7oratory 4ersonnel. T:e )an5al is co84re:ensi9e and co9ers rele9ant
to4ics o6 interest. T:e c:a4ters :a9e 7een dra6ted 7y national e!4erts.
T:e 8aterial :as 7een e!tensi9ely re9ie<ed at t<o 8eetings :eld at
NICD.
Dr. Cs:a H. 2a9eja1 Cons5ltant and +ead +I&;AIDS Di9ision and :er
colleag5es Dr. D. C:atto4ad:ya and S:. (a8es: Aggar<al deser9e
credit 6or ta=ing t:e initiati9e 6or t:e 4re4aration o6 )an5al and :ard
<or= 45t in 7y t:e8 to gi9e it s:a4e in a record ti8e. T:e National
Instit5te o6 Co885nica7le Diseases is also o7liged to all t:e e!4erts
<:o <illingly contri75ted t:e c:a4ters.

2 /otna So3e+ 4
Director
National Instit5te o6 Co885nica7le Diseases
Dated : 0";06;DD
5
ACKNOWLEDGEMENTS
The HIV Testing Manual : Laboratory Diagnosis, Biosafety and
Quality Control is intended for use by different ategories of HIV testing
laboratories!enters all o"er India as a referene and training #anual$
The #anual an be referred to, to %re%are standard o%erati"e %roedures
&'()s* and to %ratise biosafety and +uality ontrol et$, so that eah HIV
testing faility #ay unifor#ally %rodue +uality results ,ithin the
li#itations of the test$
This #anual does t,o things ,hih ha"e not been done before$
-irst it gi"es the u%dated ba.ground .no,ledge on HIV and other
transfusion trans#itted infetions along ,ith tehni+ues$ 'eond, it ta.es
ad"antage of the e/%ert .no,ledge of authorities by %resenting
ontributions fro# nearly 01 ontributors fro# different %arts of the
ountry$
I #ost gratefully a.no,ledge the tehnial guidane and su%%ort
fro# Dr$ '$)$ 2ggar,al, Diretor 3eneral of Health 'er"ies, 3o"ern#ent
of India, Ministry of Health 4 -a#ily 5elfare$
I e/%ress #y sinere gratitude and than.s to 'hri )rasada 6ao,
2dditional 'eretary and )ro7et Diretor, 82C( for enourage#ent,
finanial su%%ort and for ,riting the fore,ard for the #anual$ I a#
than.ful to Diretor, 8ICD, Ms$ 9otna 'o.hey for her enourage#ent,
su%%ort and onstruti"e o##ents during the disussions$ I than. Dr$
'udershan :u#ari, 6egional 2d"isor, Laboratory 'er"ies 5H(!';26(
for her e/%ert guidane and for %ro"iding the rele"ant literature$
I a# #ost grateful to all of the tehnial e/%erts ,ho ontributed
"arious ha%ters in a ti#ely #anner$ I es%eially than. Dr$ )$ L$ 9oshi, Dr$
L$ Dar, Dr$ 6$ 8$ Ma.roo, Dr$ :$ 6ay, Dr$ '$ )$ Tri%athi for their "aluable
o##ents and suggestions during the #eetings held to re"ie, the rough
draft$ I ,ish to than. #y olleagues Dr$ D$ Chatto%adhya, 'hri 6$ :$
2ggar,al and others for their ontributions and hel% during the t,o
#eetings for the re"ie, of the #anual$
-inally, I e/%ress #y a%%reiation and than.s for the seretarial
assistane %ro"ided by '#t$ <sha and 'h$ Mu.esh :u#ar$
Dr. (Mrs.) Ush K. B!"#
Consultant &Miro* 4 Head
Di"ision of 2ID'
NICD1 Del:i
6
Contributors
1. Dr. Pradee4 Set:
Pro6essor and +ead1
De4art8ent o6 )icro7iology
All India Instit5te o6 )edical Sciences1
Ne< Del:i 3 1100#D.
#. Dr. Cs:a H 2a9eja
Cons5ltant @)icroA and +ead
AIDS Di9ison1
National Insit5te o6 Co885nica7le
Diseases1 ##1 S:a8 Nat: )arg1
Del:i 3 1100$".
3. Dr. Hris:na (ay
Cons5ltant @)icroA and +ead
STD ,a7oratory
Sa6darjang +os4ital1 Ne< Del:i
". Dr. (. N. )a=roo
Senior Cons5ltant
Trans65sion )edicine1
A4ollo +os4ital1 Sarita &i:ar1 Ne<
Del:i
$. Dr. P. ,. Ios:i
It. Director @Tec:nicalA
National AIDS Control Organisation
@NACOA
)inistry o6 +ealt: and a8ily .el6are
Nir8al 2:a<an1 Ne< Del:i 3 110011.
6. Dr. (. N. Prasad
Senior Scientist and +ead
De4art8ent o6 2lood Prod5cts and
)icro7iology1
National Instit5te o6 2iologicals1Ne<
Del:i
7. Dr. D. C:atto4ad:ya1
Ioint Director1
AIDS Di9ision1 NICD1 Del:i 3 1100$".
E. Dr. ,alit Dar
Associate Pro6essor1
De4art8ent o6 )icro7iology1
AII)S1 Ne< Del:i 3 1100#D.
D. Dr. S. N. )is:ra
Cons5ltant1
NACO
) + - . ..1 Ne< Del:i 3 110011
10
.
Dr. )o:d. S:a5=at
De45ty Director1
NACO1
) + - . ..1 Ne< Del:i 3 110011
11. Dr. 2. 2. (e<ari
Senior P:ysician1
(.).,. +os4ital1 Ne< Del:i
1#
.
Dr. 2rajac:and Sing:
Associate Pro6essor1
De4art8ent o6 )icro7iology
Instit5te o6 )edical Sciences1
I84:al
13. S:ri (. H. Aggar<al1
Assistant Director
AIDS Di9ision
NICD1 Del:i
1"
.
Dr. (. Paranja4e
De45ty Director
National AIDS (esearc: Instit5te
P5ne1 India.
1$. Dr. S. H5l=arni
National AIDS (esearc: Instit5te
P5ne1 India.
16
.
Dr. S. P. Tri4at:y
National AIDS (esearc: Instit5te
P5ne1 India.
17. Dr. *.%.H. Sing:
AII)S1 Ne< Del:i
1E
.
Dr. %eorge 2ittra
Sa6darjang +os4ital1 Ne< Del:i
7
1a#e No)
TABLE OF CONTENTS 6 3 D
Ca5ter A6tor
1. Nat5ral :istory1 7asic e4ide8iology incl5ding Dr. P. ,. Ios:i
10 3 17
glo7al and co5ntry scenario o6 +I&;AIDS Dr. ). S:a5=at
#. &irology Dr. Cs:a H. 2a9eja 1E 3 #"
3 T:e :58an i885node6iciency 9ir5s
3 (e4lication
3 S5sce4ti7ility
3 Trans8ission
3. I885no4at:ology Dr. Cs:a H. 2a9eja #$ 3 30
3 Pri8ary in6ection illness
3 Clinically latent 4eriod
3 AIDS
3 )ec:anis8 o6 CD" cell dys65nction
3 T:e co5rse o6 4rogression o6 +I& in6ection
". 2iosa6ety in la7oratories Dr. Hris:na (ay 31 3 "E
3 (is= 4roced5res
3 Cni9ersal 4reca5tions
3 SteriliJation and disin6ection
3 Sa6e dis4osal o6 la7oratory <astes
$. Collection1 trans4ort and storage o6 s4eci8ens Dr. Hris:na (ay
"D 3 $1
6or +I& testing
3 Collection o6 7lood
3 Se4aration o6 ser58
3 Trans4ort o6 s4eci8en
3 Storage o6 s4eci8en
6. +I& anti7ody testing <it: s4ecial re6erence Dr. Cs:a H. 2a9eja
$# 3 6D
to +I&31
3 P5r4ose o6 +I& testing
3 Hinetics o6 :58oral i885ne res4onse
3 In6or8ed consent
3 Con6identiality
3 Detection o6 +I& s4eci6ic anti7odies
3 Screening tests
3 ',ISA
3 (a4id
3 Si84le
3 S544le8ental tests
3 .estern 7lot
3 ,ine I885noassay
3 C:oice o6 screening assay
3 O7jecti9e o6 testing
3 Sensiti9ity o6 test
3 S4eci6icity o6 test
3 Pre9alence o6 +I& in6ection in 4o45lation
8
3 Cost e66ecti9eness
3 A44ro4riateness to strategy o6 testing and
in6rastr5ct5re a9aila7le
7. Di66erential 6eat5res in e4ide8iology1 Dr. 2rajac:and Sing:
70 3 76
nat5ral :istory1 9irology1 4at:ogenesis and
la7oratory diagnosis o6 +I&3# in6ection
3 '4ide8iology
3 &irology
3 Nat5ral :istory and 4at:ogenesis
3 D5al assays
3 Co87ined assays
3 Assays 6or detection o6 +I&3# anti7odies
E. Direct e9idence o6 +I& in6ection Dr. (. Paranja4e 77 3 E3
3 Poly8erase c:ain reaction Dr. S. P. Tri4at:y
3 4#" antigen assay Dr. S. H5l=arni
3 Isolation o6 +I&
3 (eagents 6or PC(
3 'B5i48ents 6or PC(
D. +I& testing 4olicy Dr. S. N. )is:ra E" 3 D3
3 O7jecti9es o6 testing Dr. P.,. Ios:i
3 +I& testing strategy
3 5nlin=ed anony8o5s
3 9ol5ntary con6idential
3 8andatory testing
3 )andatory testing 9ers5s standard
4reca5tions in a :ealt: care settings.
3 +I& testing in a :ealt: care setting incl5ding
legal and et:ical considerations
3 ,ist o6 7lood testing and re6erence centres @NACOA
10. K5ality ass5rance in +I& testing Dr. Pradee4 Set:
D" 3 10E
3 Necessity and i84ortance
3 %5idelines to i84ro9e
B5ality o6 testing
3 '!ternal B5ality control
o6 la7oratory
3 (ecord =ee4ing
3 ,a7oratory as4ects o6
B5ality control
11. '9al5ation o6 +I& =its and 4re4aration Dr. Cs:a H. 2a9eja
10D 3 11$
o6 ser58 4anel Dr. D. C:atto4ad:ya
3 Pre4aration o6 ser58 4anel
3 Co84osition o6 4anel S:. (. H. Aggar<al
3 (e6erence test
3 Testing conditions @sensiti9ity1 s4eci6icity etc.A
1#. ,a7oratory tests 6or 8onitoring stage Dr. ,alit Dar
116 3 1#6
and 4rogression o6 +I& in6ection Dr. *. %. H. Sing:
3 &iral 8ar=ers
3 S5rrogate 8ar=ers
3 Cost 6actor and a9aila7ility o6 reagents
9
13. Trans65sion trans8itted diseases e!ce4t +I& Dr. (.N. )a=roo
1#7 3 136
in6ection
3 ,ist o6 diseases
trans8itted t:ro5g: 7lood
3 Trans65sion associated :e4atitis 9ir5ses
@+A&1 +2&1 +C&1 +D&1 +%&A
3 )alaria
3 Sy4:ilis
3 +58an T cell ly84:otro4ic 9ir5ses
@+T,&I and +T,&IIA
3 Cyto8egalo9ir5s @C)&A
1". In6ections trans8itted t:ro5g: 7lood 4rod5cts Dr. (. N. Prasad
137 3 1"3
3 ,ist o6 in6ections
3 2acterial conta8ination o6 7lood co84onents
and 7lood 4rod5cts
3 Pre9ention o6 in6ections associated <it:
trans65sion o6 7lood co84onents and
7lood 4rod5cts
1$. Co5nselling and +I& testing Dr. %eorge 2ittra
1"" 3 1"6
3 De6inition
3 Di66erence 7et<een co5nselling and :ealt:
ed5cation
3 O7jecti9es o6 +I&;AIDS co5nselling
3 .:y is co5nselling necessary
3 .:o reB5ires co5nselling L
3 .:ere can co5nselling 7e 4ro9ided
3 .:o can 4ro9ide co5nselling
3 Ty4es o6 co5nselling
3 Contents o6 4re9enti9e co5nselling.
3 Co5nselling
16. In6rastr5ct5re 6or +I& testing la7oratories Dr. Cs:a H. 2a9eja
1"7 3 1$0
3 Categories o6 +I& testing la7oratories ; Dr. D. C:atto4ad:ya
centres along <it: jo7 de6initions S:ri. (. H. Aggar<al
3 Screening la7oratories in 7lood 7an=s
3 &ol5ntary 7lood testing and s5r9eillance la7oratories
3 National +I& re6erence centres
3 ,a7oratory 8onitoring stage and 4rogression o6 +I& in6ection
17. Post e!4os5re 4ro4:yla!is @P'PA g5idelines Dr. 2. 2. (e<ari
1$1 3 1$7
3 De6inition o6 occ54ational e!4os5res Dr. S. N. )is:ra
3 Ste4s to 7e ta=en on e!4os5re to +I&
in6ected 7lood ; 7ody 6l5ids
3 Ty4es o6 occ54ational e!4os5res 6or <:ic: P'P
is reco88ended
3 Deter8ination o6 e!4os5re code
3 Deter8ination o6 +I& stat5s code
3 Pre and 4ost test co5nselling and testing
3 Dr5gs reco88ended 6or P'P
3 D5ration 6or <:ic: dr5gs need to 7e ta=en
3 Pregnancy and P'P
10
3 Sa6ety and side e66ects o6 dr5gs
3 Ste4s to 7e ta=en 7y t:e in6ection control o66icer
1E. ',ISA tests <:ic: :a9e 7een e9al5ated 7y .+O .+O doc58ent
1$E 3 161
colla7orating centre on AIDS in t:e De4art8ent
o6 )icro7iology1 Instit5te o6 Tro4ical )edicine1
Ant<er41 2elgi58
1D. ,ist o6 re6erences 6or 65rt:er st5dy
16#
=> )rofor#as to be used for sending re%orts to 0?@ A
0?B
Diretor, 'tate 2ID' 'oiety and
Additional Director Tec:nical1
National AIDS Control Organisation1 Ne< Del:i
,ist o6 6ig5res and anne!5res
1703171
11
$. BASIC E%IDEMIOLOGY AND NATURAL HISTORY
OF HIV&AIDS
$.$ I'(r)*+,(-)' : The 2+uired I##unodefiieny 'yndro#e &2ID'* is
aused by Hu#an I##unodefiieny Virus &HIV*$ It is a serious disorder
of the i##une syste# in ,hih the bodyCs nor#al defenes against
infetion brea. do,n, lea"ing it "ulnerable to a host of life threatening
infetions, inluding unusual #alignanies$
$.. Th" %r)/0"1 :
$...$ G0)/0 : By the end of 0BB1, aording to esti#ates fro# the 7oint
<nited 8ations )rogra##e on HIV!2ID' &<82ID'* and the 5orld Health
(rganiDation &5H(*, the nu#ber of %eo%le li"ing ,ith HIV ,ill ha"e
gro,n to @@$E #illion, 0>F #ore than 7ust one year ago$ The e%ide#i
has not been o"ero#e any,here$ Virtually e"ery ountry in the ,orld
has seen ne, infetions in 0BB1 and the e%ide#i is fran.ly out of ontrol
in #any %laes$
2ording to <82ID'!5H( esti#ates, 00 #en, ,o#en and hildren
around the ,orld ,ere infeted %er #inute during 0BB1 &near to ?
#illion*$ More than BGF of all HIVAinfeted %eo%le no, li"e in the
de"elo%ing ,orld, ,hih has li.e,ise e/%eriened BGF of all deaths to
date fro# 2ID', largely a#ong young adults ,ho ,ould nor#ally be in
their %ea. %roduti"e and re%roduti"e years$
'ubA'aharan 2fria is ho#e to H>F of the %eo%le ,ho bea#e infeted
during last year &0BB1*$ 2fria, the global e%ienter, ontinues to d,arf
the rest of the ,orld on the 2ID' balane sheet$ 'ine the start of the
e%ide#i, 1@F of all 2ID' deaths so far ha"e been in this region$ 2t
least BGF of all 2ID' or%hans ha"e been 2frian$ 5hile no ountry in
2fria has esa%ed the "irus, so#e are far #ore se"erely affeted than
others$ The 'outhern %art of the 2frian Continent holds the #a7ority of
the ,orldCs hard hit ountries$ In Bots,ana, 8a#ibia, ',aDiland and
Ii#bab,e, esti#ates sho, that bet,een =>A=?F of %eo%le aged 0GAEB
are li"ing ,ith HIV or 2ID'$ HIV %re"alene in %regnant #others ,as
found to be o"er =>F in #a7ority of sentinel sites in this ountry$
Inreasingly the s%otlight is on the s%read of HIV through the 2sian
Continent, es%eially in 'outh 2sia and ;ast 2sia, ,hile rates re#ain lo,
relati"e to so#e other regions, ,ell o"er H #illion 2sians are already
infeted and HIV is learly beginning to s%read in earnest through the
"ast %o%ulations of India and China$
In ;astern ;uro%e and in Latin 2#eria and the Caribbean, infetions are
onentrated in #arginaliDed grou%s though, learly not li#ited to the#$
'o#e studies suggest that u% to @>F of #en ,ho ha"e se/ ,ith #en
#ay be infetedJ a#ong drug infetions in 2rgentina and BraDil the
%ro%ortion #ay be lose to half of abo"e$
In 8orth 2#eria and 5estern ;uro%e, ne, o#binations of antiAHIV
drugs ontinue to redue 2ID' deaths signifiantly$ Ho,e"er, beause
ne, infetions ontinue to our ,hile antiretro"iral drug o.tails .ee%
already infeted %eo%le ali"e, the %ro%ortion of the %o%ulation li"ing ,ith
HIV has atually agro,n$
Table 0 sho,s the regional HIV!2ID' statisti and features$
12
T/0" $ R"2-)'0 HIV&AIDS s((-s(-,s '* 3"(+r"s, D","1/"r $445
R"2-)' E6-*"1-
,
s(r("*
A*+0(s 7
,h-0*r"'
0-!-'2
8-(h
HIV&AID
A
A*+0(s 7
,h-0*r"'
'"809
-'3",("*
8-(h HIV
A*+0(s
6r"!0"'
,"
r(" (:)
6"r,"'(
)3 HIV;
6)s-(-!"
*+0(s
8h) r"
8)1"'
M-' 1)*"
(s) )3
(r's1-ss-
)' (<)
3)r *+0(s
0-!-'2
8-(h
HIV&AIDS
'ubA
'aharan
2fria
Late
KH>sA
early
K1>s
==$G
#illion
E$> #illion 1$>F G>F Hetero
8orth 2fria
4 Middle
;ast
Late K1>s =0> >>> 0B >>> >$0@F =>F ID<, Hetero
'outh 4
'outhA;ast
2sia
Late K1>s ?$H #illion 0$= #illion >$?BF =GF ID<, Hetero
M'M
;ast 2sia
4 )aifi
Late K1>s G?> >>> =>> >>> >$>?1F 0GF M'M,
ID<, Hetero
Latin
2#eria
Late
KH>sA
early
K1>s
0$E #illion 0?> >>> >$GHF =>F Hetero,
M'M
Caribbean Late
KH>sA
early
K1>s
@@> >>> EG >>> 0$B?F @GF Hetero,
M'M
;astern
;uro%e
4 Central
2sia
early
KB>s
=H> >>> 1> >>> >$0EF =>F ID<, M'M
5estern
;uro%e
Late
KH>sA
early
K1>s
G>> >>> @> >>> >$=GF =>F M'M, ID<
8orth
2#eria
Late
KH>sA
early
K1>s
1B> >>> EE >>> >$G?F =>F M'M,
ID<, Hetero
2ustralia
4 8e,
Iealand
Late
KH>sA
early
K1>s
0= >>> ?>> >$0F GF M'M, ID<
TOTAL ==.>
1-00-)'
?.5 1-00-)' $.$@ >=@
13
M T:e 4ro4ortion o6 ad5lts @1$ to "D years o6 ageA li9ing <it:
+I&;AIDS in 1DDE1 5sing 1DD7 4o45lation n587ers.
N )S) @se!5al trans8ission a8ong 8en <:o :a9e se! <it: 8enA1
IDC @trans8ission t:ro5g: injecting dr5g 5seA1 +etero @:eterose!5al
trans8issionA.
<82ID'!B1$@G 5H(!;MC!VI6!B1AEA5H(!2'D!B1$@
$.... I'*- : 'ine the detetion of HIV infetion in o##erial se/ ,or.ers
&C$'$5s*
in Ta#il 8adu in 0B1?, the infetion is gro,ing "ery fast in the ountry$ 2s on
9anuary 0BBB, 1=H10 HIV infeted %ersons and ?H>@ 2ID' ase ,ere
re%orted to 82C($ 2lthough the #agnitude of disease has been found to be
"aried in "arious %arts of the ountry, the states of Maharashtra, Ta#il 8adu,
2ndhra )radesh, :arnata.a and Mani%ur are hard hit states$ Maharashtra
aounts for lose to G>F of all re%orted HIV and 2ID' ases in India$
'entinel sur"eillane data olleted during 2ugustA(tober, 0BB1 re"ealed
that in the abo"e #entioned 'tates the infetion in %regnant #others, in high
ris. grou%s li.e 'TD lini attenders, C$'$5s, in7eting drug users et$ has
sho,n gallo%ing inreases$ Moreo"er, the esti#ated nu#ber of HIV infetions
in the ountry ha"e been re%orted to be @AG #illion$ -ig$ 0 through ? sho,
the HIV!2ID' features and statistis in India$
$.= E6-*"1-)0)2-,0 3"(+r"s :
$.=.$ A2"'( 3,()rs :
i* A2"'( : HIV, the "irus ,hih auses 2ID' is a lenti"irus, one of sub
fa#ily of retro"iruses$ They ha"e a uni+ue enDy#e, re"erse
transri%tase, ,hih o%ies the "iral ribonulei aid &682* into
deo/yribonulei aid &D82*, ,hih e"entually integrates into host ell
hro#oso#e$ Hene, HIV %ersists ,ithin ells for years and annot be
eradiated fro# host ells ,ith any of the urrently a"ailable antiA
retro"iral drugs$ There are t,o ty%e of HIVJ HIVA0 and HIVA=$ Both ha"e
#any strains$
ii* R"s"r!)-r )3 -'3",(-)' : )eo%le harbouring HIV in their body are the
reser"oir of infetion$ They #ay be asy#%to#ati healthy arriers or full
blo,n 2ID' ases$ 2ording to latest obser"ations in de"elo%ed
ountries, G>F of %eo%le ,ith HIV infetion are li.ely to de"elo% 2ID'
,ithin 0> years after first beo#ing infeted$ In de"elo%ing ountries the
inter"al bet,een infetion and disease is %robably shorter$
iii* S)+r," )3 -'3",(-)' : Conta#inated blood, se#en, "aginal fluids are the
%ri#e soure of infetion$ Conta#inated breast #il. #ay be soure of
infetion for trans#ission fro# #other to hild$ 'ali"a and tears are not
onsidered to be e%ide#iologially i#%ortant soures of infetion$
$.=.. H)s( F,()rs :
i* A2" '* s"A : Loung %eo%le are dis%ro%ortionately affeted by HIV
and 2ID'$ 2round half of the ne, HIV infetions are in %eo%le aged 0GA=E
years, the range in ,hih #ost %eo%le start their se/ual li"es$ In 8orth
2#eria, ;uro%e and 2ustralia H> %erent ases are ho#ose/uals or
bise/ual #en$ In 2fria the %iture is "ery different$ 2ID' ours al#ost
as fre+uently a#ong fe#ales as #ales$
ii* H-2h R-sB Gr)+6s : HIV %re"alene in ertain %o%ulations is an
i#%ortant fator in deter#ining on ,hih target %o%ulations the
%rogra##eCs efforts and resoures should fous$ If esti#ated le"el of
14
infetion in targeted %o%ulations, suh as in7eting drug users is high , it
is #ore li.ely that they ,ill infet eah other and also their se/ual
%artners$ 'o, this %o%ulation ,ill need to be foused for targeted
inter"ention$ 5hen %re"alene is high in a ountry, hild bearing age
beo#es #ore at ris. as also the %o%ulation reei"ing blood transfusions$
a* %r"s"'," )3 STD : There is strong e"idene that #en and ,o#en ,ith
genital uler disease or uerthral disharge are at inreased ris. of
a+uiring and trans#itting HIV$ If there is data suggesting high
%re"alene of 'TD in a %o%ulation, this ,ould be an influening fator for
inreased ris. of HIV infetion$
b* Fr"C+"',9 )3 "A6)s+r" : The %robability that a %erson has beo#e
infeted ,ith HIV se/ually is, in general, %ro%ortional to the fre+ueny of
un%roteted se/ ats and nu#ber of high ris. %artners ,ith ,ho# the
%erson has had se/ual ontat in reent years$
* M-A-'2 6(("r' )3 6)6+0(-)' : The ,ay &ris.* beha"iours are #i/ed
a#ong grou%s of %eo%le an be "ery une"enly distributed$ -or e/a#%le,
in7eting drug users #ight only share in7eting ,or.s ,ithin their o,n
grou%s but ha"e se/ual %artners that are both ,ithin and outside their
identified grou%s$
d* I11+'-(9 : It ,as found that the 2ID' "iti#s had nor#al B ell
funtion, that is their antibody le"els ,ere nor#al or ele"ated$ But their
antibodies ,ere of non neutraliDing "ariety ,hih ha"e no de#onstrable
effet on the "irus$ Ho,e"er, their T ell funtion ,as far fro# nor#al$
In a healthy i##une syste#, s%eialiDed TAells alled hel%er T ells
&CDE* assist B ells and antibodies to fight infetion ,hile their ounter
%arts, su%%ressor T ells inhibit this ati"ity$ Healthy indi"iduals ha"e
t,ie as #any hel%er ells as su%%ressor ells$ In 2ID' %atients, the
ratio is re"ersed$ (ne of the #ost stri.ing features of the i##une
syste# of %atients ,ith 2ID' is %rofound ly#%ho%aenia ,ith total
ly#%hoyte ount often belo, G>>!$##$
$.> %)0-(-,0 '* ,+0(+r0 3,()rs :
i* A,,"6(/-0-(9 )3 ,"r(-' -'*-2"')+s s"A+0 6r,(-,"s: Certain
indigenous se/ual %raties #ay be ulturally ae%table and these #ay
ontribute to HIV trans#ission$ -or e/a#%le, dry se/ #ay be a o##on
ae%ted %ratie but ontributes to HIV trans#ission$ 2lso, the national
2ID' %rogra##e #ay not be able to %ro#ote ertain safer se/ual
%raties if they are not ulturally ae%table$
ii* Wr '* ,-!-0 *-s(+r/'," : These li#it the regular i#%ortation of
o##odities, suh as 'TD treat#ent drugs, ondo#s and HIV testing
.its$ 6adio and tele"ision #essages %ro#oting safer se/, ondo#
%ro#otion, either #essages #ay not go as %lanned$
iii* L-1-((-)'s )' -'("r!"'(-)'s : These hinder the %rogress of %re"ention
inter"entions suh as distribution of ondo#s to youth or %rostitutes$
i"* S),-0 +',,"6('," )3 ,)'*)1s: This #ay be a deter#inant of ris. in
ertain %o%ulations$ 'uh %o%ulations are at high ris. of trans#ission
through unsafe se/ual %raties$
"* W)1"'Ds s((+s : May li#it ,o#enCs ability to %ratie safer se/, for
e/a#%le, ,o#en #ight not be in a %osition to hoose or #a.e deisions
suh as ondo# use ,ith their %artners$ If a ,o#an is eono#ially
15
de%endent on her %artner, it is es%eially diffiult to influene hi# to use
ondo#s$
"i* N(-)'0 6)0-,-"s : That ser"e as barriers to the i#%le#entation of
i#%ortant inter"entions for e/a#%le, restrited a"ailability of needles and
syringes ,ould li#it the usefulness of an inter%osition to %ro#ote safe
drug in7etion %raties$
"ii* N)r1s '* 6r,(-,"s : Can inrease the ris. of beo#ing infeted in
ertain %o%ulations$ -or e/a#%le, sharing needles to belong to a grou%
of drug in7etors #ight be a o##on ritual that needs to be onsidered
before designing %re"ention inter"entions for the drug in7eting
o##unity$ 2lso, it #ay not be ulturally ae%table to disuss
ho#ose/uality$
"iii* C+0(+r" '* "(h'-, 6r,(-,"s : 'uh as iru#ision in #ales and
fe#ales, tattooing, and sarifiation #ay be ,ell ae%ted but an also
ontribute to the ris. of beo#ing infeted in ertain %o%ulations beause
of use of %oorly steriliDed %iering e+ui%#ent$
i/* Mr2-'0-s"* 6)6+0(-)' : ;ono#ially de%ressed %o%ulations #ay
ne"er be able to benefit fro# %re"ention efforts beause the soial
syste# refuses to reognise the#$ ;/a#%le of suh %o%ulations are
%rostitutes and in7eting drug users$
$.? S),-0 '* ",)')1-, 3,()rs :
i* L)8 0-("r,9 : May li#it aess to ,ritten, ris. redution infor#ation$
ii* Ur/'-E(-)' : -or eono#i reasons #any %eo%le #ay #o"e to the
larger ities, ,here they #ay indulge in high ris. beha"iours suh as
o##erial se/ and in7eting drug use et$
iii* I16r-s)'1"'( : May restrit #enCs aess to ,o#en and enourage
#en to ha"e se/ ,ith #en$
i"* H-2h 1)/-0-(9 : (f ertain target %o%ulations inreases the geogra%hi
s%read of HIV trans#ission$ -or e/a#%le, tru. dri"ers #ay inrease the
s%read by engaging in se/ ,ith %rostitutes at se"eral tru. sto%s$
"* M-2r(-)' '* s"6"r(-)' 3r)1 31-0-"s : Industries suh as fishing,
tru.ing and #ining #ay fore %eo%le to tra"el to another ountry or
region of the ountry to find ,or.$ The resulting se%eration fro# fa#ilies
and situations #ay dri"e the# to %rostitution and asual se/$
"i* Dr+2 +s" : Drug use #ay i#%air 7udge#ant and li#it the ability to
%ratise safer se/$
"ii* A0,)h)0 +s" : 2lohol use #ay i#%air 7udge#ent and li#it the ability to
%ratise safer se/$
$.F M)*" )3 HIV (r's1-ss-)' : ;%ide#iologial studies throughout the
,orld ha"e sho,n three #odes of HIV trans#ission$
i* S"A+0 -'("r,)+rs" : 5hether heterose/ual or ho#ose/ual, is the #a7or
route of trans#ission$ HIV an be trans#itted through any indi"idual at
of un%roteted se/ual interourse that is any %enetrati"e se/ual at in
,hih a ondo# is not used ,here one %artner is infeted ,ith HIV$ The
ris. of beo#ing infeted through an at of un%roteted se/ual
16
interourse de%ends on four #ain fators :
Th" 0-B"0-h))* (h( (h" s"A 6r('"r -s -'3",("* : The %robability that
a %erson has beo#e infeted ,ith HIV is in general %ro%ortionate to the
nu#ber &fre+ueny* of un%roteted se/ ats and the nu#ber of high ris.
%artners ,ith ,ho# the %erson has had se/ual ontat in reent years$
Th" (96" )3 s"A ,( : 2ll un%roteted ats of se/ual %enetration &anal,
"aginal, oral* arry a ris. of HIV trans#ission beause they bring se/ual
seretions diretly into ontat ,ith e/%osed #uous #e#brane$ In7ury
to the #uous #e#brane of the retu#, the "agina or the #outh #ay
hel% the "irus to enter into the blood strea#$ 6ee%ti"e %artners are
thus at a greater ris. than Inserti"e %artners in ats of interourse$
Ho,e"er, HIV an be trans#itted e"en through unbro.en #uous
#e#brane$
Th" 1)+'( )3 !-r+s 6r"s"'( -' (h" /0))* )r s"A+0 s",r"(-)'s
(s"1"', !2-'0 )r ,"r!-,0 s",r"(-)'s) )3 (h" -'3",("* 6r('"r :
Indi"iduals ,ith HIV infetion beo#e #ore infetious as they %rogress to
HIV related diseases and 2ID'$ There is also an early %eriod of high
infetiousness around the ti#e of seroon"ersion$
Th" 6r"s"'," )3 )(h"r s"A+009 (r's1-(("* *-s"s"s '*&)r
2"'-(0 0"s-)'s -' "-(h"r 6r('"r : It is i#%ortant to be a,are that HIV
an be trans#itted se/ually e"en ,hen neither %artner has any of the
other se/ually trans#itted diseases$ Ho,e"er, there is strong e"idene
that #en and ,o#en ,ith genital uler disease or urethral disharge are
at inreased ris. of a+uiring and trans#itting HIV$
ii* HIV infeted blood, blood %roduts, trans%lanted organs or tissues and
the use of i#%ro%erly steriliDed needles and syringes that ha"e been in
ontat ,ith infeted blood an trans#it HIV$
iii* HIV -'3",("* 8)1' ,' (r's1-( HIV () h"r 3)"(+s )r -'3'(
/"3)r", *+r-'2, )r 3("r /-r(h : 2 %regnant ,o#en ,ith HIV infetion
has an a%%ro/i#ately @>F hane of %assing the "irus to her foetus or
ne,born baby$ There is e"idene that infetion an our as early as the
first 0=A0G ,ee.s of gestation$ G>F of %erinatal infetions are in utero or
during the birth %roess$ It is esti#ated that G>F of %erinatal infetions
our through breast feeding$
$.G N(+r0 h-s()r9 )3 HIV&AIDS : )eo%le infeted ,ith HIV are both
infeted and infetious for life, e"en ,hen they loo. and feel healthy, they
an trans#it the "irus to others$ The signs and sy#%to#s of infetion
,ith HIV are "aried and o#%le/$ -our stages of HIV infetion an be
desribed
i* %r-1r9 -'3",(-)' : Infetion ,ith HIV results in ra%id %roliferation of
the "irus in blood and ly#%h nodes$ The infeted %erson #ay e/%eriene
a seroon"ersion illness, ,hih usually resol"es ,ithin ,ee.s$ The CDE
ell ount delines ra%idly before "irus is ontrolled by the i##une
syste#, ,hereu%on the ount returns to near nor#al$
ii* Er09 -11+'" *"3-,-"',9 (CD> ,"00 ,)+'(H?II&ML) : During this
%hase the i##une syste# has ontrolled the "irus, ,hih is largely
restrited to ly#%hoid tissue$ In this %hase, da#age inflited by the
"irus is li#ited to the regenrati"e a%aity of the i##une syste# and
%eo%le ,ith HIV are usually ,ithout sy#%to#s$
17
iii* I'("r1"*-(" -11+'" *"3-,-"',9 (CD> ,"00 ,)+'( .II;?II&ML) :
Viral re%liation is "ery high and CDE ell turno"er is ra%id$ 'ubtle signs
and sy#%to#s indiating o#%ro#ise of i##une systen begin to a%%ear$
i"* A*!'," -11+'" *"3-,-"',9 (CD> ,"00 ,)+'( J.II&ML) : The "irus
,hih %roliferates throughout the body o"ero#es the i##une syste#$
Ma7or o%%ortunisti infetions and #alignanies beo#e inreasingly
o##on and re+uire inreasing #edial inter"ention$
Fi#) %)7 de5ict! te nat6ral i!tor+ of HI08AIDS)
18
Fi#) %)7 Nat6ral Hi!tor+ of HI0 Infection in ad6lt!
E91OSU*E TO HI0
INFECTION
Serocon9ersion illness Asy84to8atic 4eriod S57tle Sy84to8s and
Signs o6
I885node6iciency
5ll 2lo<n AIDS
Sy84to8s and Signs
o6 o44ort5nistic
In6ections
ANTIBOD: TIT*E AND
INFECTI0IT:
Not Detecta7le
@.indo< PeriodA
+ig:ly In6ectio5s
Detecta7le anti7odies
In6ectio5s
Detecta7le anti7odies
In6ectio5s
Anti7odies 8ay or 8ay
not 7e detecta7le
+ig:ly In6ectio5s
TIME 1E*IOD 1# .ee=s 33$ years #33 years 13# years
STAGE OF HI0 DISEASE Pri8ary In6ection
@CD" cell co5nt near
nor8alA
'arly
I885node6iciency
@CD" cell co5nt O$00
cells; lA
Inter8ediate
i885node6iciency
@CD" cell co5nt P$00
O#00 cells ;l
Ter8inal illness
@CD" cell co5nt P#00
cell ;lA
19
") 0I*OLOG:
")% Introd6ction; It is 9ery i84ortant to 5nderstand t:e ca5sati9e agent o6 AIDS
@AcB5ired I885node6iciency Syndro8eA1 t:e agent <:ic: :as ca5sed a
4ande8ic and a disease <:ic: :as enor8o5s social1 econo8ic and
7e:a9io5ral i84act on indi9id5als1 6a8ilies1 co885nities and t:e <:ole <orld.
AIDS :as s:attered t:e glo7al econo8y <it: no s5ccess65l treat8ent and
9accine in sig:t.
")" Hi!tor+ and ori#in ; AIDS <as o66icially recognised 6or t:e 6irst ti8e in I5ne1
1DE1 at t:e Centers 6or Disease Control1 C.S.A. in 4re9io5sly :ealt:y
:o8ose!5al 8en dying <it: Pne58ocystis carinii 4ne58onia and candidiasis.
Since t:en AIDS :as 7een re4orted 6ro8 all t:e continents. T:e 9ir5s ca5sing
AIDS <as inde4endently identi6ied 7y a tea8 o6 renc: scientists led 7y Dr.
,5c )ontagnier o6 Paste5r Instit5te and A8erican scientists lead 7y Dr.
(o7ert C. %allo o6 National Cancer Instit5te in 1DE331DE". T:e 9ir5s :as 7een
called 7y di66erent na8es ,A& i.e. ,y84:adeno4at:y Associated &ir5s 7y t:e
renc: and +T,& III i.e. +58an T ,y84:ocytotro4ic &ir5s ty4e III 7y t:e
A8ericans. T:e International Co88ittee on No8enclat5re o6 &ir5ses na8ed
it t:e > +58an I885node6iciency &ir5s? @+I&A and to date t<o ty4es1 +I&31
and +I&3# are identi6ied. Pro7a7le origin o6 +I&s is de4icted in ta7le #.1.
")< Cla!!ification ; +58an i885node6iciency 9ir5ses @+I&A 7elong to t:e class
(etro9ir5ses and 6a8ily ,enti9irinae. T<o ty4es are recognised +I&31 and
+I&3#. 2ot: di66er in geogra4:ical distri75tion1 7iological and 8olec5lar
c:aracteristics and e!tent o6 trans8issi7ility. T:ese 9ir5ses store t:eir
genetic in6or8ation as ri7on5cleic acid @(NAA. (NA 85st 7e con9erted to
DNA 7y a s4ecial enJy8e re9erse transcri4tase. +I&31 :as 3 gro54s1 +I&31
8ajor gro54 @+I&13)A1 o5tlier @+I&13OA and +I&13N gro54. T:e strains o6
+I&31 isolated 6ro8 4eo4le in C.S.A. and '5ro4e are genetically di9erse 6ro8
strains isolated in A6rica and Asia. +I&31 8ajor gro54 can 7e 65rt:er
classi6ied into s57ty4es or clades designated A t:ro5g: H. S5c: s57ty4es
:a9e en9elo4e gene seB5ences t:at 9ary 7y #0Q or 8ore 7et<een s57ty4es.
T:e s57ty4es di66er in geogra4:ical distri75tion1 7iological c:aracteristics and
8ajor 8ode o6 trans8ission etc. +I&31 s57ty4es O and N are 8ore distant to
all ot:er +I&31 s57ty4es 75t less so co84ared to +I&3#. So t:ese are
classi6ied 5nder +I&31 only and :a9e li8ited distri75tion in .est A6rica. +I&3
# :as also 7een re4orted 6ro8 ot:er co5ntries and t:is also co84rises o6
:eterogeno5s gro54 o6 9ir5ses. Ta7le #.# s:o<s t:e distri75tion o6 co88on
s57ty4es.
Ta=le $")" HI0$% Ma>or S6=t+5e!)
Ma>or e5idemic 5attern
+eterose!5al :osts
Geo#ra5ic *e#ion
S57 3 Sa:aran A6rica
So5t: 'ast Asia
India
HI0$% S6=t+5e
A 2 C D '
R 3 R R R
3 3 3 3 R
R R R 3 3
Intra9eno5s dr5g 5sers
and
:o8ose!5al :osts
Nort: A8erica
.estern '5ro4e
So5t: 'ast Asia
India
3 R 3 3 3
3 R 3 3 3
3 R 3 3 3
3 R R 3 3
20
Ta=le $ ")%) 1ro=a=le Ori#in of HI0
SI& C:i84anJee SI& )anga7ey Nat5ral :ost A6rican
Nat5ral :ost A6rican a4e 8on=ey. No disease1 :ig: trans8ission
+I&31 Progenitor
Asian 8on=ey +58an
+I&13A +I&132 +I&13C +I&13D +I&13' +I&13 +I&130 +I&13N So8e disease L So8e disease
@SI& 8acA So8e trans8ission
@+I&3# A
21
")( Str6ct6re ; +I& is 1#0 n8 icosa:edral1 en9elo4ed1 (NA 9ir5s. +I& co84rises o6
an o5ter en9elo4e consisting o6 a li4id 7ilayer <it: 5ni6or8ly arranged 7#
s4i=es or =no7s o6 g4 1#0 and g4 "1. %lyco4rotein @g4A 1#0 4rotr5des o5t on
t:e s5r6ace o6 t:e 9ir5s and g4 "1 is e87edded in t:e li4id 8atri!. Inside is
t:e 4rotein core s5rro5nding t<o co4ies o6 (NA. Core also contains 9iral
enJy8es re9erse transcri4tase1 integrase and 4rotease1 all essential 6or 9iral
re4lication and 8at5ration. Proteins 47 and 4D are 7o5nd to t:e (NA and are
7elie9ed to 7e in9ol9ed in reg5lation o6 gene e!4ression @ig3#.1A.
")' Genetic !tr6ct6re ; T:e genetic str5ct5re o6 9ir5s contains 7ot: :ig:ly
conser9ed and :ig:ly 9aria7le regions. T:e :ig: 9aria7ility o6 t:e 9ir5s
acco5nts 6or dr5g resistance and e9asion 6ro8 i885ne res4onse. T:is also
4oses 4ro7le8s 6or de9elo48ent o6 a s5ccess65l 9accine. In an in6ected
indi9id5al1 B5asis4ecies o6 a 4artic5lar 9iral s57ty4e 8ay 7e 6o5nd on acco5nt
o6 constant 9aria7ility.
+I& :as str5ct5ral and reg5latory genes coding 6or str5ct5ral and
reg5latory 4rod5cts1 res4ecti9ely. Str5ct5ral genes direct t:e synt:esis o6
4:ysical co84onents o6 t:e 9ir5s and are also res4onsi7le 6or 9iral siJe1
s:a4e1 str5ct5ral integrity and its co84art8entaliJation in :ost cell. T:e
reg5latory genes direct synt:esis o6 4roteins t:at e66ect t:e synt:esis o6 9iral
co84onents and 9iral re4lication @ig3#.#A. Str5ct5ral genes are gag 4ol1 en9
and ot:ers s:o<n in t:e 6ig5re are t:e reg5latory genes. +I& also :as so8e
accessory genes.

")?) *e5lication ; %lyco4rotein 1#0 o6 +I& 7inds to a rece4tor;rece4tors on +I&
4er8issi9e :ost cell. Predo8inant rece4tor is t:e CD" 8olec5le 4resent on T
ly84:ocytes and 8acro4:ages1 t:o5g: ot:ers s5c: as galactosyl cera8ide
@gal CA :a9e also 7een 4ro4osed. (ece4tors are 8olec5les @4roteins and or
glyco4rotinsA 4resent on t:e s5r6ace o6 :ost cells <:ic: 6acilitate t:e
attac:8ent and entry o6 9ir5ses in to t:e cell. 'ntry o6 9ir5s into t:e :ost cell
reB5ires certain cell5lar corece4tors;6actors e.g. CC(3$1 CSC(3"1 CC(3# and
CC(3 etc designated collecti9ely as cell in6ecti9ity 6actor @CIA. CI 8ay 7e a
corece4tor or enJy8e :el4ing in 9ir5s interaction <it: :ost cell. )ost
con9incing candidate is t:e c:e8o=ine rece4tor related 4rotein1 65sin @CSC(3
"A. Once t:e g4"1 o6 t:e 9ir5s 65ses <it: t:e :ost cell 8e87rane t:e ca4sid
is 5ncoated and a ri7on5cleo4rotein co84le! ca4a7le o6 re9erse transcri4tion
is 6or8ed. D5ring t:e 4rocess o6 re9erse transcri4tion cDNA is 6or8ed 5nder
t:e e66ect o6 9iral enJy8e1 t:e re9erse transcri4tase. (e9erse transcri4tion is
ine66icient in B5iescent cells s5ggesting t:e in9ol9e8ent o6 :ost co84onents
in t:e 4rocess. T:e n5cleo4rotein co84le! 6or8ed a6ter transcri4tion
co84rises o6 linear do57le stranded DNA1 t:e gag 8atri! @)AA 4rotein1 t:e
accessory 94r 4rotein and t:e 9iral integrase @INA. T:is is called
4reintegration co84le! and is trans4orted into t:e :ost cell n5cle5s. IN
8ediates a co84le! series o6 enJy8atic ste4s and integration occ5rs at
cell5lar loci <it: o4en c:ro8atin str5ct5re. Integration 4ro7a7ly is an
essential ste4 6or 9iral re4lication. T:e integrated 9ir5s is called 4ro9ir5s.
T:e 9ir5s 8ay not 7e e!4ressed in 8any cells and is considered latent. &ir5s
e!4ression can 7e sti85lated 7y 8any 9iral1 cell5lar and e!ogeno5s 6actors.
Ot:er1 co3e!istent 9iral in6ections e.g. C)&1 :er4es 9ir5s etc. can 8a=e t:e
non 4er8issi9e cells 4er8issi9e. )at5ration o6 9ir5s also ta=es 4lace a6ter
9ir5s asse87ly and 75dding.
")?)% HI0 *e5lication $ !6mmar+
3 g4 1#0 7inds to :ost cell rece4tors
3 (e9erse transcri4tion
3 Pro9iral DNA synt:esis
3 Integration <it: :ost cell DNA
3 &iral 4roteins synt:esis
20
3 &ir5s asse87ly and 75dding
3 )at5ration o6 core34roteins
")7 Hetero#eneit+8#enetic diver!it+ ; St5dies cond5cted on di66erent isolates o6 +I&
re9ealed t:at t:e +I& is :ig:ly :eterogeneo5s in a 9ariety o6 7iological1
serological and 8olec5lar 6eat5res . T:ese incl5de :
3 ,e9el o6 9ir5s 4rod5ction
3 (e4lication =inetics in 9i9o and in 9itro
3 Cell5lar tro4is8 @ty4es o6 cells in6ectedA
3 Cyto4at:icity
3 Syncyti58 6or8ing a7ility
3 ,atency and ind5ci7ility
3 Sensiti9ity to ne5traliJing;en:ancing anti7odies
3 %enetic str5ct5re.
T:ere are t<o 8ain ty4es +I&31 and +I&3#. +I&3# <:ic: <as isolated
in Port5gal 6ro8 4atients <it: AIDS <:o :ad co8e 6ro8 .est A6rica s:o<ed
seB5ence di66erence o6 8ore t:an $$Q 6ro8 earlier isolated strains o6 +I&1 so
<as designated a di66erent ty4e stat5s. +I&3# seB5ences are closer to SI&
t:an +I&31. +I&31 and to a lesser e!tent +I&3# are 5ndergoing 85tation
d5ring eac: re4licati9e cycle. K5asis4ecies are 7eing t:ro<n constantly into
circ5lation o6 t:e in6ected indi9id5al. +I&31 strains 6ro8 di66erent
geogra4:ical areas are di66erent genetically @O#0Q seB5ence di66erencesA.
T:ree gro54s 8ajor1 o5tlier @OA and N :a9e 7een identi6ied 6or +I&31 ty4e.
)ajor gro54 :as 7een classi6ied into di66erent s57ty4es;clades A t:ro5g: H on
t:e 7asis o6 genetic1 geogra4:ical1 8olec5lar and 7iological di66erences as
a7o9e. 'ac: s57ty4e again co84rises o6 genetically :ertergeneo5s strains on
acco5nt o6 :ig: rate o6 9aria7ility o6 t:e 9ir5s e9en in t:e sa8e :ost.
+eterogeneity o6 9ir5s :as i84lications 6or de9elo48ent o6 a s5ccess65l
9accine and a t:era4e5tic agent.
")@ H6man cell! 8 cell line! and ti!!6e! !6!ce5ti=le to HI0 ; +I& 4ractically 85lti4lies in
all cells 75t t:e e!tent o6 re4lication 9aries in di66erent cells :
Haemato5oietic !+!tem ; T ly84:ocytes1 2 ly84:ocytes1 )acro4:ages1 NH cells1
)ega=aryocytes1 Dendritic cells1 Pro8yelocytes1 Ste8 cells1 T:y8ic
e4it:eli581 ollic5lar dendritic cells.
Brain ; Ca4illary endot:elial cells1 Astrocytes1 )acro4:ages @8icrogliaA1
Oligodendrocytes1 C:oroid 4le!5s1 %anglia1 Ne5ro7lasto8a cells1 %lio8a cell
lines and Ne5rons @LA.
S3in : i7ro7lasts and ,anger:ans cells @LA.
Bo.el : Col58nar and go7let cells1 'nteroc:ro8a66in cells and Colon
carcino8a cells.
Oter! : )yocardi581 (enal t575lar cells1 Syno9ial 8e87rane1 +e4atic
sin5soid e4it:eli581 +e4atic carcino8a cells1 H5466er cells1 P5l8onary
6i7ro7lasts1 oetal adrenal cells1 Adrenal carcino8a cells1 (etina1 Cer9i!
e4it:eli58 @LA1 Prostate1 Testes1 Osteosarco8a cells1 (:a7do8yosarco8a
cells1 oetal c:orionic 9illi1 Placental tro4:o7last cells.
")A Mecani!m of cell deat
3 Increase in cell 4er8ea7ility d5e to 75dding o6 9ir5s. &ir5s 45nc:es :oles and
=ills t:e cell.
3 Increase in cell 4er8ea7ility d5e to to!ic e66ects o6 9ir5s re4lication.
3 Syncytia 6or8ation 3 in9ol9ing 5nin6ected cells.
3 A4o4totic cell deat: o6 acti9ated T cells.
3 A5toi885ne 4:eno8enon in9ol9ing CD" 8olec5le.
3 ADCC i.e. anti7ody de4endent cell cytoto!icity.
21
")%& S6!ce5ti=ilit+ of HI0 ; ort5nately +I& is a 9ery 6ragile 9ir5s. It is s5sce4ti7le
to :eat1 a te84erat5re o6 $6
0
C 6or 30 8in5tes or 7oiling 6or a 6e< seconds
=ills t:e 9ir5s. )ost o6 t:e c:e8ical ger8icides 5sed in :os4ital;la7oratories
and :ealt: care settings =ill +I& at 85c: lo<er concentrations. T:5s 0.$Q to
1Q sodi58 :y4oc:lorite1 70Q et:anol1 #Q gl5taralde:yde1 acetone1 et:er1
7eta 4ro4iolactone @1:"00 dil5tionA and sodi58 :ydro!ide @"0 8 )ol;litreA
inacti9ate t:e 9ir5s.
")%&)% SteriliBation and di!infection $ !6mmar+ of metod! .ic 3ill all microor#ani!m!
incl6din# HI0 2detail! in ca5ter (4
SteriliBation
3 A5tocla9ing at 1#1
0
C1 1$ l7s 4ress5re 6or #0 8in5tes.
3 Dry :eat 170
0
C 6or 1 :r.
3 2oiling 6or #0330 8in5tes.
Cemical di!infection
3 Sodi58 :y4oc:lorite : $g8;litre. @0.$ to 1Q ordinarily1 $310Q 6or :ig:
organic
8atter content e.g. discarding tiss5es etc.A
3 Calci58 :y4oc:lorite : 1." g8;litre.
3 C:lora8ine : #0g8;litre
@A9aila7le c:lorine 0.1QA
3 't:anol : 70Q
3 or8alin : 33"Q
3 %l5taralde:yde : #Q 6or 30 8in5tes
3 Poly9idone iodine @P&IA
")%% Tran!mi!!ion ; (is= 6actors 6or +I& in6ection incl5de 85lti4le :o8ose!5al or
:eterose!5al 4artnersF conta8inated 7lood trans65sionF injections <it:
conta8inated needles and syringes and in6ected 8ot:er to 6oet5s;in6ant1
@7e6ore1 d5ring or s:ortly a6ter 7irt:A. T:e e66iciency o6 trans8ission o6 +I& is
deter8ined 7y t:e a8o5nt o6 9ir5s in a 7ody 6l5id and t:e e!tent o6 contact.
+ig: concentrations o6 6ree in6ectio5s 9ir5s and 9ir5s in6ected cells :a9e 7een
re4orted in 7lood1 genital 6l5ids and cere7ros4inal 6l5id. 2reast 8il= and
sali9a yield 9arying n587ers1 <:ereas1 ot:er 7ody 6l5ids :a9e a lo< 9iral
content. +ig: le9els o6 9ir5s are al<ays associated <it: sy84to8s and
ad9anced disease.
Sali9a in ad5lts contains so8e nons4eci6ic in:i7itory s57stances li=e
6i7ronectins and glyco4roteins <:ic: co5ld 4re9ent cell to cell trans6er o6
9ir5s. T:5s1 sali9a is not a li=ely 9e:icle o6 trans8ission. Crine1 s<eat1 8il=1
7ronc:o3al9eolar la9age 6l5id1 a8niotic 6l5id1 syno9ial 6l5id1 6aeces and tears
:a9e 7een re4orted to yield Jero or a 6e< +I& 4articles. +ence1 t:ese 9e:icles
also do not a44ear to 7e i84ortant in 9ir5s trans8ission.
2reast 8il= at t:e ti8e o6 4ri8ary in6ection in a 6eeding 8ot:er :as a
:ig: content o6 9ir5s and 8ay trans8it t:e in6ection to t:e 7a7y.
Cere7ros4inal 6l5id @CSA1 on t:e ot:er :and1 also :as a :ig: content o6 9ir5s
4artic5larly in indi9id5als <it: ne5rological disease1 75t1 CS is not a nat5ral
so5rce o6 9ir5s trans8ission.
Ta=le; ")<) Efficienc+ of different ro6te! of HI0 tran!mi!!ion and teir contri=6tion to
total n6m=er of ca!e!)
33333333333333333333333333333333333333333333333333333333333333333333333333333333333333333333
3333333333333333
'!4os5re (o5te Percent '66iciency Percentage o6 total
@.orld o9erA @.orld o9erA @ IndiaA
22
33333333333333333333333333333333333333333333333333333333333333333333333333333333333333333333
3333333333333333
2lood trans65sion D03D$ $ 7.0$
Perinatal #03"0 10
Se!5al interco5rse 0.1 to 1 7$ 7".1$
@:eterose!5alA
&aginal @60A
0.$E @:o8ose!5alA
Anal @1$A
Injecting dr5gs 5se 0.$31.0 10
7.3
Needle stic= e!4os5re P 0.$ 0.1 0
Ot:ers 10.D#
33333333333333333333333333333333333333333333333333333333333333333333333333333333333333333333
3333333333333333@Percentage o6 total in India 6ro8 NACO doc58ent entitled
>Co5ntry Scenario1? 1DDEA
T:e 8ost e66icient 9e:icle o6 +I& trans8ission is 7lood @Ta7le3#.3A.
+o<e9er1 t:e ris= o6 in6ection 9ia 7lood trans65sion is no< e!tre8ely lo< d5e
to strict +I& screening o6 donated 7lood. T:e 8ost co88on ro5te o6
trans8ission is 5n4rotected1 4enetrati9e se!5al contact. Di66erent 6or8s o6
se!5al 4ractices carry a 9aria7le ris= gradient o6 acB5iring +I&. Cell
associated rat:er t:an 6ree 9ir5s is res4onsi7le 6or disease trans8ission. Anal
interco5rse carries a :ig: ris= o6 trans8ission 7eca5se o6 7o<el 85cosa
<:ic: acts as a 4ortal o6 entry 6or 9ir5s1 and also 7eca5se o6 a greater c:ance
o6 inj5ry to t:e 85cosa. (is= to inserti9e 4artner is t:ro5g: in6ection o6
ly84:ocytes and 8acro4:ages in t:e 6ores=in or along t:e 5ret:ral canal. In
6e8ales +I& trans8ission occ5rs <:en in6ected cells in t:e se8en gain entry
into t:e 6e8ale genital tract1 and in6ect t:e resident ly84:ocytes1
8acro4:ages and 4ro7a7ly t:e 5terine e4it:elial cells.
T:e trans8ission 6ro8 in6ected 8ot:er to c:ild a44ears to occ5r in #03
"0Q c:ildren 7orn to +I& 4ositi9e 8ot:ers. T:e so5rce o6 9ir5s in t:e
ne<7orn is contro9ersial. +I& in6ection can occ5r 9ia a8niotic 6l5id1 genital
secretions1 8aternal 7lood and t:ro5g: t:e 7reast 8il=. Trans8ission to t:e
7a7y can occ5r in35tero1 and d5ring or a6ter deli9ery.
Trans8ission o6 +I& in6ection to +ealt: Care .or=ers @+C.A is
e!tre8ely 5nco88on. Pooled data 6ro8 #0 4ros4ecti9e st5dies s5ggests t:at
ris= associated <it: needle inj5ry 6ro8 +I& in6ected 7lood is a44ro!i8ately
0.$ 4ercent. 5rt:er1 t:e ris= associated <it: 85coc5taneo5s contact is too
lo< to 7e relia7ly esti8ated. T:e ris= 6ro8 85cosal or non3intact s=in is also
8ini8al.
ig. #.3 de4icts t:e di66erent ro5tes o6 trans8ission o6 +I&.
So 6ar1 t:ere :as 7een no re4ort o6 +I& trans8ission t:ro5g: a cas5al
social contact1 7y t:e enteric or res4iratory ro5te1 and t:ro5g: an insect
e.g.8osB5ito 7ite. Pros4ecti9e st5dies o66er a concl5si9e e9idence t:at 6a8ily
8e87ers and close :o5se:old contacts o6 +I& in6ected indi9id5als are not at
ris= o6 acB5iring +I& in6ection t:ro5g: cas5al :58an contact @s:a=ing :ands1
=issing and 7y s:aring o6 5tensils1 toilet1 linen1 7ed etc.A or 7y 4ro9iding
ro5tine n5rsing care.
23
Fi#) ")< Tran!mi!!ion c+cle of HI0
+I& @&ir5sA
+58an 7eing is
t:e reser9oir
or :ost
+I& Trans8ission t:ro5g:

2lood;7lood 4rod5cts
&aginal 6l5id
Se8inal 6l5id
2reast 8il=
Ot:er 7ody 6l5ids
In6ects ot:er 5nin6ected 4ersons t:ro5g:

Cn4rotected se!5al contact

Trans65sion o6 conta8inated 7lood and 7lood
4rod5cts
Cse o6 5nsterilised syringes1 needles and ot:er
instr58ents
)ot:er to C:ild
@in35etro1 d5ring deli9ery and 7reast 6eedingsA
+I& In6ected 4ersons increase
AIDS cases
Deat:s
Trans8ission o6 +I& t:ro5g: cas5al contact1 s:aring 5tensils1 la9atories and t:ro5g:
insect 7ites :as not 7een doc58ented so 6ar
24
<) IMMUNO1ATHOLOG:
<)% Introd6ction ; To 5nderstand t:e e66ects o6 +I& in6ection a re8inder o6
7ody de6ence is a 85st. T ,y84:ocytes and 2 ,y84:ocytes toget:er
de6end t:e 7ody against all =inds o6 assa5lts t:e 7ody is e!4osed to1
at all ti8es. 2ot: T and 2 cells 8igrate 6ro8 7one 8arro<1 75t T cells
8at5re in t:y85s1 <:ere t:ey de9elo4 s4ecial 65nctions. 2ot: =inds
o6 ly84:ocytes <:en acti9ated 7y antigen1 85lti4ly and c:ange. 2
cells de9elo4 into 4las8a cells1 4rod5ce s4eci6ic anti7odies <:ic:
tra4 and =ill 8icroorganis8s @7acteria 8ostlyA. T:is is >:58oral
i885nity?. T ly84:ocytes are res4onsi7le 6or cell38ediated
i885nity1 9ery i84ortant de6ence against 65ngi1 4rotoJoa1
8yco7acteria and 9ir5ses. T ,y84:ocytes incl5de :el4er T cells
@CD"A and cytoto!ic;=iller T cells @CDEA. T:ey release sol57le
6actors ly84:o=ines @cyto=inesA1 can reg5late :58oral s544ressor
cells and can 7eco8e 8e8ory cells. So CD" cell 4o45lation is central
in de6ending t:e 7ody. It is t:e =ey cell and t:is is t:e cell +I&
in6ects and destroys 4rogressi9ely. In6ection <it: +I& irres4ecti9e o6
ty4e @+I&31 or +I&3#A s57ty4e and ro5te o6 in6ection leads to
4rotracted disease and de4letion o6 CD" cells in 8ost cases res5lting
in AIDS. T:e rate o6 4rogression o6 disease de4ends 54on 9iral
c:aracteristics on t:e one :and and :ost 6actors on t:e ot:er :and
and 8ay ta=e 6ro8 1 year to 8ore t:an 1$ years.
<)" 0iral Entr+; Cell 6ree or cell associated +I& enters t:e 7ody d5ring :ig:
ris= 4ractices t:ro5g: any ro5te 9ia 7lood1 se8en and 9aginal
secretions 6ro8 an in6ected 4erson. +I& in6ection is 6acilitated 7y
4resence o6 5lcerati9e and to a lesser e!tent non5lcerati9e se!5ally
trans8itted in6ections. +I& i88ediately targets on to cells
dis4laying co84le8entary rece4tors @CD"1 CC(3$ and CSC(3";65sinA
<:ic: 8ay 7e CD" cells1 resident 8acro4:ages or ,anger:ans cells
de4ending 54on t:e site o6 e!4os5re. &ir5s g41#0

6its on t:e rece4tor
li=e a loc= and =ey syste8. &iral re4lication starts i88ediately a6ter
entry into t:e cell and disse8ination occ5rs t:ro5g: circ5latory and
ly84:oid syste8s.
<)< 1rimar+ HI0 Infection ; D5ring t:is stage +I& and +I& in6ected cells
reac: t:e ly84: nodes and t:e ot:er ly84:oid tiss5es1 <:ere acti9e
i885ne res4onse to 9iral antigens occ5rs and at t:e sa8e ti8e
intense re4lication o6 9ir5s occ5rs in acti9ated T ly84:ocytes. T:is is
a 4arado! 7eca5se ly84:ocytes are acti9ated on acco5nt o6 in6ection
and +I& re4licates 7etter in t:ese acti9ated cells. T:e 4ea= in
n587er o6 9ir5s e!4ressing cells and s4read o6 9ir5s t:ro5g:o5t t:e
ly84:oid tiss5e 4recedes t:e increase in 4las8a 9irae8ia i.e. t:e
9ir5s in t:e 7lood. T:e 9ir5s s4ills o9er 6ro8 ly84: nodes. T:ese
4:eno8ena occ5r d5ring t:e 6irst #33 <ee=s a6ter in6ection and t:ere
is intense 9ir5s s4reading d5ring t:is 4eriod so t:is is called t:e !ta#e
of vir6! di!!emination. Clinically it coincides <it: >6l5 li=e illness? also
=no<n as ac5te +I& disease. T:ere is signi6icant 6all in CD" cells and
9iral le9els 8ay 7e as :ig: as 10
6
310
7
9iral co4ies;8l. T:e ne!t !ta#e
i! tat of do.n re#6lation of viraemia. T:is coincides <it: ro75st1
intense i885ne res4onse 7y t:e :ost. 2ot: e66ecti9e cell 8ediated
i885ne res4onse carried o5t 7y +I& s4eci6ic cytoto!ic T ly84:ocytes
@CT,A and :58oral res4onse carried o5t 7y
25
Fi#) <)% 0iral 1ato#ene!i! and imm6ne re!5on!e
Infection
0iral di!!emination to l+m5oid
ti!!6e
0iral di!!emination
via =lood
0iral di!!emination to
CNS ti!!6e
Tra55in# of vir6! in follic6lar
dendritic cell net.or3
Imm6ne re!5on!e to
HI0
SeC6e!tration of vir6! in l+m5
node!

0ir6! activel+ re5licatin# in l+m5oid
ti!!6e, CD( cell! and follic6lar
dendritic cell!
Di!r65tion of follic6lar dendritic cell
net.or3, .it increa!in# De!ca5e of
vir6!E
Increa!ed demand! on
imm6ne re!5on!e
De!tr6ction of te
imm6ne !+!tem d6e to
con!tant viral
re5lication
Increa!ed ri!3 of
infection! and t6mo6r!
A55earance of clinical
!+m5tom!
AIDS
26
co84le8ent 6i!ing and ne5traliJing +I& s4eci6ic anti7odies co8es
into 4lay. T:e 4eriod 6ro8 t:e entry o6 +I& in t:e :ost and t:e
a44earance o6 detecta7le le9els o6 +I& s4eci6ic anti7odies is called
F.indo. 5eriodG. D5ring t:is 4eriod indi9id5al is in6ected1 is in6ectio5s
to ot:ers 75t is seronegati9e i.e. +I& tests 6or detecting anti7odies
are negati9e. .indo< 4eriod ranges 6ro8 3 <ee=s to 3 8ont:s on
a9erage1 can 7e longer so8eti8es. 2ot: +I& s4eci6ic anti7odies and
CT, =ill t:e 9ir5s in6ected cells. As a res5lt t:e 9irae8ia dro4s and
CD" cells 7o5nce 7ac= to le9els slig:tly lo<er t:an t:e 4re9io5s
nor8al le9el. )ost o6 t:e 9ir5s tra44ing and =illing occ5rs in
ollic5lar Dendritic Cells @DCA o6 t:e ly84: nodes and ly84:oid
tiss5e. T:is 8ay 7e one reason o6 generalised ly84:adeno4at:y seen
in +I& disease. A44earance o6 ne5traliJing +I& s4eci6ic anti7odies
:eralds t:e transition 6ro8 ac5te to cronic !ta#e of HI0 di!ea!e.
Alt:o5g: t:e i885ne res4onse s5cceeds in do<nreg5lating t:e
9irae8ia1 +I& is ne9er co84letely eli8inated and 4rogression to
c:ronic 4:ase o6 +I& disease occ5rs in 8ost cases. .:at deter8ines
t:e 4rogression o6 t:e +I& disease is t:e B5ality o6 T cell res4onse.
T:is 8ay 7e gentically deter8ined.
ig. 3.1 de4icts t:e co5rse o6 e9ents 6ollo<ing 9iral entry in t:e 7ody.
<)<)% 1rimar+ infection !6mmar+
3 Disse8ination o6 +I& to ly84:
nodes and
ot:er ly84:oid organs.
3 &irae8ia
3 .indo< 4eriod
3 Ac5te +I& disease
3 CT, res4onse
3 Anti7ody res4onse
Ac5te gland5lar 6e9er li=e syndro8e
Ac5te 8eningoence4:alo4at:y @less
co88onA
<)( Clinicall+ latent 5eriod8Cronic illne!! ; T:is stage is 8ar=ed 7y
disa44earance o6 sy84to8s o6 ac5te 9iral disease1 do<n reg5lation o6
9irae8ia1 CD" cell co5nt 7eco8es al8ost nor8al and t:e ne5traliJing
and co84le8ent @C
$
A6i!ing 9ir5s s4eci6ic anti7odies a44ear in t:e
7lood. All 9irological 4ara8eters in t:e 4eri4:eral 7lood @9iral (NA
co4ies;9iral load1 9ir5s e!4ressing 8onon5clear cells1 etc.A are 9ery
lo< . +o<e9er1 acti9e and contin5o5s 9ir5s re4lication goes on in
t:e ly84: nodes and ly84:oid organs <:ic: e!4ress 9ir5s 133 logs
:ig:er t:an t:e 4eri4:eral 7lood. As long as t:e CD" co5nts are
:ig:er t:an $00 cells;l1 t:e i885ne res4onse 8o5nted 7y t:e
ly84:oid tiss5es is e66ecti9e1 t:ere is 6ollic5lar :y4er4lasia o6
ger8inal centres indicating i885ne acti9ation o6 ly84: nodes. T:e
i84ortant 4arado! to note is t:at cell5lar acti9ation seen in ly84:
nodes is critical 6or 9iral re4lication i.e. 9ir5s re4lication is 7etter in
acti9ated CD" cells. T:ere is grad5al red5ction in n587er o6 CD"
cells and increase in 9ir5s load d5ring t:e long asy84to8atic stage.
Increase in 9ir5s load in 4eri4:eral 7lood indicates 6ail5re o6 and
4rogressi9e deterioration o6 e66ecti9e i885ne res4onse.
+58oral i885nity is intact d5ring t:e asy84to8atic stage t:at
is s4eci6ic anti7odies are 4rod5ced against di66erent 9iral 4roteins1
75t t:e anti7odies are not 4rotecti9e1 are not a7le to inter6ere <it:
27
cell to cell trans8ission and in6ecti9ity o6 9ir5s on acco5nt o6 constant
9ariation o6 9ir5s. T:is 4eriod on a9erage lasts 6or E310 years.
Progressi9e i84air8ent o6 +I& s4eci6ic and nons4eci6ic cell 8ediated
and :58oral i885ne res4onses :eralds t:e onset o6 AIDS. T:e CD"
cell co5nts range 7et<een O#00 to $00 cells;l. in 4eri4:eral 7lood.
<)' AIDS ; Ad9anced stage o6 +I& in6ection is c:aracterised 7y increase
in all 9irological 4ara8eters @9ir5s load1 4#" antigen etc.A in 7ot:
4eri4:eral 7lood and ly84: nodes. ,y84:oid tiss5e is totally
destroyed and re4laced 7y 6i7ro5s tiss5e. &ir5s tra44ing 7y <:ate9er
ly84:oid tiss5e re8ains is 8ini8al or nil. T:ere is 4ro6o5nd i885ne
s544ression and o44ort5nistic in6ections 8ay 4ro9e 6atal at t:is
stage. T:e CD" co5nt is 5s5ally less t:an #00 cells;l. and
4rogressi9ely 6alls @ig. 3.1 and 3.#A.
<)')% 1oint! to 1onder
3 In6ection <it: +I& is li6elong .
3 Se9ere i885node6iciency de9elo4s in in6ected 4ersons <it:in 1031# years
on a9erage.
3 ConseB5ent to s566icient i885ne da8age1 s5sce4ti7ility to o44ort5nistic
in6ections and
cancers increases.
3 Cancers and o44ort5nistic in6ections act as s5rrogate clinical indicators o6
AIDS.
3 S5r9i9al a6ter diagnosis o6 AIDS is s:ort1 13# years on a9erage.
<)? Mecani!m of CD(

cell de5letion and d+!f6nction ; CD" cells are t:e 8ain
targets o6 +I& and 4rogressi9e destr5ction o6 t:ese cells is
c:aracteristic o6 all stages o6 +I& disease. CD" cells ser9e as
s5rrogate 8ar=ers to 8onitor t:e 4rogression o6 +I& in6ection.
T:ese cells can 7e destroyed 7y t<o 8ec:anis8s :
iA Direct da8age 7y t:e 9ir5s.
iiA I885ne 8ec:anis8s triggered d5ring t:e co5rse o6 +I& in6ection.
+I& can =ill cells singly or a6ter giant cell and syncytia 6or8ation.
Single cell =illing occ5rs d5e to acc585lation o6 5nintegrated 9iral
DNA and in:i7ition o6 cell5lar 4rotein synt:esis. Syncyti58 6or8ation
is ind5ced 7y 9ir5lent strains o6 +I& in a 85ltiste4 8ec:anis8. CD"
cells e!4ressing 9iral antigens on t:e s5r6ace attract CD" 5nin6ected
cells and t:e 8e87ranes o6 t:ese 65se 4rod5cing giant cells and
syncytia. One s5c: +I& in6ected cell can eli8inate :5ndreds o6
5nin6ected cells 7y syncyti58 6or8ation. %lyco4rotein 1#0 and ot:er
intracell5lar ad:esion 8olec5les 7ring a7o5t t:e cell5lar ad:esion
and s57seB5ent da8age.
T:e non 9irologic 8ec:anis8s <:ic: can da8age;destroy CD" cells
incl5de a5toi885ne 8ec:anis8s1 anergy1 s54erantigens1 a4o4tosis
@4rogra88ed cell deat:A and 9ir5s s4eci6ic i885ne res4onses. A
n587er o6 :y4ot:esis and co84le! i885ne 8ec:anis8s :a9e 7een
4ost5lated 6or CD" cell de4letion in9ol9ing one or 8ore o6 a7o9e
8entioned 4at:<ays.
28
ig. 3.# de4icts t:e =inetics o6 CD" T ly84:ocytes d5ring t:e co5rse
o6 +I& in6ection 9is3a39is 9iral load.
<)7 Te co6r!e of 5ro#re!!ion HI0 infection ; T:ree do8inant 4atterns o6 +I&
disease 4rogression :a9e 7een descri7ed. T:ese are 7ased on t:e
=inetics o6 i885nologic and 9irologic e9ents descri7ed a7o9e.
iA E0Q3D0Q o6 +I& in6ected are >ty4ical 4rogressors? <it: a 8edian
s5r9i9al ti8e o6 10 years1 a44ro!i8ately.
iiA $Q to 10Q o6 +I& in6ected indi9id5als are >ra4id 4rogressors? <it: a
8edian s5r9i9al ti8e o6 33" years a44ro!i8ately.
iiiA A7o5t $Q o6 +I& in6ected indi9id5als do not e!4erience disease
4rogression 6or an e!tended 4eriod o6 ti8e and are called > long ter8
non 4rogressors? @,TNPsA.
<)7)% T+5ical 5ro#re!!or! ; T:e ty4ical co5rse o6 +I& in6ection incl5des t:ree
4:ases: 4ri8ary in6ection @serocon9ersionA1 clinical latency and
clinically a44arent disease.
Pri8ary 4:ase 8ay 7e totally ina44arent or 8ay 7e associated
<it: ac5te 6l5 li=e or 8onon5cleosis li=e syndro8e in $0Q to 70Q
indi9id5als. Occ5rs <it:in 336 <ee=s o6 in6ection and 8ay last 6or D3
1# <ee=s. T:ere is :ig: le9el o6 9ir5s in t:e 7lood. T:e co5rse o6 +I&
disease is as descri7ed a7o9e. Progression o6 +I& in6ection to AIDS
on a9erage occ5rs in E310 years a44ro!i8ately.
<)7)" *a5id 1ro#re!!or! ; In a7o5t $Q to 10Q +I& in6ected ra4id 4rogression
to AIDS occ5rs <it:in #33 years a6ter serocon9ersion. I885ne
res4onse is de6ecti9e in t:ese indi9id5als. ,e9els o6 ne5traliJing and
C
$
6i!ing +I& s4eci6ic anti7odies are lo< and CDE

cell 8ediated
s544ression o6 +I& re4lication is /i84aired0. As a res5lt 4rogression
to AIDS is ra4id in t:ese indi9id5als.
<)7)< Lon# term non$5ro#re!!or! 2LTN1!4 ; A s8all 4ercentage @$QA o6 +I&
in6ected indi9id5als do not e!4erience clinical 4rogression o6 +I& and
:a9e sta7le CD" cell co5nts o9er long 4eriod <it:o5t any t:era4y.
CD" co5nts stay at aro5nd O$00 cells;l.1 cell 8ediated and :58oral
i885ne res4onses are co84arati9ely strong in t:ese indi9id5als.
A7sol5te n587er o6 CDE cells is also 4ersistently :ig: in t:ese
4ersons. In addition t:e cytoto!ic T ly84:ocytes retain t:eir
cytoto!ic acti9ity against +I&. T:e titer o6 ne5traliJing anti7odies
against +I& is also :ig:er in t:ese indi9id5als. T:e 9ir5s s4eci6ic
4ara8eters li=e 9ir5s load1 9ir5s re4lication in 4eri4:eral 7lood and
ly84: nodes are 6o5r 6old to #036old lo<er in long ter8
non4rogressors. Also t:e 9ir5s in6ecting t:ese 4ersons 8ay 7e o6 lo<
4at:ogenicity. So8e :ost genetic 6actors 8ay also 7e res4onsi7le 6or
t:ese ,TNPs.
ig. 3.3 de4icts t:e 4rogression o6 +I& in6ection in di66erent :osts.
<)@ T+5e % and T+5e " re!5on!e! in HI0 infection and eH5o!6re ; T:e loss o6
:el4er cell 65nction in asy84to8atic indi9id5als is seen e9en 7e6ore
t:e 6all in n587er o6 CD" cells. T:is is e9ident 6ro8 in39i9o and in3
9itro st5dies. Progression o6 +I& in6ection 6ro8 asy84to8atic to
29
sy84to8atic stage and t:en AIDS in9ol9es s:i6t o6 T:1 ty4e
res4onses @cell 8ediated i885ne res4onsesA to T:# res4onses
@:58oral i885ne res4onsesA.
T:1 or Ty4e 1
Do8inent
cell5lar
res4onse
T:# or Ty4e #
Do8inant
anti7ody
res4onse
IN3* I,3"
Cyto=ines <:ic: 8od5late t:ese
res4onse
I,3# I,3$
I,31# I,310
I,31$ I,313
Cyto=ine ind5ced i885ne dysreg5lation is a central co84onent
in 4rogression to AIDS . T:is s:i6t in ty4e o6 i885ne res4onse and
conseB5ent 4rogression to AIDS 8ay 7e on acco5nt o6 t:e 6act t:at
+I& in6ects Ty4e31 cells easily co84ared to Ty4e3# cells. T:e s:i6t in
i885ne res4onse is associated <it: s:i6t in cyto=ine 4ro6ile also
@6ro8 Ty4e31 to Ty4e3#A and 4rogression to AIDS. T:is c:ange in
cyto=ine 4ro6ile res5lts in ina7ility o6 :ost 8acro4:ages to get rid o6
o44ort5nistic 4at:ogens res5lting in o44ort5nistic in6ections and
neo4las8s <:ic: 5lti8ately 4ro9e 6atal.
30
>. BIOSAFETY IN LABORATORIES
()%) Bac3#ro6nd information ; Since t:e ad9ent o6 AcB5ired I885no3
de6iciency Syndro8e @AIDSA e4ide8ic1 e!traordinary e66orts all o9er
t:e <orld :a9e 7een 8ade to 4re9ent la7oratory acB5ired in6ections
<it: +58an I885no3de6iciency &ir5s @ +I&A.T:e recent statistics
457lis:ed 7y CDC relati9e to trans8ission ris=s 6or +I& and +e4atitis
2 9ir5s @ +2&A are t:at t:e ris= 6or acB5iring +I& 6ollo<ing needle
stic= 4ric= @4er3c5taneo5s e!4os5reA 6ro8 a so5rce 4atient <it: +I&
is 9ery lo< @0.#$ 3 0.3QA. T:is is 7eca5se t:e 9ir5s generally
circ5lates at a 85c: lo<er concentration @103100 in6ectio5s doses
;8,A and 7eca5se it is not a7le to s5r9i9e as <ell in t:e en9iron8ent
o5tside t:e 7ody. In contrast1 acB5iring +2& in6ection 6ollo<ing
needle stic= 4ric= 6ro8 a +2& carrier ranges 6ro8 D 3 30Q as +2& is
6o5nd in 9ery :ig: titres @O1010001000 in6ectio5s doses ;8,A in t:e
7lood o6 8any ac5tely and c:ronically in6ected indi9id5als and can
s5r9i9e 6or long 4eriods in dried 7lood at roo8 te84erat5re. T:e
c:ance o6 acB5iring +e4atitis C 9ir5s @ +C&A is 3 3 10Q. T:e ris= o6
trans8ission o6 +I& t:ro5g: 85co3c5taneo5s e!4os5re is 0.0$Q. In
di66erent st5dies carried o5t in t:e C.S.1 t:ere <ere $# doc58ented
sero3con9ersions in t:e conte!t o6 occ54ational e!4os5re 54 to I5ne
1DD7. O5t o6 t:ese1 "$ <ere 4er3c5taneo5s e!4os5res1 $ <ere 85co3
c5taneo5s e!4os5res11 :ad 7ot: and 1 :ad 5n=no<n ty4e o6
e!4os5re. T:ere <ere "6 e!4os5res to 7lood11 to 7ody 6l5id11 <it:
5ns4eci6ied 6l5id and t:ree in9ol9ed 9iral c5lt5res in t:e la7oratory.
In all1 1D la7oratory tec:nicians :ad got occ54ationally acB5ired +I&.
T:e ris= o6 occ54ational +I& trans8ission de4ends on :
3 Pre9alence o6 in6ected indi9id5als in t:e 4o45lation
3 T:e 6reB5ency o6 e!4os5re to conta8inated 8edical instr58ent
3 T:e relati9e in6ecti9ity and concentration o6 t:e 9ir5s
3 I6 t:ere is an e!4os5re to a large B5antity o6 7lood
3 Dee4 needle stic= inj5ry
3 Inj5ry <it: :ollo< 7ore needles and
3 So5rce 4atient <:o died <it:in 60 days as a res5lt o6 AIDS

In India, there is no %ublished data a"ailable on HIV trans#ission due to
needle sti. in7ury ! #uoAutaneous e/%osure in health are setting$ The la.
of %ro%er infrastruture, lo, le"els of a,areness about safety %reautions and
a ertain degree of o#%laeny a#ongst laboratory staff are so#e of the
onerns for the laboratory #anagers$
>... I16)r('," )3 /-)s3"(9 6r,(-,"s : There is no "aine for %re"ention of
HIV! 2ID', treat#ent is e/%ensi"e and is not easily a"ailable$ Hene
%re"ention of infetion is the ornerstone of ontrol of this e%ide#i$ In the
health are settings also, %re"ention is onsidered #ost i#%ortant$ The
irrational fear of aidental infetion fro# HIV!2ID' %atients in"ariably
influenes the beha"iour of the health are ,or.ers &s%eially those ,ho
routinely handle blood and body fluids* to,ards their %atients and their
s%ei#ens$ Therefore, on the one hand, it is e/tre#ely i#%ortant that the
laboratory ,or.ers should be a,are of the ris.s in"ol"ed in their day to day
,or. and a%%ly effeti"e infetion ontrol %raties to redue the %ossibility of
trans#ission of these fatal infetions to a #ini#u#$ (n the other hand, they
should not be %re7udied and should not sho, disri#ination to,ards %atients
of HIV!2ID'$ Table E$0 sho,s the different high ris. %roedures for the health
31
are ,or.ers &H$C$5$* and the "arious #odes of a+uiring infetion$
T/0" >.$ M)*"s )3 "A6)s+r" () /0))* /)r'" 6(h)2"'s -' (h"
0/)r()r9
Lab$ )roedure H$C$5$ at 6is. 'oure!Modes of
Trans#ission
Colletion of
blood!Body fluids
Laboratory
tehniian!8ursing staff
8eedle sti. in7ury,
Bro.en s%ei#en
ontainer, Blood
onta#ination of hand
,ith s.in lesion!breah$
Transfer of s%ei#en Laboratory tehniian
and trans%ort ,or.er
Conta#inated e/terior of
the ontainer!re+uisition
sli%$
)roessing of
s%ei#en
Laboratory %ersonnel )unture of s.in or
onta#ination of
s.in!#uous #e#brane
fro#
A Conta#inated ,or.
surfae
A '%ill!s%lash of s%ei#en
A Bro.en s%ei#en
ontainer
A -aulty tehni+ues
Cleaning!,ashing 'u%%ort staff )unture!onta#ination of
s.in fro#
A Conta#inated
glass,are
A 'har%s
A Conta#inated ,or.
surfae
Dis%osal of ,aste Laboratory %ersonnel
su%%ort staff
Contat ,ith infetious
,aste s%eially shar%s
Trans%ort of s%ei#en
to distant laboratory
Trans%ort!)ostal staff Bro.en!lea.ing ontainer
()<) Bio!afet+ in a HI0 te!tin# la=orator+ 8ay 7e disc5ssed 5nder t:e
6ollo<ing :eadings :
()<)%) Preca5tions in relation to 7lood and 7ody 6l5ids 3 Univer!al -or3
1reca6tion!
()<)") '66ecti9e 5se o6 sterilisation and disin6ection
()<)<) Sa6e dis4osal o6 :os4ital <aste
>.=.$. U'-!"rs0 /0))* '* /)*9 30+-* 6r",+(-)'s 3)r 0/)r()r9 8)rB"rs
&6eo##ended by CDC 3uidelines, 0B1H* : 2lthough the ris. of trans#ission
of infetion due to blood borne %athogens in the laboratory annot be
eli#inated altogether beause hanes of aidents are there, a%%liation of
uni"ersal %reautions #ini#ise these to a large e/tent$ U'-!"rs0
6r",+(-)'s refer to %reautions onsistently used for all %atients regardless
of their blood borne infetion status$ <nder uni"ersal %reautions, blood and
body fluids of all %atients are onsidered %otentially infetious for HIV, HBV,
32
HCV and other blood borne %athogens$ 'i#ilarly, all instru#ents and other
e+ui%#ents that ha"e o#e into ontat ,ith blood are assu#ed to be
%otentially onta#inated ,ith blood borne %athogens and #ust be %ro%erly
handled, leaned and steriliDed! disinfeted or safely dis%osed of$ <ni"ersal
%reautions are intended to %re"ent aidental %arenteral, #uous #e#brane
and nonintat s.in e/%osures of health are ,or.ers to the abo"e blood borne
%athogens in %atients$ 6outine and !or #andatory testing of %atients for HIV
antibody or HBs2g is not an effeti"e strategy for ontrolling the trans#ission
of these infetions in health are settings$
>.=.$.$.B)*9 30+-*s () 8h-,h +'-!"rs0 6r",+(-)'s 6609
A Blood M the single #ost i#%ortant soure of HIV, HBV and other blood borne
%athogens in the ou%ational setting
A 2ll other body fluids ontaining "isible blood
A 'e#en 4 "aginal seretions Mnot i#%liated in ou%ational trans#ission
A Cerebros%inal fluid
A 'yno"ial fluid
A )leural fluid
A )eritonial fluid
A )eriardial fluid
A 2#nioti fluid
>.=.$...B)*9 30+-*s () 8h-,h +'-!"rs0 6r",+(-)'s *) ')( 6609
The ris. of HIV trans#ission is e/tre#ely lo, or negligible unless these
ontain "isible blood$ These inlude :
A -aees
A 8asal seretions
A '%utu#
A ',eat
A Tears
A <rine
A Vo#itus
>.=.$.=.U'-!"rs0 6r",+(-)'s -',0+*" :
i* Barrier %rotetion
ii* Hand ,ashing
iii* 'afe tehni+ues
i"* 'afe handling of shar% ite#s
"* 'afe handling of s%ei#en &blood et$*
"i* 'afe handling of s%ill of blood!body fluid
"ii* <se of dis%osable!sterile ite#s
"iii* 'afe tehni+ues inluding #ehanial %i%etting de"ie
i/* I##unisation ,ith He%atitis B "aine
-) Brr-"r 6r)(",(-)' : )roteti"e barriers redue the ris. of e/%osure of the
laboratory ,or.erCs s.in or #uous #e#brane to %otentially infeti"e
#aterials inluding blood and the body fluids$
Glove! ; Can red5ce t:e incidence o6 conta8ination o6 :ands 75t
cannot 4re9ent 4enetrating inj5ries 7y needles and ot:er s:ar4
instr58ents.
33
%lo9es !o6ld =e ;
3 .orn <:ile collecting;:andling 7lood s4eci8ens1 7lood soiled ite8s
or <:ene9er t:ere is a 4ossi7ility o6 e!4os5re to 7lood or 7ody 6l5ids
containing 7lood.
3 .orn <:ile dis4osing la7oratory <aste
3 .ell 6itting dis4osa7le 9iny 1 and 85st 7e c:anged i6 9isi7ly
conta8inated <it: 7lood;7reac:ed
3 +ea9y d5ty general 45r4ose r577er glo9es 6or <as:ing in6ected
glass<are;s:ar4s. T:ese 5tility glo9es 8ay 7e deconta8inated and
re5sed 75t s:o5ld 7e discarded i6 t:ey are 4eeling1 crac=ed or
discolo5red or i6 t:ey :a9e 45nct5re1 tears etc.
3 (e8o9ed 7e6ore :andling door =no7s1 tele4:ones1 4ens1 4er6or8ing
o66ice <or= and lea9ing t:e la7oratory.
La=orator+ #o.n! ;
3 ,a7oratory go<ns or 5ni6or8s @4re6era7ly <ra43aro5nd go<nsA
s:o5ld 7e <orn <:en in t:e la7ortory and s:o5ld 7e re8o9ed 7e6ore
lea9ing.
3 Plastic a4rons s:o5ld 7e 5sed <:ile cleaning in6ected re35sa7les and
d5ring dis4osing <astes.
Facial 5rotection ;
3 Si84le and c:ea4 de6lector 8as=s and 4rotecti9e glasses 8ay 7e
<orn i6 s4las:ing or s4raying o6 7lood;7ody 6l5ids is e!4ected.
Occl6!ive =anda#e ;
3 All s=in de6ects eg. c5ts1 scratc:es or ot:er 7rea=s 85st 7e co9ered
<it: <ater34roo6 dressing 7e6ore 4atient care.
ii4 Hand .a!in# ; T:is is an ideal sa6ety 4reca5tion and is one o6 t:e
8ost i84ortant in 4re9enting +I& trans8ission in :ealt: care
settings.
3 +ands s:o5ld 7e <as:ed t:oro5g:ly in r5nning <ater <it: soa4
<it:o5t 8issing any area @ig.".1and ".#.A.
3 .as:ing o6 :ands is 8andatory
3 i88ediately a6ter conta8ination <it: 7lood ; 7ody 6l5ids
3 a6ter re8o9ing go<ns ; coats and glo9es
3 7e6ore eating ; drin=ing and lea9ing t:e la7oratory
3 Ideally1 liB5id soa4 dis4ensors s:o5ld 7e 4ro9ided to t:e la7oratories1
<:ic: s:o5ld 7e reg5larly cleaned and 8aintained. I6 not 6easi7le1
soa4 7ars a6ter <as:ing s:o5ld 7e le6t in a dry tray to 4re9ent
34
conta8ination <it: so8e 8icroorganis8s <:ic: gro< in 8oist
conditions
3 A 8oist5rising :and crea8 s:o5ld 7e 5sed a6ter e9ery :and <as:
3 %lo9es s:o5ld not 7e regarded as s57stit5te 6or :and <as:ing
iii4 Safe tecniC6e! ;

3 Cse o6 7iological sa6ety ca7inets3 2iological sa6ety ca7inets Class #
s:o5ld 7e 5sed 6or :andling 8aterials containing :ig:er
concentration o6 in6ectio5s agent t:an 5s5ally 4resent in clinical
s4eci8ens eg. <:ile carrying o5t +I& c5lt5re.
3 All 4roced5res and 8ani45lations o6 4otentially in6ectio5s 8aterial
s:o5ld 7e 4er6or8ed care65lly to 8ini8ise t:e 6or8ation o6 dro4lets1
aerosols1 s4las:es or s4ills.
3 )o5t: 4i4etting s:o5ld 7e strictly 4ro:i7ited. )ec:anical 4i4etting
de9ices s:o5ld 7e 5sed 6or 4i4etting o6 all liB5ids in t:e la7oratory.
3 Centri65gation s:o5ld 7e done in t57es <it: sa6ety ca4s.
-!) S3" H'*0-'2 )3 shr6s :
3 '!tre8e care s:o5ld 7e 5sed to a9oid a5to3inoc5lation.
3 All c:i44ed or crac=ed glass<are s:o5ld 7e discarded in a44ro4riate
containers.
3 2ro=en glass s:o5ld 7e 4ic=ed 54 <it: a 7r5s: and 4an. +ands 85st
ne9er 7e 5sed.
3 T:e dis4osa7le needles s:o5ld ne9er 7e 8ani45lated1 7ent1 7ro=en1
reca44ed or re8o9ed 6ro8 syringes.
3 T:e 5sed s:ar4s s:o5ld ne9er 7e 4assed directly 6ro8 one 4erson to
anot:er.
3 Al<ays one s:o5ld dis4ose o6 :is;:er o<n s:ar4s.
3 Csed needles s:o5ld 7e discarded in 45nct5re34roo6 rigid containers
@4lastic or card7oard 7o!esA a6ter disin6ection in 0.$31Q 6res:ly
4re4ared sodi58 :y4oc:lorite sol5tion @co88on 7leac:A and ne9er in
ot:er <aste containers. I6 a needle s:redder is a9aila7le1 only t:e
needles or t:e needles along <it: syringe noJJle 8ay 7e s:redded
de4ending 54on t:e ty4e o6 t:e s:redder
3 S:ar4 dis4osa7le containers s:o5ld 7e located close to t:e 4oint o6
5se.
3 S:ar4 dis4osal containers s:o5ld 7e sent 6or dis4osal <:en t:ree3
6o5rt: 65ll.
!) S3" h'*0-'2 )3 s6",-1"'s ( /0))* "(,.) :
35
3 S4eci8ens1 s4ecially 7lood and 7ody 6l5ids s:o5ld 7e collected in 4re3
sterilised scre<3ca44ed 4lastic containers 4ro4erly sealed to 4re9ent
s4illage or lea=age.
3 A5tocla9ed ; 4re3sterilised dis4osa7le syringes and needles 6or
9ene45nct5re or lancets ; c5tting needles 6or 6inger 4ric= s:o5ld 7e
5sed.

3 C5ts in :ands s:o5ld 7e 4ro4erly co9ered <it: <ater4roo6 ad:esi9e
7andages.
3 Dis4osa7le glo9es s:o5ld 7e <orn <:ile collecting 7lood ; 7ody 6l5ids
and 4ro4er ase4sis s:o5ld 7e 8aintained.
3 I6 a sa84le s:o<s e9idence o6 7rea=age @in case not collected in t:e
a7o9e containerA1 lea=age or soiling1 it s:o5ld 7e trans6erred <it: a
glo9ed :and into a second sterile container. Any 4ertinent
in6or8ation s:o5ld 7e re<ritten 6ro8 t:e old to t:e ne< container.
3 I6 t:e reB5isition sli4 is conta8inated <it: 7lood1 it s:o5ld 7e
rejected. In case o6 e8ergency1 t:e conta8inated sli4 8ay 7e
:andled 5sing glo9es.
3 +ands s:o5ld 7e t:oro5g:ly <as:ed <it: soa4 and <ater 4artic5larly
a6ter :andling s4eci8ens.

3 2lood and ot:er s4eci8en containers s:o5ld 7e la7elled <it: a
s4ecial <arning sign >2io:aJard 4reca5tion?. I6 t:e o5tside o6 t:e
container is 9isi7ly conta8inated <it: 7lood it s:o5ld 7e cleaned <it:
disin6ectant. All 7lood s4eci8ens s:o5ld 7e 4laced in s8all lea=34roo6
i84er9io5s 4lastic t57es 6or trans4ortation to t:e la7oratory.
vi4 Safe Handlin# of =lood8=od+ fl6id !5ill! ; In case o6 a s4ill o6 7lood ; 7ody
6l5id in t:e la7oratory1 t:e area s:o5ld 7e 6looded <it: a disin6ectant
sol5tion. eg. 6res:ly 4re4ared 0.$31Q Sodi58 :y4oc:lorite sol5tion
and le6t 6or 30 8in5tes. A6ter <earing glo9es1 t:e area s:o5ld 7e co9ered
<it: 4a4er to<els or ga5Je s4onges to a7sor7 t:e liB5id 6ollo<ed 7y a
t:oro5g: <as: <it: soa4 and <ater. All conta8inated 8aterials s:o5ld
7e dis4osed o6 as in6ectio5s <aste.
He5atiti! B 0accine ; A44ro4riate 5se o6 +2& 9accine 4lays a =ey role in
4re9ention o6 trans8ission o6 +2& 6ro8 4atient to :ealt: care <or=er.
T:ere6ore it is i84ortant t:at all t:e +C.0s incl5ding t:e la7oratory
<or=ers s:o5ld 7e i885niJed as reco88ended 7y .+O.
L+'*r9 '* 0-'"' : 2lthough soiled linen has been identified as a soure of large
nu#bers of ertain %athogeni organis#s, the ris. of atual disease
trans#ission is negligible$
'oiled linen #ay be handled as little as %ossible and ,ith #ini#u# agitation to
%re"ent gross #irobial onta#ination of the air and of %ersons handling the
linen$

36
All soiled linen 85st 7e :andled <it: glo9ed :ands and i6 6easi7le1
deconta8inated in 0.$3 1Q sodi58 :y4oc:lorite in t:e la7oratory
7e6ore sending to t:e la5ndry.
Soiled linen a6ter deconta8ination1 s:o5ld 7e 45t in :ea9y 4lastic
7ags <:ic: are tied and sent to t:e la5ndry. In t:e la5ndry1
deconta8ination in 7leac: is reco88ended in case not done earlier.
T:is s:o5ld 7e 6ollo<ed 7y <as:ing in :ot <ater @O70
0
C A <it:
detergent. Cs5al <as: cycles in <as:ing 8ac:ines =ill +I&.
M'2"1"'( )3 A,,-*"'(0 EA6)s+r" () HIV -' (h" L/)r()r9:
2idental e/%sosure and %ostAe/%osure %ro%hyla/is is disussed in Cha%ter
01$
%eneral 7iosa6ety g5idelines 6or la7oratory <or=ers
i4 Acce!! to la=orator+ !o6ld =e limited at te di!cretion of la=orator+$ in$
car#e .en eH5eriment! are in 5ro#re!!) Te door !o6ld ave a !+m=ol of
2IO+AGA(DF NO AD)ITTANC'. La=oratorie! !o6ld =e .ell ventilated !o
a! to en!6re tat te 5er!onnel do not =reate in contaminated air)
--) E(-'2, *r-'B-'2, s1)B-'2 '* 660-,(-)' )3 ,)s1"(-,s r" 6r)h-/-("*
-' (h" 0/)r()r9. S'*0s '* )6"' s(90" sh)"s *) ')( 33)r* 6r)6"r
3))( 6r)(",(-)' '* r" ')( () /" +s"*. C)'(,( 0"'s"s "s6",-009 (h"
s)3( )'"s sh)+0* ')( /" 8)r'.
---) L/)r()r9 '* 8)rB (/0"s sh)+0* /" s,r+6+0)+s09 ,0"'"* 8-(h
0-C+-* *"("r2"'(s '* *-s-'3",('(s. L/)r()r9 8)rB s+r3," sh)+0*
/" *",)'(1-'("* )'," *9 3("r ,)160"(-)' )3 *9Ds ,(-!-(9 '*
-11"*-("09 3("r s6-00 )3 !-/0" 1("r-0 8-(h *-s-'3",('(.
iv4 Biolo#ical !afet+ ca=inet! and oter 5rimar+ containment device! e)#)
centrif6#e !afet+ ca5! !o6ld =e 6!ed .enever andlin# aBardo6!
!5ecimen! and .en it i! li3el+ to 5rod6ce aero!ol! or infectio6! dro5let!)
v4 Blood and oter !5ecimen container! !o6ld =e la=elled .it a .arnin# !i#n)
Te o6t!ide of te !5ecimen container !o6ld =e cleaned .it !odi6m
+5oclorite !ol6tion in ca!e of vi!i=le contamination)
vi4 Glove! !o6ld =e .orn .ile dealin# .it =lood !5ecimen!, =lood$!oiled
item!, =od+ fl6id!) eHcretion!, !ecretion!, !6rface material! and o=>ect!
eH5o!ed to tem)
vii4 Go.n! 8 la=orator+ coat! m6!t =e .orn .ile .or3in# .it 5otentiall+
infective material! and removed =efore leavin# te la=orator+)
viii4 Hand! !o6ld =e .a!ed immediatel+ after contact .it =lood and =efore
leavin# te la=orator+)
iH4 Mecanical 5i5ettin# device! !o6ld =e 6!ed) Mo6t 5i5ettin# i! !trictl+
5roi=ited) Caref6l tecniC6e! !o6ld =e follo.ed to minimi!e te creation of
aero!ol!)
H4 Accidental .o6nd! from !ar5 in!tr6ment! !o6ld =e avoided) EHtreme
ca6tion i! .arranted .en andlin# needle! and !ar5! to avoid a6to$
37
inoc6lation) Needle! !o6ld =e 5rom5tl+ 5laced in a 56nct6re re!i!tant
container immediatel+ after decontamination)
Hi4 1a5er .or3 !o6ld not =e done on 5otentiall+ contaminated !6rface)
Hii4 All 5otentiall+ contaminated material! and .a!te! from te la=orator+ !o6ld
=e di!5o!ed after decontamination 5refera=l+ =+ a6toclavin#) A la=el .it a
#lo=all+ acce5ted =iolo#ical aBard !i#n !o6ld =e a55lied)
>.=... E33",(-!" +s" )3 s("r-0-E(-)' '* *-s-'3",(-)'
>.=...$.D"3-'-(-)'s :
C0"'-'2 is a %roess ,hih re#o"es foreign #aterial &e$g$ soil, organi
#aterial, #iroAorganis#s* fro# an ob7et$
D-s-'3",(-)' is a %roess ,hih redues the nu#ber of %athogeni #iroA
organis#s, but not neessarily baterial s%ores fro# inani#ate ob7ets or
s.in, to a le"el ,hih is not har#ful to health$
38
H-2h 0"!"0 *-s-'3",(-)' is often used for a %roess ,hih .ills #yobateriu#
tuberulosis and entero"iruses in addition to other "egetati"e bateria, fungi
and #ore sensiti"e "iruses$
S("r-0-E(-)' is a %roess ,hih destroys all #iroAorganis#s inluding
baterial s%ores$
The le"el of deonta#ination should be suh that there is no ris. for infetion
,hen using the e+ui%#ent$
>.=.....C0ss-3-,(-)' )3 -'3",(-)' r-sB 3r)1 "C+-61"'( )r "'!-r)'1"'( -'()
(hr"" ,("2)r-"s '* s+22"s("* 0"!"0s )3 *",)'(1-'(-)'.
; L)8 r-sB : Ite#s in ontat ,ith nor#al and intat s.in, or the inani#ate
en"iron#ent not in ontat ,ith the %atient &e$g$ ,alls, floors, eilings,
furniture, sin.s and drains*$ Cleaning and drying is usually ade+uate e/e%t
,hen there is s%ill of blood! body fluids$
; I'("r1"*-(" r-sB : ;+ui%#ent ,hih does not %enetrate the s.in or enter
sterile areas of the body but is in ontat ,ith #uous #e#branes or nonA
intat s.in, or other ite#s onta#inated ,ith "irulent or trans#issible
organis#s &e$g$ res%iratory e+ui%#ent, gastrointestinal endoso%es, "aginal
instru#ents, ther#o#eters*$ High le"el disinfetion is usually ade+uate$
; H-2h r-sB : Ite#s %enetrating sterile tissues, inluding body a"ities and the
"asular syste# &e$g$ surgial instru#ents, intraAuterine de"ies, "asular
atheters, syringes and needles et$*$ Deonta#ination follo,ed by leaning
and sterilisation is re+uired$ High le"el disinfetion #ay so#eti#es be
a%%ro%riate if sterilisation is not %ossible$
>.=...=.M"(h)*s

C0"'-'2 )3 20ss8r"s :
A Thorough leaning and drying ,ith detergents and ,ater re#o"e #ost
organis#s fro# an ob7et!surfae and should be arried out #etiulously
before steriliDation$
A It #ay be #anual or #ehanial &ultrasoni leaners or ,ashers!
disinfetors * Manual A2ll onta#inated ite#s after %rior disinfetion should
be dis#antled before leaning$ Cold ,ater is %referred as it ,ill re#o"e #ost
of the %rotein #aterials &blood, s%utu#, et$*$ The #ost si#%le, ost
effeti"e #ethod is to thoroughly brush the ite# .ee%ing the brush belo, the
surfae of the ,ater to %re"ent the release of aerosols$ The brush should be
deonta#inated after use and dried$ The ite#s should be rinsed finally in
lean, ,ar# ,ater and dried$ Ite#s are then ready for use for sterilisation$
)ersonnel handling onta#inated ite#s should ,ear good +uality glo"es for
%ersonal %rotetion$
E'!-r)'1"'(0 ,0"'-'2 :
A -loors, surfaes, sin.s and drains should be leaned ,ith ,ar# ,ater and
detergent$ 6outine use of disinfetants is unneessary$
A If there is s%illage of blood, body fluids or s%utu#, disinfetion before leaning
39
is reo##ended$ In high ris. areas or follo,ing s%illage fro# a .no,n
infeted %atient, the surfae is leaned using freshly %re%ared >$GA0F
sodiu# hy%ohlorite solution as desribed under E$@$0$@$ 3lo"es should be
,orn$ 6elease of hlorine gas fro# disinfetion of large s%illage an be
haDardous to staff$ If s%illage is i##ediately re#o"ed, general disinfetion
of the roo# is not neessary$ Thorough leaning ,ill suffie$
D-s-'3",(-)' : Most of the disinfetants used in health are settings in this
ountry, if used in %ro%er onentration and for suitable %eriod of ti#e are
effeti"e against HIV$ ;ither ther#al or he#ial %roesses are used for
disinfetion$ Ther#al disinfetion is %referred ,hene"er %ossible$ It is
generally #ore reliable, lea"es no residue, is #ore easily ontrolled and is
nonAto/i$ (rgani #atter &seru#, blood, %us or faeal #atter* interferes ,ith
the anti#irobial effiieny of either #ethod$ The larger the nu#ber of
#irobes %resent, the longer it ta.es to disinfet$ Boiling &0>>
>
C* for =>A @>
#inutes &holding ti#e* is a "ery si#%le and reliable #ethod for the
inati"ation of all #iroAorganis#s inluding he%atitis B "irus, HIV and
#yobateria$ Carefully done, it is a highAle"el disinfetion %roedure$ The
#ain use of he#ial disinfetants is for heat labile e+ui%#ents ,here single
use is not ost effeti"e$ The %ro%erties of o##on he#ial disinfetants are
sho,n in TableA E$=$
D",)'(1-'(-)'&*-s-'3",(-)' )3 +s"* '""*0"s '* s9r-'2"s :

A The needle is not detahed fro# syringe$
A The disinfetant is as%irated into the syringe$
A The needles and syringes are i##ersed ,ith disinfetant horiDontally in a flat
tray for @> #inutes &-ig E$@*$
A The disinfetant solution is disharged fro# the syringe and needle and rinsed
,ith disinfetant by filling and e#%tying a nu#ber of ti#es$
A The dis%osable syringes and needles are dis%osed as in E$@$@$
A The reusable syringes and needles are autola"ed!boiled for @> #inutes$
A Che#ial disinfetion should ne"er be used as a #ethod of steriliDation
%artiularly for syringes and needles$
40
S("r-0-s(-)' :
A 'terilisation is arried out by stea# under %ressure, dry heat, gas or li+uid
he#ials$
A The hoie of the #ethods li.e autola"ing, use of hot air o"en et$de%ends
on a nu#ber of fators inluding ty%e of #aterial of the ob7et, nu#ber and
ty%es of organis#s in"ol"ed and ris. of infetion to %atients or staff$
A HIV is a fragile "irus and is ade+uately inati"ated by standard sterilisation
%roedures li.e autola"ing at 0G %ounds %ressure for => #inutes at 0=0
>
C or
hot air o"en at 0?>
>
A 01>
>
C for 0 hour &holding ti#e*$
A )ressure oo.er &household %ressure oo.er or 5H(!<8IC;- #odifiation*
ahie"es 0G %$s$i$ e+ui"alent to 0=0
>
C for =>A@> #inutes and #ay be used in
s#all laboratories$
A 2ny steriliDation %roedure should be #onitored routinely by #ehanial,
he#ial and biologial tehni+ues$
A 'terile ite#s should be %roteted against reonta#ination$
41
T/0" >.. D-s-'3",('( 6r)6"r(-"s )3 ,)11)'09 +s"* *-s-'3",('(s.
2nti #irobial ati"ity (ther %ro%erties
'%ore Myo
bateri
a
(ther
bateria
Virus 'table Inati"ation
(rg$ #atter
Corrosi"e!
da#aging
Irritant!
sensitisin
g
;n"$ 8$;n"$
0$ 2lohol
&H>F*NNNN
&0A0> #in$*
8one 3ood 3ood 3ood Moderat
e
Les in
losed
ontainer*
Les
&-i/ati"e*NN
N
'light &Len
e#ent*
8o
=$ Chlorin rel$
#%ds$ &>$GA0F*
&0>A?>
#in$*NNNNN
3ood 3ood 3ood 3ood 3ood 8o &O 0
day*
Les Les LesNNNNN
@$ 3lutaraldehyde
=F &G #in A @
hrs*NNNNN
3ood
&@
hours*
3oodN
&=>
#in$*
3ood
&GA0>
#in$*
3ood
&GA0>
#in$*
3ood
&GA0>
#in$*
Moderate
&0EA=1
days*
8o
&-i/ati"eNN
N*
8o LesNNN
E$ Clear soluble
%henoli &0A
=F*NN
8one 3ood
to
#odera
te
3ood Modera
te
)oor Les NN8o 'light Les
N Less ati"e against M$a"iu# intraellulare ;Aen"elo%ed
NN )otentially to/i, not to be used in neonatal ,ard 8;Anon en"elo%ed
NNN )oor %enetration
NNNN -la##able
NNNNN Irritant!sensitiDing in high onentration
42
>.=.=. S3" *-s6)s0 )3 0/)r()r9 8s("s
Laboratory ,astes are %otential haDards$ Infetious ,aste an trans#it
nu#erous diseases in the o##unity and %ut those ,ho handle ,aste and
li"e on itCs %ro/i#ity, at ris.$ Besides, the inreasing use of dis%osables in
health are is also %osing an additional burden on the ,aste #anage#ent
faility$ It is e/tre#ely i#%ortant that the reyling of these ite#s is
%re"ented$ (nly a s#all %erentage &O0>F* of the ,aste generated in health
are settings are infetious ,hile another GF is nonAinfetious but haDardous$
The #ost %ratial a%%roah to the #anage#ent of bio#edial ,aste is to
identify and segregate infetious ,aste &,ith a %otential for ausing infetion
during handling and dis%osal*, for ,hih so#e s%eial %reautions a%%ear
%rudent$ This ,ill drastially redue the ost of the dis%osal #ethods in health
are settings$
>.=.=.$. S"((-'2 +6 )3 /-)1"*-,0 8s(" 3,-0-(9
;"ery hos%ital, nursing ho#e, "eterinary institution, ani#alAhouse, blood
ban.s, researh institutes generating bio#edial ,astes should install an
a%%ro%riate bio#edial ,aste faility in the %re#ises or should set u% a
o##on faility in aordane ,ith the diretions gi"en by the a%%ro%riate
authority$ Bio#edial ,aste should not be generated ,ithout authorisation$
;"ery hos%ital should ha"e a ,aste #anage#ent %rogra##e 5aste sur"ey is
an i#%ortant %art of the ,aste #anage#ent %rogra##e and hel%s in
deter#ining both the ty%e and +uantity of ,aste being generated in the
hos%ital inluding the laboratory and deter#ine the feasible #ethods of
dis%osal$
>.=.=...C0ss-3-,(-)' )3 0/)r()r9 8s("s :
5astes generated in the laboratory are lassified in TableAE$@

T/0";>.= L/)r()r9 8s("
8on infetious &B>F$* Infetious&0>F* &BioAhaDardous*
)a%er, Cardboard, Ther#ool bo/,
)lasti! )VC, Disarded reagents
'har%s 8on shar%s
8eedles, 'yringes, Blades,
$ 'al%els, 3lass,are
'olids Li+uids
Hu#an ,aste, blood !
blood %roduts, body
fluids, ani#al ,aste
#irobiologial! biotehnologial
,aste, soiled linen! gauDe! otton
>.=.=.=.C)'(-'-'2 8s(" ( 2"'"r(-)' 6)-'( : 2t the generation %oint ie$ the
laboratory, ,aste is #anaged in the follo,ing ,ay :
i * Colletion
ii * 'egregation and ,eighing
iii * 'torage
Ws(" s"2r"2(-)' is the .ey to any ,aste #anage#ent she#e$ It
onsists of %laing different ty%es of ,aste in different ontainers or olourA
odedAbags at the site of generation$ This hel%s in reduing the bul. of
infetious ,aste and ontains s%read of infetion to general ,aste$ This
43
%ratie redues the total treat#ent ost, the i#%at of ,aste in the
o##unity and the ris. of infeting ,or.ers$ )ro%er segregation should
identify ,aste aording to soure and ty%e of dis%osal! disinfetion$
5aste should be segregated into different ategories &TableAE$@* at the site of
generation ie$ in the laboratory and ,eighed se%arately at the ti#e the ,aste
is being dis%osed$
S)0-* N)' -'3",(-)+s ,aste is olleted in bla. bags and dis%osed as
household ,aste$
Infetious ,aste #ust be se%arated at the %oint of generation itself and
should be deonta#inated %rior to itCs storage, trans%ort and dis%osal$ 'olid
infetious ,aste are dis%osed as follo,s :
A 'har%sA a* 8eedles and syringe noDDleA shredded in needleAdestroyer &if
a"ailable*J if not a"ailable deonta#inated as in b*$
A b* 'al%el blades! lanets ! bro.en glassAshould be %ut in se%arate ontainer
,ith bleah, transferred to %lasti! ardboard bo/es, sealed to %re"ent
s%illage and trans%orted to ininerator$
A 3lass ,aresA should be disinfeted, leaned and sterilised$
A Culture %lates ,ith "iable ultureA should be autola"ed in %lasti autola"able
bags, #edia are re#o"ed and olleted in yello, bags and dis%osed of by
inineration! #iroA,a"ing! hot air o"en$ The %lates an be reused after
steriliDation$
A ',absA should be he#ially disinfeted follo,ed by inineration$
A Dis%osable ite#s inlude single use %roduts &syringes, glo"es, shar%s et$*A
2s these ite#s are often reyled and ha"e the ris. of being reused illegally,
these should be disinfeted by di%%ing in freshly %re%ared 0F 'odiu#
hy%ohlorite for @> #inutes to 0 hour$ Bins ,hih an be used for this
%ur%ose are a set of t,in bins, one inside the other ,ith the inner one being
%erforated and easily e/tratable$ This #ini#iDes ontat ,hen the ontents
are being re#o"ed$
Dis%osable ite#s li.e to glo"es, syringes et$ should be shredded ut
or #utilated before dis%osal follo,ed by dee% burial or %ro%erly aounted
before dis%osal$ ;/tre#e are should be ta.en ,hile handling the needles$
L-C+-* 8s("s generated by the laboratory are either %athologial or
he#ial in nature and are dis%osed of as follo,s:
44
A 8onAinfetious he#ial ,aste should first be neutralised ,ith reagents and
then flushed into on"entional se,er syste#$

A The li+uid infetious ,aste should be treated ,ith a he#ial disinfetant for
deonta#ination then neutraliDed and flushed into the se,er$
C)00",(-)' /2s. 'olid ,astes are olleted in lea.Aresistant single hea"y
duty bags or double bags #ay be used$ Bags ha"ing different olour odes
&Table E$E* ,ith red labels #entioning date and details of ,aste are
reo##ended$ The bags are tied tightly after they are threeAfourths full$
%,B-'2 , s()r2" '* (r's6)r(
A 2ll segregated and disinfeted ,aste should be %a.ed in %ro%er ontainers
and olourAoded bags & Table E$E* ,ith red labels #entioning details of
bio#edial ,aste and biohaDard signs &-igAE$E$*$ 2ll ontainers used for
storage of suh ,aste shall be %ro"ided ,ith a %ro%erly o"ered lid$
A 'uh ontainers should be inaessible to sa"engers and %roteted against
insets, birds, ani#als and rain$
A There should not be any s%illage during handling and transit of suh ,aste$
A The ,aste shar%s, after %reAtreat#ent should be bro.en before %a.ing in the
ontainer$
A The ,aste should be trans%orted in "ehiles authorised for this %ur%ose only$
A 8o suh ,aste should be stored in the %lae ,here it is generated for a %eriod
of #ore than t,o days$
>.=.=.>. Tr"(1"'( '* *-s6)s0
D-s6)s0 1"(h)*s: Dis%osal #ay be done by :
A Muni%al or%oration
A 'anitary landfill
A If ininerator is not a"ailable, dee% burial in ontrolled landfill sites is
reo##ended$ Deonta#ination should be arried out before burial$
A Inineration &Te#%$HG>
>
C*
InineratorA burns !redues the infetious ,aste to ashes and therefore
fa"oured by hos%itals$ It #ay be of t,o ty%es Mo##on or indi"idual$ There
are so#e disad"antages li.e %ollution !ino#%lete #elting of needles$
Hos%itals ,ith #ore than @> beds or P0>>> %atients %er #onth should ha"e
an ininerator$ )lastis annot be ininerated$
45
>.=.=.?. G+-*"0-'"s 3)r 8s(" *-s6)s0 :
A 2ll bio#edial ,astes should be treated and dis%osed of stritly in aordane
,ith the o%tions #entioned in the table E$E$
A 5aste ,hih annot be ininerated, &%lastis* should be %reAtreated by
disinfetion and dis%osed of in an en"iron#entally sound #anner$
A 5aste should not be du#%ed, disharged or dis%osed in any %lae other than
a site identified for the %ur%ose$
A 2ll %reautions and %ersonal safety #easures should be ta.en &inluding
%ro"ision of %roteti"e lothing, #as.s, glo"es, gu#boots, goggles, et$ as
#ay be neessary*$ He%atitis B "aine is reo##ended for affording
%rotetion to all %ersonnel engaged in handling bio#edial ,aste, or being
e/%osed to suh ,astes against infetion fro# handling or e/%osure$
A 2ll treat#ent and dis%osal failities should be loated at a s%eified area a,ay
fro# the general ser"ie area of the hos%ital, %ubli %laes and residential
areas$
A 5hen the treat#ent o%tion is burial, the %its should be loated at sites a,ay
fro# agriultural land, residential areas, groundAand safe ,ater soures$
There should be no lea.age fro# the %its in to surrounding areas$
A 2ll %lastis should be disinfeted, shredded and dis%osed of in an
en"iron#entally friendly #anner$ 6eyling of dis%osables eg$ syringes,
needles, glo"es, transfusion bags et$ should be %re"ented$
A 2ll li+uid ,aste should be disinfeted and flushed in the sin.s at the %oint of
generation$
A Bio#edial ,aste should not be dis%osed of on o%en land and #unii%al
dustbins$ <ntreated li+uid ,aste should not be let into se,ers$
>.=.=.F.M-'("''," )3 r",)r*s : 'e%arate reords for lassifiation of ,aste and
their regular dis%osal should be #aintained in the laboratory$ The ,aste
dis%osal %rogra##e should be su%er"ised and #onitored regularly$

>.=.=.G.R"6)r(-'2 )3 ,,-*"'(s : In the e"ent of an aident ourring at any
loation or site ,here bio#edial ,astes are handled or during trans%ortation,
the a%%ro%riate authorities #ust be infor#ed and needful ation ta.en$
>.>. Tr-'-'2 : Training regarding need of and national guidelines of biosafety
%raties is e/tre#ely i#%ortant and should be %ro"ided at regular inter"als
for different le"els of health are ,or.ers$ 3uidelines for biosafety should be
%ro"ided ,hih #ay be #odified fro# ti#e to ti#e aording to re+uire#ent$
46
T/0";>.> C)'(-'"r '* ,)0)+r ,)*-'2 3)r *-s6)s0 )3 /-);1"*-,0 8s("s
5aste
atego
ry
5aste lass Ty%e of ontainer Colou
r
odin
g
Treat#ent !
Dis%osal
o%tion
8o$ 0 Hu#an anato#ial
,aste, blood 4
body fluids
'ingleAuse
ontainers!
)lasti holding bags
6ed Inineration !
Dee% Burial
8o$ = 2ni#al 4 slaughter
house ,aste
'ingleAuse
ontainers!
)lasti holding bags!
sas
(ran
ge
Disinfetion 4
Dee% Burial
8o$ @ Mirobiology 4
bioAtehnology
,aste
'ingleAuse
ontainers!
)lasti holding bags
Lello
,
2utola"ing !
Miro,a"ing 4
Inineration
8o$ E 5aste shar%s 6eAusable! singleA
use sturdy
ontainers of %lasti,
glass or #etal
Blue 'hredding 4
Dee% Burial
8o$ G Disarded
#ediines
6eAusable! sturdy
ardboard! glass!
%lasti holding bags
Blue 'hredding 4
Dee% Burial
8o$ ? 'oiled ,astes
&linen*
)lasti bags! sas Lello
,!
Bla.
Disinfetion
and #ahine
leaning
8o$ H Dis%osables &other
than shar%s*
6eAusable! sturdy
ontainers! %lasti
holding bags
Lello
,!
Bla.
DisinfetionA
Che#ial !
2utola"ing,
'hredding,
Burial
8o$ 1 Li+uid ,astes 82 82
8o$ B Che#ial ,astes 'turdy ontainers!
%lasti holding bags
Lello
,!
Bla.
>.?. I160"1"'((-)' )3 /-)s3"(9 6r,(-,"s -' 0/)r()r9 : 2lthough
guidelines regarding uni"ersal %reautions and other biosafety %raties are
a"ailable sine long,strit i#%le#entation is not in %ratie in healthare
settings in India e"en in the a%ital ity$ 5ith inrease in the %re"alene of
HIV infetion, there is a definite need that the HC5Cs ta.e biosafety %raties
seriously$ -or effeti"e o#%liane, the laboratory #anagers should ensure
ade+uate su%%ly of %ersonal %roteti"e e+ui%#ents, a"ailability of #aterials
for hand,ashing, disinfetants and set u% an effeti"e ,aste dis%osal
%rogra##e for dis%osal of bio#edial ,astes$
>.F S)+r," )3 0-("r(+r" '* s+22"s("* 3+r(h"r r"*-'2s
i* Medial Manage#ent of HIV Infetion 0BB1 edition$ ;ditor 9ohn 3$ Bartlett
9ohns Ho%.ins <ni"ersity Di"ision of Infetious Diseases, <$'$2$
iiA %lo7al Progra88e o6 AIDS3(e4ort o6 a .+O Cons5ltation on t:e
4re9ention o6 +58an I885node6iciency &ir5s and +e4atitis 2 9ir5s
trans8ission in :ealt: care setting %ene9a1 1131# A4ril 1DD1.
iiiA )inistry o6 'n9iron8ent and orests noti6ication Ne< Del:i t:e 16
t:
Octo7er1 1DD7.
i9A S:ris:ti: I84le8enting +os4ital .aste )anage8ent: A g5ide 6or
:ealt: care 6acilities. Se4te87er1 1DD6.
47
9A .+O ,a7oratory 2iosa6ety )an5al #
nd
'dition1 1DD3 .orld +ealt:
Organisation1 A.I.T.2.S. P57lis:ers and Distri75tors.
9iA .+O )anaging )edical .aste in De9elo4ing Co5ntries1 1DD" .orld
+ealt: Organisation1 %ene9a.
9iiA AIDS in India. Ne<s letter o6 t:e National AIDS Control Organisation1
Se4te87er1DD3.
9iiiA Pers4ecti9es in Disease Pre9ention and +ealt: Pro8otion 54date
Cni9ersal Preca5tions 6or 4re9ention o6 trans8ission o6 +58an
I885node6iciency &ir5s1 +e4atitis 2 &ir5s and ot:er 2lood37orne
Pat:ogens in +ealt:3care Settings. )).( 37@#"A1 37733EE.
i!A Centers 6or Disease Control. (eco88endations 6or 4re9ention o6 +I&
trans8ission in :ealt:3care settings. )).( 1DE7:36 @s544l no.#S A.
48
?. COLLECTION AND TRANS%ORT OF S%ECIMENS
FOR HIV TESTING
2l#ost all the laboratory %roedures for HIV testing are %erfor#ed on
%atientsC bloodJ seru# or %las#aJ hene the olletion of blood is desribed
belo,$
?.$ %"r3)r1-'2 !"'"6+',(+r" :
3 %lo9es s:o5ld 7e <orn and sterilised ;dis4osa7le syringes and
needles s:o5ld 7e 5sed.
3 or a9oiding soiling1 a 4iece o6 linen <it: a layer o6 dressing 4ad @ a
s:eet o6 a7sor7ent cotton 7et<een t<o layers o6 ga5Je 4ieceA or
si84ly a 7ig 4iece o6 a7sor7ent cotton 8ay 7e 4laced 7elo< t:e
6orear8 7e6ore co88encing 9ene345nct5re.
3 A6ter collecting $ 8l o6 7lood ase4tically1 it s:o5ld 7e care65lly
trans6erred 6ro8 t:e syringe <it:o5t sB5irting in to a sterile 4lastic
lea= 4roo6 s4eci8en container 4re6era7ly scre< ca44ed. T:e
containers s:o5ld 7e la7elled 7e6ore co88ence8ent o6
9ene45nct5re. I6 t:e 9ial :as anticoag5lants1 t:en a second 4erson
<earing glo9es <o5ld :a9e to :el4 in s:a=ing t:e 9ial 6or 8i!ing t:e
7lood <ell <it: t:e anticoag5lants. T:e ca4 8ay 7e tig:tly scre<ed
a6ter t:e 7lood :as 7een trans6erred to t:e 9ial.
3 A6ter 7lood is collected1 t:e to5rniB5et is re8o9ed and t:e needle is
<it:dra<n. T:e 4atient is gi9en a dry sterile cotton s<a7 to 4ress
o9er t:e site o6 9ene345nct5re. 'l7o< 8ay 7e 6le!ed to =ee4 t:e
cotton s<a7 in 4lace till t:e 7lood sto4s. Any 7lood s4ill is care65lly
<i4ed <it: 70Q et:anol.
3 All t:e s<a7s and cotton 4ieces are 4laced in 4lastic 7ags 6or
dis4osal. I6 t:e o5tside o6 t:e 9ial is 9isi7ly conta8inated <it: 7lood1
it s:o5ld 7e cleaned <it: 10Q 6res:ly 4re4ared sodi58 :y4oc:lorite
sol5tion.
3 T:e 7lood is allo<ed to clot 6or 30 8in5tes @not 8ore t:an # :o5rsA at
roo8 te84erat5re. T:e clot 8ay 7e gently 7ro=en i6 necessary 5sing
sterile 4aste5r 4i4ettes.
')" Se5aration of !era !am5le! ;
3 T:e 9ial is centri65ged at 1#00g @3000 r48A 6or 10 8in5tes to
se4arate ser58 to a9oid :ae8olysis. I6 no centri65ge is a9aila7le1 t:e
7lood <it: clot 8ay 7e le6t in t:e re6rigerator at R"
0
C 6or o9ernig:t.
T:e clot <ill retract and get se4arated 6ro8 ser58.
3 T:e s4eci8en 9ial is 5n3sto44ered1 t:e ser58 is dra<n o66 7y sterile
4aste5r 4i4ette and trans6erred to a sterile 4lastic scre< ca44ed lea=
4roo6 t57e.
49
')< Addition of 5re!ervative ;
3 Preser9ati9es s:o5ld not 7e added since it inacti9ates conj5gates and
gi9es rise to 6alse serological res5lts.
3 I6 necessary1 $ 7ro8o1 $ nitro1 133 dio!ane in 4ro4ylene glycol at a
6inal concentration o6 0.0$Q is reco88ened as 4reser9ati9e @c:a4ter
10 and 11A.
3 T:io8ersal at a 6inal concentration o6 0.01Q is e66ecti9e only 6or a
6e< <ee=s as it loses acti9ity <:en e!4osed to lig:t.
')( Stora#e of !er6m !5ecimen! ;

3 T:e sera sa84les are 4laced in lea= 4roo6 4lastic containers in t:e
re6rigerator at R"
0
C1 6or storage.
3 T:e o5tside o6 t:e container is c:ec=ed 6or 9isi7le conta8ination <it:
7lood <:ic: s:o5ld 7e cleaned as in $.1.
3 T:en t:e s4eci8en 9ials are 4ac=ed in a second tig:tly ca44ed
5n7rea=a7le container s5rro5nded 7y adeB5ate 4ac=ing 8aterial.
or storage 6or a long ti8e1 dee436reeJing at T70
0
C is ad9ised.
')') Tran!5ort of !era !am5le! ;
3 T:e s4eci8en t57e in <:ic: ser58 is to 7e trans4orted1 s:o5ld not
:a9e crac= or lea=age. Pre6era7ly1 it s:o5ld 7e 8ade o6 4lastic and
s:o5ld 7e scre< ca44ed. T:e o5tside o6 t:e container is c:ec=ed 6or
any 9isi7le conta8ination <it: 7lood <:ic: s:o5ld 7e disin6ected as
in $.1.

3 T:e t57e is la7elled and t:en 4laced in a second tig:tly ca44ed
5n7rea=a7le container @ig.$.1A s5rro5nded 7y adeB5ate 4ac=ing
8aterial @li=e tiss5e 4a4er1 adsor7ent cotton etc.A to a7sor7 liB5id1 i6
lea=age occ5rs accidentally.

3 T:e secondary container s:o5ld also :a9e a la7el. T:is is 4laced in a
t:er8ocol 7o! :a9ing ice 4ac=s to 8aintain 4ro4er cold c:ain syste8
d5ring transit.

3 A 4ro6or8a <it: details i.e. na8e1 age 1 se! 1 ris= 6actors1 :istory o6
4re9io5s testing etc .s:o5ld acco84any t:e s4eci8en.
3 A 7io:aJard sy87ol @ig3$.#.A 85st 7e 6i!ed o5tside t:e t:er8ocol
7o!. T:is 7o! can no< 7e sent to a distant la7oratory.
')?) Tran!5ort of .ole =lood for vir6! i!olation and CD(, CD@ !t6die!;
3 T:e 5ni9ersal 4reca5tions as 8entioned a7o9e s:o5ld 7e 6ollo<ed.
3 2lood is collected in :e4ariniJed ; 'DTA scre< ca44ed 9ials.
3 It is t:en 4laced in a second 5n7rea=a7le container <it: adeB5ate
4ac=ing 8aterial.

3 S:o5ld 7e trans4orted in <et ice so as to reac: t:e la7oratory <it:in
#" :o5rs.
50
3 or 9ir5s isolation1 i6 t:e s4eci8en cannot 7e trans4orted
i88ediately a6ter collection1 it s:o5ld 7e 6roJen
51
?) HI0 anti=od+ te!tin# .it !5ecial reference to HI0$%
?)% Introd6ction ; +I&;AIDS is not li=e ot:er in6ectio5s diseases. It is 6ar
8ore co84le! 7eca5se +I& in6ection is li6e long1 o5tco8e is in9aria7ly
6atal and no c5re or 9accine is a9aila7le so 6ar. Since1 co88only
+I&;AIDS is acB5ired t:ro5g: se!5al contact1 indi9id5als =no<n to 7e
+I& in6ected are stig8atiJed and discri8inated against. A n587er o6
8oral1 et:ical1 legal and 4syc:osocial iss5es are related <it: a 4ositi9e
+I& stat5s. So1 anyone atte84ting to assess t:e +I& stat5s o6 an
indi9id5al 85st 7e con9ersant <it: t:ese iss5es1 strategies o6 +I&
testing1 4rotocols o6 testing1 rationale o6 5sing test =its1 correct 8et:od
o6 in6or8ing t:e client1 co5nselling1 i84ortance o6 con6identiality1
tec:nical and ot:er 4it6alls and B5ality ass5rance to na8e so8e.
Con6identiality o6 a 4ositi9e test res5lt is o6 5t8ost i84ortance.
Co5nselling s:o5ld 7e 5nderta=en to 8oti9ate t:e indi9id5al to tell t:e
s4o5se;6a8ily and ind5ce 7e:a9io5r c:ange.
Anot:er i84ortant 4oint to 5nderstand is t:at la7oratory
diagnosis is t:e only 8et:od deter8ining t:e +I& in6ection stat5s o6 an
indi9id5al d5ring t:e long asy84to8atic 4eriod.
?)" 16r5o!e of HI0 te!tin# ;
3 In6or8ation is 5se65l 6or 4ro4:yla!is1 8edical 8anage8ent and
treat8ent o6 +I& and related illnesses.
3 To ass5re 7lood sa6ety and donation sa6ety.
3 To assess t:e e66icacy o6 targeted inter9ention in a de6ined
co:ort.
3 To 8onitor trends o6 e4ide8ic @sentinel s5r9eillance etc.A.
3 Identi6ication o6 asy84to8atic indi9id5als @4ractising :ig: ris=
7e:a9io5rA.
3 To 4lan 4ersonal and 6a8ily0s 65t5re i6 t:e res5lt is 4ositi9e.
3 To 8oti9ate 6or 7e:a9io5r 8odi6ication t:ro5g: co5nselling
a8ongst t:ose <:o test negati9e and <:o 4ractise :ig: ris=
7e:a9io5rs.
3 To ind5ce 7e:a9io5r c:ange and 4re9ent trans8ission 7y
co5nselling in t:ose <:o test 4ositi9e.
3 To diagnose clinically s5s4ected cases.
3 or 4eace o6 8ind o6 indi9id5als 4ractising :ig: ris= 7e:a9io5r.
?)< Iinetic! of 6moral imm6ne re!5on!e ; An 5nderstanding o6 t:e seB5ence
o6 e9ents t:at 6ollo< t:e entry o6 9ir5s into t:e 7ody <ill :el4 to
5nderstand t:e o4ti8al 5sage o6 9ario5s +I& tests d5ring di66erent
stages o6 +I& disease.
&iral entry into t:e 7ody leads to a transient 4eriod o6 4las8a
9irae8ia @5s5ally :ig: le9elA and 4#" antigenae8ia. +o<e9er1 t:e
le9els o6 t:ese co84onents co8e do<n <it: conco8itant i885ne
res4onse. +58oral res4onse is e9idenced 7y 6or8ation o6 anti7odies o6
di66erent classes against di66erent str5ct5ral 4roteins @gag: 41$1 4171
4#"1 4$$F en9: g4 "11 g4 1#01 g4 160F and 4ol: 4311 4$1 and 466A1
reg5latory 4roteins @ne61 re91 tatA and accessory 4roteins @9i6 and 945A.
All str5ct5ral co84onents are strongly i885nogenic and ind5ce
6or8ation o6 anti7odies1 <:ereas i885nogenicity o6 reg5latory and
accessory 4roteins 4roteins is 9aria7le.
T:e anti7odies a44ear in 7lood <it:in #3E <=s a6ter in6ection 75t
5s5ally 7eco8e detecta7le a6ter 3 <=s to 1# <=s. <it: t:e assays
52
a9aila7le 4resently. T:is 4eriod 6ollo<ing t:e entry o6 +I& into t:e
7ody and t:e a44earance o6 detecta7le le9els o6 anti7odies <it: t:e
a9aila7le test =its is called t:e >.indo. 5eriod?. D5ring t:is 4eriod t:e
indi9id5al is in6ected1 in6ectio5s and non3reacti9e <it: t:e anti7ody
detection tests. T:e anti7odies to gag 4rotein @4#" and 4$$A are 6irst to
a44ear 5s5ally1 t:o5g: anti7odies to en9 4roteins and 4ol 4roteins 8ay
also 7e 4rod5ced si85ltaneo5sly. As in6ection 4rogress to AIDS
anti7ody to 4#" 5s5ally declines as 4#" antigen le9els rise conco8itant
<it: 4rogression o6 disease to AIDS. +o<e9er1 anti7odies to en9
4roteins 4ersist t:ro5g:o5t t:e in6ection @ig. 6.1A.
Anti3+I& anti7odies 8ay 7e IgA1 Ig) and Ig%. T:e IgA and Ig)
res4onses are inconsistent. Ig% res4onse is consistent and 7etter
5nderstood. Ig) res4onse a44ears earlier t:an Ig% 75t t:e sensiti9ity
o6 assays a9aila7le to detect t:is class o6 i885noglo75lins is lo< and
also Ig) is detecta7le 6or a s:ort 4eriod. Detection o6 Ig) is 9al5a7le
6or identi6ying early serocon9ersion 4artic5larly 6ollo<ing needle stic=
inj5ry @#311 daysA and in6ection in ne<7orn. IgA is t:e 4redo8inant
i885noglo75lin in sero85co5s secretions @sali9a1 colostr581
genito5rinary secretions etc.A. Since IgA does not cross 4lacenta and
t:ere6ore detection o6 anti3+I& IgA 7y +I&3IgA assay in a ne<7orn is
diagnostic o6 congenital +I& in6ection. +o<e9er1 'nJy8e ,in=ed
I885nosor7ent Assay @',ISAA and .estern 2lot @.2A to detect IgA
and Ig) need 65rt:er e9al5ation.
?)( Informed con!ent after 5rete!t co6n!ellin# ; @See c:a4ter 1$ 6or co5nselling
6or +I& testingA. +I& testing 6or t:e 45r4ose o6 identi6ication o6 an
indi9id5al 85st al<ays 7e 5nderta=en a6ter 4re test co5nselling and
in6or8ed consent. Testing <it:o5t e!4licit consent :as 4ro9en to 7e
co5nter 4rod5cti9e and :as dri9en t:e +I& 4ositi9e indi9id5als
5ndergro5nd. T:is 8a=es instit5tion o6 4re9ention and inter9ention
8eas5res 8ore di66ic5lt. Pretest co5nselling also 4re4ares t:e
indi9id5al to 6ace t:e +I& test res5lt.
?)' Confidentialit+ ; T:e con6identiality o6 t:e test res5lt @7ot: negati9e as
<ell as 4ositi9eA s:o5ld 7e strictly 8aintained in 8ost cases. T:is is to
res4ect t:e 4ri9acy and rig:ts o6 t:e indi9id5als and to 4rotect t:e8
6ro8 discri8ination1 9icti8iJation and stig8atiJation. T:e test res5lt1
na8e o6 t:e indi9id5al etc. 85st ne9er 7e disc5ssed loosely. T:e test
re4ort 85st 7e 4laced in a sealed en9elo4e and s578itted to t:e
clinician <:o reB5isitioned t:e test. T:e en9elo4e s:o5ld 7e 8ar=ed
>con6idential?. T:e records in t:e la7oratory 85st also 7e =e4t sec5re
to 4re9ent access 7y 5na5t:orised 4ersons. As 6ar as 4ossi7le t:e
res5lts ne9er 7e co885nicated 9ia tele4:ones.
?)? Detection of HI0 !5ecific anti=odie! ;Detection o6 anti3+I& anti7odies is t:e
8ainstay o6 testing 6or +I& and diagnosis o6 +I&.
Tests to detect s4eci6ic +I& anti7odies can 7e classi6ied into :
3 Screenin# te!t! 2ELISA8EIA, *a5id, Sim5le4
3 S655lemental te!t!
Screenin# te!t! are 4er6or8ed to screen 5nits o6 donated 7lood and 7lood
4rod5cts and 6or s5r9eillance. T:ese incl5de a 8yriad o6 ',ISAs <:ic:
5s5ally ta=e #33 :rs to yield res5lts. (a4id screening tests gi9e res5lts
<it:in 8in5tes and incl5de 9is5al assays li=e dot37lot tests1 4articles
@gelatin1 (2C1 late!1 8icro7eadsA aggl5tination1 +I& s4ot and co87 test
53
and 6l5oro8etric 8icro4article tec:nologies. Si84le screening tests are
also 7ased on ',ISA 4rinci4le 75t ta=e a7o5t :al6 an :o5r or so.
S655lemental te!t! are 4er6or8ed on ser58 sa84le reacti9e in screening
test 6or t:e 45r4ose o6 diagnosis and identi6ication o6 t:e indi9id5al.
.:en a ser58 s4eci8en is reacti9e 7y any one o6 t:e screening tests it
:as to 7e tested again 7y a di66erent syste8 5sing di66erent +I&
antigens or di66erent 4rinci4le o6 test to con6ir8 t:e diagnosis. I6 a
s4eci8en is reacti9e in # di66erent syste8s it :as to 7e tested again
5sing one o6 t:e s544le8ental tests <:ic: 8ay 7e a t:ird
',ISA;(a4id;Si84le test or a .estern 2lot test @.2A as t:e case 8ay
7e.
A :ealt:y indi9id5al reacti9e in t:ree di66erent syste8s o6 testing
is con6ir8ed to 7e :a9ing +I& in6ection. T:e ot:er s544le8ental tests
li=e ..2.;I are 5sed to resol9e discordant res5lts o6 ',ISA and
researc:1 as 6ar as India is concerned as t:ese tests are e!4ensi9e1 ti8e
cons58ing and need e!4ertise.
.:ic:e9er co88erical =it is selected it s:o5ld 7e ens5red t:at it
detects anti7odies against 7ot: +I&311 +I&3# and t:eir s57ty4es.
Presence o6 +I& anti7odies indicates t:e indi9id5al is in6ected
<it: +I& and can trans8it in6ection to ot:ers t:ro5g: 5nsa6e ris=
7e:a9io5r.
?)7 S5ecimen! to =e collected for detection of anti$HI0 anti=odie! ; )ost co88on
s4eci8en is 7lood. +I& test =its are a9aila7le 6or detecting +I&
anti7odies in 9ario5s =inds o6 s4eci8ens li=e 7lood1 4las8a1 ser581
sali9a and 5rine.
Anti7ody detection :
3 2lood;ser58;4las8a
3 Sali9a not as good
3 Crine as 7lood
?)@ HI0 anti=od+ !creenin# a!!a+! ; T:e list o6 t:e screening assays 6or +I&
testing is gi9en 7elo<
3 ',ISA @#33 :o5rsA
3 (a4id tests @)in5tesA
3 Dot 7lot assays
3 Particle aggl5tination
3 +I& s4ot and co87 tests
3 l5ori8etric 8icro4article tec:nologies
3 Si84le : 2ased on ',ISA 4rinci4le
Ta=e 1;# an :o5r
',ISA is t:e 8ost co88only 4er6or8ed screening test. Screening
assays 85st detect all 4ositi9e sera i.e. s:o5ld 7e :ig:ly sensiti9e e9en
i6 so8e 6alse 4ositi9es do occ5r. +o<e9er1 res5lts o6 a screening test
s:o5ld ne9er 7e 5sed as t:e 6inal inter4retation o6 +I& stat5s1 and
indi9id5al is ne9er identi6ied on t:e 7asis o6 one screening assay as
tec:nical errors can occ5r. T:e ser58 reacti9e in screening assay is
s57jected to con6ir8atory tests @as 4er 4olicy and strategy o6 testingA
to 7e classi6ied as reacti9e only i6 it is reacti9e in re4eated assays.
Ot:er screening tests <ere also introd5ced s57seB5ently. T:ese
incl5de late!1 red cell and gelatin 4article aggl5tination1 co87 tests and
54
dot3 7lot assays. T:ese tests are easy to 4er6or81 are ra4id1 do not
reB5ire so4:isticated eB5i48ent1 tec:nical e!4ertise and are 8ostly
cost e66ecti9e. So8e o6 t:e81 4artic5larly co87 tests and dot3 7lot
assays1 are also discri8inatory 6or +I& 1 and +I& # anti7odies.

?)A ELISA a! te !creenin# a!!a+
',ISA is t:e 8ost co88only 4er6or8ed screening test. It is easy
to 4er6or81 ada4ta7le to large n587er o6 sa84les1 is sensiti9e and
s4eci6ic and cost e66ecti9e. So8e o6 t:e assays ta=e $ 8in5tes and
ot:ers as long as ## :rsF so8e need only 1l ser581 ot:ers 8ay need as
85c: as E00l F so8e need so4:isticated instr58ents1 ot:ers need
noneF so8e are easy to 4er6or8 and ot:ers need e!4ertise and are
di66ic5lt to 4er6or8F so8e are e!4ensi9e and so8e are not so e!4ensi9e1
so t:e a44ro4riate test c:oice can 7e 8ade ta=ing into consideration
t:e a9aila7le reso5rce1 storage 6acility1 tec:nical e!4ertise a9aila7le1
in6rastr5ct5re a9aila7le1 o7jecti9e o6 testing1 4re9alence o6 in6ection
and 4er6or8ance c:aracteristics o6 test =its etc.
?)A)% Different t+5e! of HI0 3it! 2=a!ed on t+5e of HI0 anti#en4 availa=le
commerciall+ ; irst generation o6 ',ISA de9elo4ed <ere 9ery sensiti9e
75t not s4eci6ic 7eca5se <:ole 9iral lysates <ere 5sed as antigen.
T:ese lysates 5s5ally contained s8all a8o5nts o6 :ost cell co84onents
<:ic: ga9e rise to 6alse 4ositi9e reactions. T:e ',ISA tec:nologies
<ere i84ro9ed and #nd and 3rd generation =its <ere de9elo4ed 5sing
reco87inant and synt:etic 4e4tides as antigens. So1 ',ISA assays
a9aila7le in t:e 8ar=et 8ay 7e:
3 irst generation : 5se antigens deri9ed 6ro8 detergent disr54tion o6
9ir5ses gro<n in :58an ly84:ocytes.
3 Second generation :5se arti6icially deri9ed reco87inant antigens
e!4ressed 6ro8 7acteria or 65ngi.
3 T:ird generation : 5se c:e8ically synt:esiJed oligo4e4tides o6 a7o5t
1$3"0 a8ino acids @synt:etic 4e4tidesA.
?)A)" 1rinci5le! of ELISA
On t:e 7asis o6 t:e 4rinci4le o6 t:e test ',ISA can 7e di9ided into :
3 Indirect
3 Co84etiti9e
3 Sand<ic:
3 Ca4t5re assays
All ',ISA consist o6 eit:er +I& antigen or anti7ody @de4ending 54on
t:e 4rinci4leA attac:ed on a solid 4:ase @8atri! or s544ortA and1
incor4orate a conj5gate and s57strate detection syste8. &iral antigens
8ay 7e <:ole 9ir5s lysates1 reco87inant or synt:etic 4e4tides. T:e
8atri! can 7e ><ells? or >stri4s? o6 a 8icro4late1 4lastic 7eads or
nitrocell5lose 4a4er. Conj5gates are 8ost o6ten anti7odies @Ig%1
so8eti8es Ig) and IgA alsoA co54led to enJy8es @al=aline
4:os4:atase or :orseradis: 4ero!idaseA1 6l5oroc:ro8es or ot:er
reagents t:at <ill s57seB5ently 7ring a7o5t a reaction t:at can 7e
9is5alised. In case o6 enJy8e conj5gates t:e signal generated is a
colo5r reaction and in case o6 6l5oroc:ro8e it is 6l5orescence. T:e
s57strates 5sed are "3nitro4:enyl4:os4:ate 6or al=aline 4:os4:atase
and o34:yn!ylencdia8ine di:ydroc:loride @OPDA and T)2 6or
:orseradis: 4ero!idase1 <:ic: 4rod5ce colo5r on 7eing acted 54on 7y
55
t:e res4ecti9e enJy8es and t:e colo5r can 7e eit:er detected 9is5ally
or 8eas5red on a ',ISA (eader as OD 9al5es.
All =it controls as s4eci6ied in t:e 4ac=age insert o6 t:e =it 85st 7e
incl5ded <it: eac: test r5n to 9alidate t:e test. Internal controls also
85st 7e incl5ded <it: eac: r5n to ens5re B5ality res5lts. Pre4aration
o6 internal controls 6or 4ractising B5ality ass5rance is detailed in t:e
c:a4ter 10 and 11 .
?)A)")% Indirect ELISA
+I& antigens are attac:ed co9alently to t:e solid 4:ase s544ort
allo<ing +I& anti7odies 4resent in t:e s4eci8en to 7ind1 and t:ese
7o5nd anti7odies are s57seB5ently detected 7y enJy8e la7elled anti3
:58an i885noglo75lin and s4eci6ic s57strate syste8. I6 t:e test
s4eci8en contained anti7odies colo5r reaction <ill ta=e 4lace.
Indirect ',ISA is t:e 8ost co88only 5sed syste8.
General 5roced6re
ig. 6.# gi9es t:e diagra88atic re4resentation o6 indirect ',ISA.
T:e instr5ctions gi9en in t:e 4ac=age insert 4ro9ided in t:e =it are read
care65lly and 6ollo<ed to t:e letter. All controls as s4eci6ied 85st 7e
incl5ded <it: eac: test r5n to 9alidate t:e test res5lt.
3 Dil5te s4eci8en a44ro4riately1 add to t:e solid 4:ase and inc57ate 6or a
s4eci6ied ti8e and at s4eci6ied te84erat5re.
3 Solid 4:ase is <as:ed to re8o9e 5n7o5nd anti7odies
3 A44ro4riately dil5ted enJy8e conj5gate is added and inc57ated as
s4eci6ied
3 Solid 4:ase is <as:ed to re8o9e e!cess conj5gate
3 S57strate 8ade a44ro4riately is added
3 Colo5r c:ange 4rod5ced is 8eas5red a6ter s4eci6ied ti8e 5sing an
',ISA (eader at <a9elengt: s4eci6ied
3 T:e res5lt is calc5lated as detailed in t:e 4ac=age insert 6ro8 t:e
9ario5s OD 9al5es o7tained.
T:e indirect ',ISA 4rod5ces a colo5r c:ange directly 4ro4ortional to t:e
concentration o6 s4eci6ic anti7odies in t:e s4eci8en.
?)A)")" Com5etitive ELISA
In t:is assay t:e +I&3anti7odies 4resent in t:e s4eci8en co84ete <it:
t:e enJy8e conj5gated anti7odies in t:e reagent 6or 7inding to t:e
antigen on t:e solid 4:ase. I6 t:e test s4eci8en contains +I&
anti7odies1 t:ese <ill co84ete <it: t:e la7elled anti7odies in t:e
reagent 6or 7inding to antigen. So t:at less or no la7elled anti7odies
can attac: to t:e solid 4:ase. +ence1 6aint or no colo5r is 4rod5ced on
addition o6 s57strate i6 s4eci8en contained +I& anti7odies.
56
Cnli=e indirect ',ISA1 :ere red5ction and or a7sence o6 colo5r indicates
t:e 4resence o6 +I& anti7odies in t:e test s4eci8en. De9elo48ent o6
strong colo5r 8eans s4eci8en is non reacti9e 6or +I& anti7odies @ig.
6.3A.
1roced6re ;
ollo< t:e instr5ctions gi9en in t:e 4ac=age insert.
3 A dil5tion o6 test s4eci8en and t:e a44ro4riate a8o5nt o6 enJy8e
la7elled +I& anti7ody conj5gate are 8i!ed and added si85ltaneo5sly
to t:e solid 4:ase antigen s544ort. Inc57ated 6or s4eci6ied ti8e at
s4eci6ied te84erat5re.
3 Plate is <as:ed and s57strate is added.
3 Colo5r c:ange de9elo4ed is 8eas5red 7y ',ISA reader and
e!4ressed as OD 9al5e.
Co84etiti9e ',ISA ta=es less ti8e t:an indirect ',ISA and no
4redil5tion o6 test s4eci8en is reB5ired.
?)A)")< Anti#en and anti=od+ !and.ic ELISA
T:is is a 8odi6ication o6 indirect ',ISA to i84ro9e sensiti9ity and
s4eci6icity o6 t:e test. Antigen 7o5nd to t:e solid 4:ase 7inds
anti7ody in t:e test s4eci8en in 6irst ste4. Since anti7ody 8olec5les
are 7i9alent t:ey are still a7le to 7ind to anot:er 8olec5le. T:e ne!t
ste4 is addition o6 si8ilar enJy8e la7elled +I& antigen i.e. sa8e
antigen as on solid 4:ase. T:is <ill attac: to t:e anti7ody 8olec5le
<:ic: is already 7o5nd to t:e solid 4:ase antigen <it: one ar8 @ig.
6."A. T:5s 6or8s a sand<ic: o6 antigen R anti7ody R enJy8e
la7elled antigen co84le!. T:e ne!t ste4 is addition o6 s4eci6ic
s57strate <:ic: res5lts in de9elo48ent o6 colo5r <:ic: is 8eas5red
7y ',ISA reader. One 7ig ad9antage o6 sand<ic: ',ISA is t:at all
classes o6 +I&3anti7odies can 7e detected.
T:ere is also anti7ody sand<ic: ',ISA <:ic: is done to detect 4#"
antigen @descri7ed in C:a4ter 1#A.
57
1roced6re ;
Sa8e as indirect ',ISA only di66erence 7eing t:at in t:is case enJy8e
la7elled antigen is added in 4lace o6 enJy8e la7elled anti3:58an
i885noglo75lins.
?)A)")( Anti#en and anti=od+ ca5t6re ELISA
Antigen ca4t5re ',ISA can 7e 7ased on 4rinci4le o6 indirect or
co84etiti9e ',ISA1 only di66erence 7eing in t:e initial ste4 o6
attac:ing antigen to t:e solid 4:ase.
A 8onoclonal anti7ody directed against an +I& antigen is 7o5nd to
t:e solid s544ort. Ne!t ste4 is addition o6 +I& antigen s544lied as
reagent. T:is antigen is ca4t5red 7y t:e 8onoclonal anti7ody 7o5nd
to t:e solid 4:ase. Test s4eci8en a44ro4riately dil5ted is added
ne!t. +I& anti7odies i6 4resent in t:e s4eci8en 7ind to +I& antigen
on solid s544ort. (e8aining 4rinci4le is sa8e as indirect ',ISA @ig.
6.$ - 6.6A.
Only ad9antage o6 antigen ca4t5re ',ISA is t:at it is 8ore s4eci6ic
t:an indirect assay.
Anti7ody ca4t5re assays <ere de9elo4ed to test s4eci8ens <it: lo<
concentration o6 +I& anti7odies @e.g. 5rine and sali9aA or to detect
s4eci6ic class o6 anti7odies @e.g. Ig%1 Ig) or IgAA. In t:is test an
anti3:58an i885noglo75lin @anti3Ig%1 Ig) or IgAA is attac:ed to
solid s544ort. T:e 4atient s4eci8en is added. T:e concentrated
i885noglo75lin in 4atient0s s4eci8en 7inds to t:e anti3glo75lins on
solid 4:ase. Ne!t la7elled antigen is added <:ic: 7inds to +I&
anti7odies o6 t:e 4atient 7o5nd to solid s544ort. Ne!t1 t:e s57strate
is added and t:e OD 9al5e is read on t:e ',ISA (eader.
?)A)< Inter5retation of re!6lt!
'ac: test r5n 85st 7e 9alidated according to t:e 9alidation criteria
gi9en in t:e 4ac=age insert o6 t:e =it.
C5t3o6639al5e
3 'ac: =it 8an56act5rer :as de9ised a 8et:od o6 calc5lation t:at
4rod5ces a c5t o66 9al5e1 to classi6y a test sa84le as 4ositi9e or
negati9e.
3 T:is c5t3o66 9al5e can 7e 7ased on an a9erage o6 t:e negati9e
controls 85lti4lied 7y a 6actor or is 7ased on a relations:i4 o6 4ositi9e
controls1 to o4ti8iJe sensiti9ity and s4eci6icity o6 assay.
%rey Jone identi6ication
3 All in6ected indi9id5als start <it: lo< le9els o6 anti7ody and grad5ally
t:e le9el o6 +I& anti7ody increases as t:e in6ection 4rogresses.
T:ere6ore1 so8e la7oratories testing :ig: ris=1 :ig: incidence
4o45lations instit5te >grey Jone? identi6ication <:ere s4eci8ens
<:ic: are j5st @10QA 7elo< t:e c5to66 @i.e.1 c5t3o66 85lti4lied 7y 0.DA
are classi6ied as >grey Jone reacti9e?1 <:ic: are retested and
s57jected to con6i8atory testing1 li=e re4eatedly reacti9e sa84les.
T:is 8et:od 8ay identi6y serocon9erting 4atients early in t:e co5rse
o6 disease.
58
*e5eat te!tin#
3 (eacti9e and >grey Jone? reacti9e s4eci8ens are nor8ally re4eated
in d54licate in ',ISA and consens5s res5lts @# or 3 test res5lts A are
acce4ted.
?)A)( 1erformance varia=le!8tro6=le!ootin#
',ISA tests are generally easy to 4er6or8 75t reB5ire care65l
ad:erence to 4roced5resF any de9iation in inc57ation ti8es and ; or
te84erat5re and 4i4etted 9ol58e can dra8atically c:ange test
res5lts @see C:a4ter 10 alsoA.
?)A)()% S5ecimen 5ro=lem! $ tin#! to avoid
3 +ae8olysis
3 (e4eated 6reeJing and t:a<ing
3 %ross li4ae8ia
3 2acterial conta8ination
3 +eat inacti9ation 3 8ay ca5se 6alsely reacti9e ',ISA
?)A)()" O5erator8tecnical error
3 I84ro4er ti8ing o6 inc57ations can gi9e
3 lo< or :ig: a7sor7ance readings
3 Inc57ation te84erat5re
3 ,o< te84erat5re 3 lo< a7sor7ance readings
3 +ig: te84erat5re 3 :ig: a7sor7ance readings
3 I6 conj5gate 4reci4itates lo< reading: i6 :eat inacti9ates enJy8es
lo< reading
?)A)()< EC6i5ment 5ro=lem!
3 Inc57ator
Poor air circ5lation @:ot s4ots1 cold s4otsA
Stac=ing 4lates @O#AF i84ro4er :eating
3 .ater 2at:
Conta8inated <ater1 s4las:es into 4lates or t57es
3 I84ro4erly 8aintained 4late <as:er
I84ro4er <as:ing1 :ig: O.D. 7ac=gro5nd1 le6t o9er conj5gate on
4lates creates colo5r c:ange.
3 I84ro4erly cali7rated;8aintained 8icro4late reader
?)A)()( O5erator tecniC6e ; &ariation <it:in or 7et<een 4lates
3 I84ro4er <as:ing
@aA Inco84lete re8o9al o6 <as: sol5tions 3 lo< a7sor7ance
readings.
@7A InadeB5ate <as:ing 3 :ig: a7sor7ance readings non3s4eci6ic
reactions.
3 Inacc5rate;incorrect 4i4etting 3 a7sor7ance readings1 :ig: or lo<.
3 ail5re to add sa84le controls1 or reagents to correct <ells.
3 Ti8ing o6 reagent additions1 es4ecially sto4 sol5tion in relation to
s57strate.
3 (eagents reconstit5ted or dil5ted in <rong dil5ent or <rong 9ol58es.
59
?)A)' Fal!e 5o!itive and fal!e ne#ative ELISA re!6lt! ; T:ere are so8e
conditions ot:er t:an +I& in6ection <:ic: 8ay gi9e a reacti9e +I&
res5lts i.e. 6alse 4ositi9e res5lt. Also1 so8eti8es t:e res5lt 8ay 7e
negati9e e9en in +I& in6ected i.e. 6alse negati9e res5lt. So8e o6 t:e
co88on conditions gi9ing erroneo5s res5lts are listed 7elo<.
3 A5to3i885ne diseases
3 )5lti4le 4regnancies
3 )5lti4le trans65sions
3 Anti7ody to Class II +,A Ag @+,A3D("A
3 +y4er ga88aglo75line8ia
3 Anti4olystyrene anti7odies
3 C:ronic alco:olics
3 Patients <it: :e4atitis
3 +e4atitis 2 i885nisation
3 Tec:nical error etc.
3 Ot:ers
alse negati9e
3 In6ected 75t not yet serocon9erted1 <indo< 4eriod
3 ,ate stage disease @i885ne colla4seA
3 Tec:nical error
?)%& *a5id a!!a+! ; A n587er o6 ra4id assays 7ased on 4rinci4le o6
aggl5tination and ',ISA :a9e 7een de9elo4ed 6or ease o6
4er6or8ance and B5ic= res5lts. T:ese assays generally reB5ire less
t:an 30 8in5tes to 4er6or8 and do not reB5ire s4ecial eB5i48ents.
?)%&)% A##l6tination a!!a+! ; Aggl5tination assays incor4orate a 9ariety o6
antigen coated carriers li=e red cells1 late! 4articles1 gelatin 4articles
and 8icro7eads. T:ese 4articles are 5sed to s544ort or carry t:e
antigen. +I& antigens are attac:ed to t:e carrier 4articles 7y non
s4eci6ic attac:8ent.
Aggl5tination assays :a9e good sensiti9ity1 do not reB5ire
so4:isticated eB5i48ent1 are easy to 4er6or81 reB5ire no <as:
4roced5res and are cost e66ecti9e. +o<e9er1 s4eci6icity is so8e<:at
co84ro8ised and 4roJone reaction 8ay 7e seen. To o9erco8e t:e
4roJone reaction1 dil5ted s4eci8en is 5sed to 4er6or8 t:e test.
D5ring t:e aggl5tination reaction +I& anti7ody co87ines <it: +I&
antigen on t:e carrier 4articles and since all anti7odies are
85lti9alent1 a sort o6 lattice net<or= is 6or8ed <:ic: can 7e
9is5alised 8acrosco4ically or 8icrosco4ically as 4er t:e directions o6
t:e 8an56act5rer.
1roced6re
T:e instr5ctions gi9en in t:e 4ac=age insert s:o5ld 7e strictly
6ollo<ed.
3 A dil5tion o6 test s4eci8en is 8i!ed <it: +I& antigen containing
carrier 4articles.
3 I6 test s4eci8en contains +I& anti7odies1 a lattice net<or= <ill 6or8
7et<een t:e antigen carrying 4articles and anti7odies and is
9is5aliJed as 6or8ation o6 cl584s @ig. 6.7A.
3 Aggl5tination is seen 9is5ally e!ce4t <:en carrier 4articles are
8icro7eads.
3 .ea= reactions read s57jecti9ely 85st 7e con6ir8ed 7y ',ISA.
?)%&)" Dot =lot a!!a+! 8 Com= te!t!
60
T:ese assays are ra4id1 easy to 4er6or81 can 5s5ally discri8inate
7et<een +I&31 and +I&3# and do not reB5ire so4:isticated
eB5i48ent. T:e res5lts are read 7y de9elo48ent o6 colo5r. Only
dra<7ac= o6 t:e test is t:e :ig: cost. Sensiti9ity and s4eci6icity o6
t:ese assays co84ares <it: ',ISA.
T:e assays 5tiliJe reco87inant or synt:etic 4e4tides s4otted on to
nitrocell5lose 4a4er;8icro 4articles. T:e antigen containing 8atri!
is :o5sed in a 4lastic de9ice @cassetteA containing adsor7ent 4ads
5nderneat: to collect reactants or 8ade as a co87 and t:e antigens
are s4otted onto t:e toot: o6 t:e co87 card @C:a4ter 7A. 'ac: assay
contains an i885noglo75lin ca4t5re control to 9alidate t:e res5lt.
T:ese assays are 9ery good 6or single test a44lication i.e. in
e8ergency1 a5to4sy roo8 and 4eri4:eral 7lood 7an=s. T:ey :a9e
not 7een a5to8ated so 6ar.
1roced6re 2com= and dot =lot te!t!4
T:e instr5ction gi9en in t:e 4ac=age insert s:o5ld 7e 6ollo<ed.
3 Dil5tion o6 test s4eci8en is added <:ic: 6ilters t:ro5g: t:e
8e87rane;8icro7eads 7y action o6 gra9ity. D5ring t:e 4rocess +I&
anti7odies in t:e test s4eci8en 7ind to +I& antigens on t:e
8e87rane.
3 .as: sol5tion is added to re8o9e 5n7o5nd anti7odies.
3 T:e a44ro4riately dil5ted conj5gate is added @anti:58an 3
i885noglo75lin conj5gated <it: enJy8e as in indirect ',ISA or
4rotein A conj5gated <it: colloidal gold 4articles. Protein A
conj5gate can 7ind to :58an Ig%s directlyA.
3 (inse <it: <as: sol5tion to re8o9e e!cess o6 reactants.
3 S57strate a44ro4riately dil5ted is added <:ic: 6or8s a 4reci4itate at
sites on 8e87rane <:ic: contain antigen anti7ody co84le!es.
3 Colo5r de9elo48ent in t:e control and test dots is read 9is5ally.
ig. 6.E - 6.D s:o< diagra88atic re4resentation o6 co87 and dot 7lot
assays
Intre4retation o6 res5lt is done as 4er t:e instr5ctions in t:e 4ac=age
insert. +o<e9er1 reacti9e res5lts :a9e to 7e con6ir8ed 7y ',ISA.
I885noco87 test is si8ilar to 7lot assay 7ased on 4rinci4le o6
indirect ',ISA. +ere also t:e instr5ctions gi9en in t:e 4ac=age insert
<it: t:e =it are 6ollo<ed to 4er6or8 t:e test.
?)%% HI0 anti=od+ detection in oter fl6id!.
T:e standard s4eci8ens 6or detection o6 +I& anti7odies are
ser58;4las8a ; 7lood. +o<e9er1 detecting +I& anti7odies in ot:er
6l5ids is also 7eing de9elo4ed. +I& anti7odies can 7e detected in oral
6l5ids @sali9a;oral 85cosal trans5datesA. T:e le9els are less t:an 1Q
o6 t:e le9els in ser581 75t can 7e detected 7y sensiti9e ',ISA
tec:nology. T:e isoty4e o6 anti7ody detected is secretory IgA1 so
a44ro4riate ',ISA s:o5ld 7e 5sed. T:e tests :a9e s:o<n :ig:
sensiti9ity and s4eci6icity as 4er t:e re4orts 6ro8 C.S.1 TanJania1
T:ailand1 2raJil and ot:er co5ntries. T:ese tests 8ay 6ind 7etter 5se
once t:e iss5es concerning con6identiality1 co5nselling and 6ollo< 54
:a9e 7een resol9ed. +I& anti7odies can also 7e detected in 5rine
5sing a44ro4riate ',ISA =its. T:ese tests :a9e also 7een 9alidated.
61
+o<e9er1 g5idelines 6or 5sing t:ese tests as <ell as strategies 6or
6ollo< 54 etc. need to 7e de9elo4ed.
+58an Ig
+I&31
+I&3#

1o!itive
control
*eactive for
=ot HI0$%
and HI0$"
Ne#ative
control
*eactive
for HI0$%
*eactive
for HI0$"
Te!t not valid
2H6man I# !5ot not
!een4
Fi#) ?)@ Dia#rammatic re5re!entation com=
te!t 2Toot of com= card !o.n4
)lasti de"ie
HIV A0 dot HIVA= dot
Control dot 8itroellulose
su%%ort
&'olid %hase*
F-2 F.4 A ,)1/-'(-)' HIV;$&HIV;. *)(;/0)(
ss9 () *-33"r"'(-(" -'3",(-)'
?)%" Coice of te !creenin# a!!a+
2e6ore c:oosing a 4artic5lar 4rotocol;=it;strategy 6or +I& testing
one :as to 7e clear a7o5t t:e 6ollo<ing 4ara8eters 7e6ore testing is
5nderta=en.
3 O7jecti9es o6 testing
3 Sensiti9ity
3 S4eci6icity
3 Pre9alence o6 +I& in6ection in t:e 4o45lation.
3 Cost e66ecti9eness.
3 A44ro4riateness to t:e strategy and national g5idelines o6 testing
and according to t:e in6rastr5ct5re 6acilities a9aila7le @C:a4ter DA.
?)%")% O=>ective! of HI0 te!tin#
iA 2lood and 7lood 4rod5cts sa6ety. T:is is ac:ie9ed 7y 8andatory
testing o6 all donated 7lood 5nits and 7lood 4rod5cts.
iiA Screening o6 donors o6 s4er8s1 organs and tiss5es.
62
iiiA Diagnosis o6 +I& in6ection in clinically s5s4ected cases.
i9A &ol5ntary testing a6ter co5nselling.
9A '4ide8iological s5r9eillance 5sing 5nlin=ed anony8o5s +I& testing.
+ere t:e res5lt o6 test cannot 7e lin=ed <it: t:e identity o6 t:e
4erson.
9iA (esearc:.
?)%")" Sen!itivit+ of HI0 te!t
It is t:e acc5racy <it: <:ic: a test can con6ir8 t:e 4resence o6 an
in6ection. Tests <it: :ig: sensiti9ity s:o< 6e< 6alse negati9es and
are 8eant to 7e 5sed to screen 7lood 4rior to trans65sion and ens5re
7lood sa6ety.
Tr5e 4ositi9e @TPA
Sensiti9ity U S100
TP R alse negati9e @NA
?)%")< S5ecificit+ of HI0 te!t
It is t:e acc5racy <it: <:ic: a test can con6ir8 t:e a7sence o6 an
in6ection. Tests <it: :ig: s4eci6icity s:o< 6e< 6alse 4ositi9es and are
to 7e 4re6erred 6or t:e diagnosis o6 +I& in6ection in an indi9id5al.
TN @Tr5e negati9eA
S4eci6icity U S100
TNRP @alse 4ositi9eA
?)%")( 1revalence of HI0 and it! im5lication for te!t re!6lt!
T:e 4ro7a7ility t:at a test <ill acc5rately deter8ine t:e tr5e in6ection
stat5s o6 a 4erson 7eing tested 9aries <it: t:e 4re9alence o6 +I&
in6ection in t:e 4o45lation 6ro8 <:ic: t:e 4erson co8es. T:e :ig:er
t:e 4re9alence1 greater is t:e 4ro7a7ility t:at a 4erson testing
4ositi9e is tr5ly in6ected i.e. greater t:e 4ositi9e 4redicti9e 9al5e o6
t:e test @PP&A.
T:e li=eli:ood t:at a 4erson s:o<ing a negati9e res5lt is tr5ly
5nin6ected t:e negati9e 4redicti9e 9al5e @NP&A decreases as t:e
4re9alence o6 +I& in6ection a8ong t:e general 4o45lation increases.
T:e details 6or calc5lation o6 PP& and NP& are gi9en in C:a4ter 11.
63
UNAIDS AND -HO recommendation! for HI0 te!tin# !trate#ie! accordin# to
te!t o=>ective and 5revalence of infection in te !am5le 5o56lation
O7jecti9e o6 testing Pre9alences o6
in6ection
Testing
strategy
Trans65sion;trans4lant
sa6ety
All 4re9alences I
S5r9eillance O10Q I
P10Q II
Diagnosis : Clinical
signs; sy84to8s
o6 +I&
in6ection
O30Q
P30Q
I
II
Asy84to8atic O10Q
P10Q
II
III
?)%")' Strate#ie! of HI0 te!tin# in India
2eca5se o6 t:e enor8o5s ris= in9ol9ed in trans8ission o6 +I&
t:ro5g: 7lood1 sa6ety o6 7lood and 7lood 4rod5cts is o6 4ara8o5nt
i84ortance. Since t:e PP& is lo< in 4o45lations <it: lo< +I&
4re9alence1 .+O;%OI :a9e e9ol9ed strategies to detect +I& in6ection in
di66erent 4o45lation gro54s and to 65l6ill di66erent o7jecti9es @Anne!5re
6.1A. T:e 9ario5s strategies1 so designated1 in9ol9e t:e 5se o6
categories o6 tests in 9ario5s 4er85tations and co87inations.
1. ',ISA;Si84le;(a4id tests @';(;SA 5sed in strategy I1 II - III
#. S544le8ental test li=e .estern 2lot and ,ine I885noassay are
5sed in 4ro7le8 cases e.g. in cases o6 indeter8inate;
discordant res5lt o6 ';(;S.
Strate#+ I ; Ser58 is s57jected once to ';(;S 6or +I&. I6 negati9e1 t:e
ser58 is to 7e considered 6ree o6 +I& and i6 4ositi9e1 t:e sa84le is
ta=en as +I& in6ected 6or all 4ractical 45r4oses. T:is strategy is 5sed
6or ens5ring donation sa6ety @7lood;7lood 4rod5cts organ1 tiss5es1
s4er8s etc.A. T:e 5nit o6 7lood testing reacti9e @4ositi9eA is discarded.
Donor is not in6or8ed.
Strate#+ II ; A ser58 sa84le is considered negati9e 6or +I& i6 t:e 6irst
',ISA re4ort is so1 75t i6 reacti9e1 it is s57jected to a second ',ISA
<:ic: 5tiliJes a syste8 di66erent 6ro8 t:e 6irst one. It is re4orted
reacti9e only i6 t:e second ',ISA con6ir8s t:e re4ort o6 t:e 6irst. T:is
strategy is 5sed 6or s5r9eillance and 6or diagnosis only i6 so8e AIDS
indicator disease is 4resent.
Strate#+ III ; It is si8ilar to strategy II1 <it: t:e added con6ir8ation o6 a
t:ird reacti9e ',ISA test 7eing reB5ired 6or a sa84le to 7e re4orted +I&
4ositi9e. T:e test to 7e 5tiliJed 6or t:e 6irst ',ISA is one <it: t:e
:ig:est sensiti9ity and 6or t:e second and t:ird ',ISAs1 tests <it: t:e
:ig:est s4eci6icity are to 7e 5sed.
Strategy II - III are to 7e 5sed 6or diagnosis o6 +I& in6ection. ',ISA #
and ',ISA 3 o5g:t to 7e tests <it: t:e :ig:est PP& 4ossi7le to
eli8inate any c:ances o6 6alse 4ositi9e res5lts. Strategy III is 5sed to
64
diagnose +I& in6ection in asy84to8atic indi9id5als ind5lging in :ig:
ris= 7e:a9io5r.
?)%< S655lemental 2confirmator+4 a!!a+!
S544le8ental;con6ir8atory test are 5nderta=en to con6ir8 t:e
+I& in6ection stat5s o6 an indi9id5al <:o is asy84to8atic and :as
:istory o6 :ig: ris= 7e:a9io5r.
S544le8ental assays are 5sed 6or 45r4ose o6 diagnosis o6 +I&
in6ection and identi6ication o6 t:e indi9id5al i.e. t:e indi9id5al is
in6or8ed a7o5t t:e test res5lts in t:e 4ro4er <ay @co5nselling and
con6identialityA. T:is is 7eca5se t:e screening assays are :ig:ly
sensiti9e and are 7o5nd to gi9e 7iologic 6alse 4ositi9e reactions
<:ereas s544le8ental assays are sensiti9e as <ell as s4eci6ic and are
4er6or8ed to r5le o5t 6alse 4ositi9e reactions. S544le8ental ';(;S
5sing di66erent antigen and;or 4rinci4le o6 test are 4er6or8ed to 65l6il
t:e strategy II and III o6 +I& testing 4olicy.
+o<e9er1 s544le8ental tests do not al<ays gi9e concl5si9e
res5lts in <:ic: case eit:er ot:er tests @PC(1 4#" antigen detection
etc.A :a9e to 7e resorted to or 6ollo< 54 st5dy is 4ractised.
0ario6! !655lemental a!!a+! availa=le commerciall+ are li!ted =elo. ;
3 ',ISA <it: di66erent antigen syste8 @reco87inant or synt:etic
4e4tidesA or <it: di66erent 4rinci4le o6 test <:ic: 8a=es t:e test
8ore s4eci6icF
3 .estern 7lot @.2AF
3 I885no7lot @I2AF
3 ,ine i885noassayF
3 Indirect 6l5orescent anti7ody test @IAAF
3 (adioi885no4reci4itation test @(IPAA.
As 4er t:e National +I& testing 4olicy o6 India1 s544le8ental
tests are 5sed only 6or strategy II and strategy III o6 +I& testing. or
t:e 45r4ose1 ';(;S <it: :ig:er s4eci6icity can 7e 5sed as s544le8ental
tests1 4ro9ided t:ese assays incor4orate di66erent antigen syste8
and;or di66erent 4rinci4le o6 test. Only in case o6
5neB5i9ocal;discordant res5lt .2 is to 7e 5sed. T:is is to 8a=e t:e
+I& testing cost e66ecti9e as <ell as easy to 4er6or8. T:e ot:er
s544le8ental tests i.e. IA1 (IPA are :ardly e9er 5sed in India. So1
only t:e s544le8ental assays 5sed in India are 7eing descri7ed.
?)%<)% E8*8S te!t! a! !655lemental a!!a+
S544le8ental ';(;S are reco88ended 6or s4eci8en reacti9e in
t:e screening assay 6or strategy II and III o6 +I& testing 4olicy. I6 t:e
screening assay contained 9ir5s lysate as antigen t:e second and t:ird
assays s:o5ld :a9e reco87inant or synt:etic 4e4tides as antigens. Cse
o6 reco87inant and synt:etic 4e4tides 8a=es t:e assay 8ore s4eci6ic
<:ile retaining t:e sensiti9ity. Alternati9ely t:e second and t:ird
assays can also 7e 7ased on di66erent 4rinci4le o6 ',ISA e.g. i6 t:e
screening assay <as indirect ',ISA t:e s544le8ental assays s:o5ld 7e
7ased on 4rinci4le o6 co84etiti9e ',ISA;Sand<ic: ',ISA etc.1 <:ic:
again is done to increase t:e s4eci6icity o6 t:e screening assay <:en
5sed as s544le8ental assays.
?)%<)" -e!tern =lot8Imm6no=lot 8 Line imm6noa!!a+
65
T:ese are 8ost <idely acce4ted s544le8ental assays1 are :ig:ly
s4eci6ic 75t are e!4ensi9e1 la7o5r intensi9e1 need e!4ertise to inter4ret
and 8ay gi9e 5neB5i9ocal;indeter8inate res5lts. T:e s4eci6icity o6
t:ese tests is 7ased on t<o 6actors: se4aration o6 antigens and t:eir
concentration.
I2 and .2 5se 9iral antigens 6ro8 <:ole 9ir5s lysates
electro4:oretically trans6erred to a 8e87rane s544ort. So t:ese 7lots
8ay contain conta8inating cell5lar co84onents.
(eco87inant or synt:etic +I& antigens 8ec:anically a44lied onto
t:e s544ort 8e87rane are 5sed in ,ine I885noassays @,IAA. T:ese do
not contain conta8inating cell5lar co84onents and are :ig:ly s4eci6ic.
Indeter8inate res5lts can 7e seen 7y any o6 t:e a7o9e assays.
.2;,IA +I& =its are co88ercially a9aila7le.
?)%<)")% 1roced6re
T:e instr5ctions gi9en in t:e 4ac=age insert 85st 7e diligently
6ollo<ed 7ot: 6or 4er6or8ance o6 test as <ell as inter4retation o6 res5lt.
%eneral Proced5re:
3 +I& Ag test stri4s are 4laced in indi9id5al tro5g:s o6 4lastic tray.
3 A dil5tion o6 t:e 4atient ser58;4las8a or control is added to
indi9id5al test tro5g:s and inc57ated on a roc=ing 4lat6or8.
3 At t:e end o6 inc57ation1 t:e tro5g:s are as4irated and t:e test
stri4s are <as:ed to re8o9e 5n7o5nd anti7odies.
3 An enJy8e conj5gate anti:58an i885noglo75lin is added to
7ind to :58an anti7odies 7o5nd to +I& antigens on t:e test stri4s.
3 T:e tro5g:s are as4irated and stri4s are <as:ed to re8o9e
5n7o5nd conj5gated i885noglo75lins.
3 A s57strate is added and colo5r de9elo48ent occ5rs <:ere
antigen;anti7ody co84le!es are localiJed on t:e stri4s.
3 T:e colo5r de9elo48ent is sto44edF t:e stri4s are <as:ed1 dried
and inter4reted.
ig. 6.10 gi9es diagra88atic re4resentation o6 .2 test.
?)%<)")") *eadin# and Inter5retation
T:e 4resence or a7sence o6 7ands o6 t:e 8ajor 9iral 4roteins is
scored 6or eac: stri4. Cn5s5al or aty4ical 7and locations are also
noted.
Inter5retative Criteria
(es5lts are inter4reted eit:er as 4er .+O criteria i.e. 4resence o6
at least t<o en9elo4e 7ands @g4 1#01 g41601 g4"1 etc.A or as 4er t:e
8an56act5rer0s criteria 6or 4ositi9e1 indeter8inate and negati9e.
66
In case o6 indeter8inate res5lt1 retesting o6 t:e indi9id5al is done
a6ter t<o <ee=s1 1 8ont:1 3 8ont:s and so8eti8es 54to 1 year.
D5ring t:is ti8e eit:er t:e test 7eco8es negati9e or 4ositi9e or 8ay
stay indeter8inate. Decision a7o5t t:e res5lt :as to 7e reac:ed a6ter
correlation <it: :istory o6 :ig: ris= 7e:a9io5r and clinical 4ara8eters.
?)%( La=orator+ dia#no!i! of HI0 infection in ne.=orn 2Con#enital HI0 InfectionA
Trans4lacental trans8ission o6 +I& can occ5r 6ro8 in6ected 4regnant
8ot:er to t:e 6oet5s as early as E <ee=s o6 gestation or 8ay 7e e9en
earlier. It is esti8ated t:at O E0Q o6 AIDS cases in in6ants P 1 year
old are d5e to 4erinatal trans8ission o6 +I&31. Diagnosis o6 +I&
in6ection in in6ants 7orn to sero4ositi9e 8ot:ers is di66ic5lt 7eca5se
8aternal anti7ody @Ig%A to +I&31 crosses t:e 4lacenta and can 4ersist
6or 54to 1$ 8ont:s 8a=ing t:e distinction 7et<een 8aternal and
neonatal Ig% di66ic5lt. T:e tests <:ic: can 7e 5nderta=en to diagnose
+I& in6ection in neonates 7e6ore 1$ 8ont:s o6 age are detailed 7elo<.
?)%()% Detection of I#A and8or I#M anti$HI0 anti=odie!) T:ese anti7odies do not
cross 4lacenta. T:e IgA class o6 +I& anti7ody assay 5sing .estern 7lot
tec:niB5e in in6ected c:ildren at 3 8ont:s o6 age :as a sensiti9ity o6
D7.6Q and s4eci6icity o6 DD.7Q as re4orted in a st5dy. Ig) class o6
anti7odies are 4rod5ced 7y in6ected in6ants 7y si! 8ont:s o6 age.
Prod5ction o6 Ig) is erratic1 6alse 4ositi9e res5lts are o7tained d5e to
r:e58atoid 6actor and 4oly9alent nat5re o6 Ig) leading to nons4eci6ic
7inding.
?)%()" E!timation of 5"( anti#en 2core anti#en4) T:e i885ne3co84le! dissociation
assays <:ic: in9ol9e 4retreat8ent o6 ser58;4las8a to li7erate 4#"
antigen co84le!ed <it: 4#" anti7ody 4rior to 4er6or8ance o6 ',ISA are
B5ite sensiti9e in identi6ying +I& 3in6ected in6ants.
?)%()< 1ol+mera!e cain reaction 21C*4) T:e tec:niB5e s4eci6ically a84li6ies 9iral
DNA seB5ences o6 interest. T:eoretically it is 4ossi7le to identi6y one
in6ected cell in t:e s4eci8en as also latent +I& in6ections. &ario5s
re4orts indicate t:e s4eci6icity o6 PC( to 7e in9aria7ly OD$Q regardless
o6 age o6 testing <:ile sensiti9ity ranges 6ro8 1$Q in neonates @<it:in
"E :rs. a6ter 7irt:A to 8ore t:an D$Q in in6ants o9er 1 8ont: o6 age.
?)%()( In$vitro anti=od+ 5rod6ction a!!a+) T:e 8et:od is tedio5s and in9ol9es in3
9itro c5lt5re o6 anti7ody 4rod5cing 2 ly84:ocytes 6ro8 4eri4:eral
7lood. T:e +I& anti7odies are detected in c5lt5re s54ernate 6ro8 tr5ly
in6ected in6ants.
?)%()' In$vitro i!olation of vir6! from =lood or ti!!6e!) T:e 8et:od is ti8e
cons58ing1 e!4ensi9e and not sensiti9e. T:is is d5e to t:e lo<er
n587er o6 a9aila7le cells c5lt5red and 4a5city o6 in6ected cells in t:e
sa84le t:at are 4resent. Additionally1 it 8ay ta=e 54to 6 <ee=s or
longer to o7tain a res5lt. &ery 6e< la7oratories in India are eB5i44ed to
c5lt5re +I&.
?)%()? Indirect indicator! of HI0 infection) T:ese incl5de 9ario5s nons4eci6ic
8ar=ers li=e :y4erga88aglo75line8ia1 a7sol5te ly84:o4enia1
:e8atologic a7nor8alities1 lo< CD" cell co5nts and clinical 8ar=ers li=e
se9ere 6ail5re to t:ri9e or <asting syndro8e1 rec5rrent se9ere 7acterial
in6ections1 o44ort5nistic in6ections1 +I& associated 8alignant disease1
ence4:alo4at:y1 ly84:ocytic interstitial 4ne58onia1 etc. <:ic: 4oint
to<ards acB5ired i885node6iciency syndro8e. +o<e9er1 t:ese
67
4ara8eters are e84loyed to s544le8ent t:e con9entional
serodiagnosis o6 +I& in6ection. T:e tests a9aila7le 6or diagnosis o6
9ertical trans8ission o6 +I& 6ro8 in6ected 8ot:er to t:e neonate err on
t:e side o6 :ig: s4eci6icity <it: a lo<er sensiti9ity.
?)%' La=orator+ dia#no!i! d6rin# -indo. 1eriod ; +I& in6ection d5ring <indo<
4eriod can 7e detected 7y de8onstrating t:e 4resence o6 9ir5s and
9ir5s co84onents.
3 PC( @4roced5re details gi9en in
3 4#" antigen assay @P"0QA C:a4ter E and C:a4ter 1#A
3 &iral C5lt5re
Need o6 la7oratory diagnosis in <indo< 4:ase
3 ollo<ing 5ntested 7lood trans65sion.
3 (is=y :eterose!5al;:o8ose!5al e!4os5re.
3 Needle stic= inj5ry @conta8inatedA
AnneH6re ?)" A 8odel la7oratory
4ro6or8a 6or 4er6or8ing ',ISA
AnneH6re ?)< Pro6or8a @in6or8ationA
6or reB5isitioning +I& testing.
68
Ch6("r F ; A''"A+r" .
A 1)*"0 0/)r()r9 6r)3)r1 3)r 6"r3)r1-'2 ELISA
0$ 6eferred by =$ 'a#%le nu#ber
@$ 8a#e of the .it : E$ Lot Q :
G$ Date of e/%irty :
?$ Date of test run : H$ Tehniian :
1$ 6esults : C<TA(-- V2L<; R
INSTRUCTIONS : -ill in sa#%le nu#ber and results into a%%ro%riate s%ae
0 = @ E G ? H 1 B 0> 00 0=
2
B
C
D
;
-
3
H
Co##ents :
SSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSS
SSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSS
SSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSS
(ffier Inharge &Laboratory*
69
7) DIFFE*ENTIAL FEATU*ES IN E1IDEMIOLOG:, NATU*AL HISTO*:, 0I*OLOG:,
1ATHOGENESIS AND LABO*ATO*: TESTING OF HI0$" INFECTION)
7)% E5idemiolo#+ ; +I&3# <as 6irst disco9ered in Senegal1 .est A6rica in
1DE$. D5ring t:e 4ast decade signi6icant +I&3# in6ection :as 7een
re4orted 6ro8 .est A6rica1 Port5gal1 )oJa87iB5e1 Angola1 C57a1 2raJil.
I84ortant co5ntries o6 .est A6rica na8ely %5inea32issa51
25r=ina6asso1 %:ana1 I9ory Coast1 Nigeria1 )ali re4orted :ig:
4re9alence o6 +I&3# in6ection. +I&3# :as also 7een re4orted 6ro8
'5ro4e1 C.S.A. and Canada. T:e 6irst doc58entation o6 +I&3# in India
<as 8ade 6ro8 2o87ay in 1DD0. T:ere are re4orts o6 +I&3# in6ection
6ro8 )a:aras:tra1 Del:i1 C:ennai1 &isa=:a4attana81 Cttar Prades:1
2i:ar1 P5nja7 and )ani45r.
7)" 0irolo#+ ; +I&3# is si8ilar to +I&31 7iologically and :as si8ilar :ost
range1 8or4:ology and genetic organisation. Co84arison o6 8ajor
gene 4rod5cts o6 +I&31 and +I&3# is gi9en 7elo<:
%ene +I&31 +I&3#
gag
Prec5rsor 4$$ 4$6
Core 4#" 4#6
)atri! 417 416
4D
47
en9
Prec5rsor g4160 g41"0
'!ternal g41#0 g410$;1#$
Trans8e87rane g4"1 g436;"1
4ol
(e9erse
Transcri4tase
466 46E
(e9erse
Transcri4tase
4$1 4$3
'ndon5clease 431 43"
3 Antigenically1 8ost cross reactions in +I& serological assay are d5e to
+I& anti7odies directed against gag and 4ol antigens1 res5lting 6ro8
t:e e!tensi9e n5cleotide :o8ology o6 t:ese str5ct5ral genes 7et<een
+I&31 and +I&3#.
3 en9 cross reactions are less co88on.
3 One o6 t:e reg5latory co84onents 94! @416A is 4resent in +I&3# and
not in +I&31 and :ence 5niB5e gene 6or +I&3#. It 8ay 7e res4onsi7le
6or t:e di66erent 4at:ologic e66ects noted <it: +I&3# in6ections. T:is
8ay :a9e i84ortant diagnostic a44lication since detection o6 anti 94!
<ill identi6y +I&3# in6ection es4ecially <:en t:e sera react 7y 7ot:
assays. Anti7odies to t:e ne6 reg5latory 4rotein <:ic: 8ay not
65nction as do<n reg5lator in +I&3# :a9e 7een detected in #$Q o6 +I&3
# in6ected indi9id5als.
7)< Nat6ral i!tor+ and 5ato#ene!i! ; )ode o6 trans8ission o6 +I&3# is sa8e
as +I&31 75t t:e rate o6 +I&3# trans8ission is signi6icantly lo<er. T:e
8ost co88on ro5tes o6 +I&3# trans8ission are 4erinatal and
:eterose!5al. I885nological and clinical o7ser9ations :a9e s:o<n
t:at t:ere is slo< 4rogression o6 +I&3# disease co84ared to +I&31.
T:e lo<er le9el o6 9iral load 8ay 7e related to t:e lo<er e66iciencies o6
CD" rece4tor 7inding and 8ay 7e an e!4lanation 6or t:e see8ingly
slo<er trans8ission and slo<er disease de9elo48ent in +I&3# in6ection
70
as co84ared to +I&31. So8e isolates o6 +I&3# can in6ect CD" cells 7y
5sing 65sin;CSC(" 8olec5les inde4endent o6 CD" rece4tor.
T:e di66erences in 4at:ogenesis and ot:er c:aracteristics o6 +I&3# as
co84ared to +I&31 in6ections are detailed 7elo< :
3 Slo< clinical deterioration and disease de9elo48entF
3 e<er i885nological dist5r7ancesF
3 ,o< trans8issi7ility F
3 ,onger inc57ation 4eriodF
3 ,o<er rate o6 9ertical trans8issionF
3 Possi7le 4rotection o6 +I&31 in6ectionF
3 ,o< rate o6 e66iciency o6 9ir5s isolationF
3 ,o< rate o6 o44ort5nistic in6ectionsF
3 ,o<er 4las8a t58o5r necrosis 6actor 3F
3 ,o<er 9iral loadF
3 ,o<er le9el o6 4ro9iral DNA in circ5lating ly84:ocytes.
7)( D6al 2HI0$% and HI0$"4 a!!a+! ; T:ese can 7e eit:er d5al screening
assays or d5al con6ir8atory assays.
7)()% D6al !creenin# a!!a+!; D5al screening tests are a9aila7le in t:e 6or8 o6
',ISA and ra4id tests. )ost second generation d5al screening tests
5se t:e indirect 6or8at and do not identi6y <:ic: anti7odies are
4resent @anti +I&31 or +I&3#A. A 4ositi9e reaction only indicates t:at
anti7odies to one or t:e ot:er are 4resent. T:ird generation d5al
assays :a9e 7een de9elo4ed in 7ot: 8icrotitre and 7ead 6or8ats. T:e
d5al screening tests are 5sed 6or screening o6 7lood donors or ot:er
indi9id5als at 5n=no<n ris= 6or +I&31 and;or +I&3# in6ection and 6or
clinical diagnostic testing.
)lasti de"ie
HIV A0 dot HIVA= dot
Control dot 8itroellulose
'u%%ort
&'olid %hase*
F-2 G.$ A ,)1/-'(-)' HIV;$&HIV;. *)(;/0)(
ss9 () *-33"r"'(-(" -'3",(-)'
In ',ISA1 t:e 4rinci4le is sa8e as t:at o6 +I&31 e!ce4t t:at t:e
antigens @reco87inant or synt:eticA o6 +I&31 and +I&3# 7ot: are
coated in t:e sa8e 8icro<ell. (a4id tests li=e dot 7lot co87ination
assays are a9aila7le t:at :a9e se4arate dots containing antigens 6or
eac: 9ir5s. .it: t:ese tests1 di66erentiation o6 +I&31 and +I&3#
in6ections can 7e acco84lis:ed <it: one test @ig. 7.1A.
T:e I885noco87 2is4ot +I&31 and # =it is 7ased on t:e
4rinci4le o6 t:e solid3 4:ase ',ISA. T:e 4lastic i885noco87 card
65nctions as t:e solid 4:ase. Synt:etic 4e4tide antigens o6 +I&31 and
+I&3# are s4otted on to t:e 4rojections o6 t:e card. T:ere are ra4id
71
B5alitati9e d5al screening tests @+I& S4otA <:ic: in9ol9e tra44ing o6
anti7odies to +I&31 and;or +I&3# 7y t:e ca4t5re reagents <:ic: are
a7sor7ed on a 4oro5s 8e87rane. A6ter <as:ing1 t:e 4resence o6
anti7odies is re9ealed 7y treat8ent <it: a 4ro4rietory 4rotein A3gold
conj5gate <:ic: <ill 7ind to a7sor7ed +I& anti7odies and 6or8 a red
colo5r on t:e 8e87rane. T:e ca4t5re antigens are en9elo4e 4roteins
o6 +I&31 4l5s a :ig:ly 45ri6ied 4e4tide <:ic: corres4onds to a region
o6 t:e en9elo4e trans8e87rane 4rotein o6 +I&3#. In Ca4ill5s +I&3
1;+I&3# screening test1 t:e en9elo4e 4roteins o6 +I&31 and +I&3# are
7o5nd to 4olystyrene late! 7eads and 6or8 t:e 7asis o6 a direct late!
aggregation assay 6or t:e detection o6 anti7odies to +I&31;+I&3# in
ser58 or 4las8a. T:e assay is 4er6or8ed on a 4atented ca4illary slide.
or1 diagnostic 45r4oses1 reacti9e res5lts 7y t:ese screening tests
s:o5ld 7e con6ir8ed 7y s544le8ental;con6ir8atory tests.
7)()" D6al !655lemental8confirmator+ te!t! $ -e!tern Blot and Line Imm6noa!!a+!,
etc)
.2 assays :a9e 7een de9elo4ed t:at :a9e t:e a7ility to identi6y and
di66erentiate in6ections 7y +I&31 and +I&3#. )ost incor4orate t:e 5se
o6 9iral lysates 6ro8 +I&31 and synt:etic 4e4tides o6 +I&3# arti6icially
a44lied on t:e sa8e nitrocell5lose stri4. )5lti4le +I&31 antigens and
one +I&3# s4eci6ic 7and @g436 or g4"1A are a44lied to t:e stri4F
anti7odies to t:e +I&3# antigen are considered to 7e diagnostic i6 a
reaction is 4resent. T:e 7lot also contains a control line1 t:is indicates
t:at t:e test :as 7een 4er6or8ed 4ro4erly @ig. 7.#A. 'ac: co84any
t:at 8ar=ets t:ese tests indicates in t:eir 4ac=age insert t:e criteria
reB5ired 6or 4ositi9ity. T:e criteria esta7lis:ed 7y so8e 8an56act5rers
incl5de reactions to one gene 4rod5ct 6ro8 eac: o6 t:e t:ree 8ajor
genes @gag1 4ol and en9A 6or 4ositi9ity 6or +I&31. To 7e considered
4ositi9e 6or +I&3#1 reaction to t:e +I&3# s4eci6ic antigen 4l5s a reaction
to +I&31 s4eci6ic antigens @75t <:ic: do not 8eet t:e criteria 6or
4ositi9ity o6 +I&31A 85st 7e 4resent.
D5al tests :a9e 7eco8e a9aila7le t:at contain arti6icially a44lied
antigens 6ro8 7ot: +I&31 and +I&3# in a .2 6or8at. T:e ,ine I885no
Assays @,IAsA :a9e gained t:e 4o45larity and are 7ased on t:e
a44lication o6 reco87inant and synt:etic 4e4tide antigens on a 4lastic
s544ort stri4 in a 8anner si8ilar to t:e I885no7lot Assay. T:e ,IAs
are second or t:ird generation assays and :a9e t:e 4otential 6or 5se as
s544le8ental tests. )ost ,IAs 5tilise t:e indirect ',ISA 8et:odology
6or 9is5alising t:e res5lts. T:e 8ajor di66erences 7et<een t:e ,IA and
.2 are t:at t:e antigens are a44lied1 rat:er t:an electro4:oresed1 and
are deri9ed 6ro8 reco87inant or synt:etic 4e4tide tec:nology rat:er
t:an 9iral lysates. In t:e ,IA1 o4ti8al B5antities o6 antigens can 7e
a44lied1 res5lting in an increased ease o6 reading and inter4retation.
T:e 5se o6 t:ese antigens :el4s to eli8inate t:e 4ro7le8s o6
indeter8inate and aty4ical res5lts t:at 8ay occ5r d5e to anti7odies to
cell5lar co84onents in 9iral lysate 7ased con6ir8atory assays. T:ese
assays also allo< 6or t:e a44lication o6 antigens 6ro8 8ore t:an one
9ir5s1 t:ere7y allo<ing t:e8 to act as co87ination assays and to
di66erentiate in6ection 7y +I&31 and +I&3#. T:e ,IAs also contain
control lines t:at indicate t:at t:e test is 4er6or8ed 4ro4erly1 and can
act as a g5ide in esti8ating t:e intensity o6 reactions and :el4ing in
inter4retation. One 4artic5lar co87ination assay incl5des t:ree
reco87inant +I&31 4roteins @4#"1 4171 431A and t<o synt:etic 4e4tide
antigens1 one corres4onding to t:e en9elo4e antigen o6 +I&31 @g4"1A
and one to t:e en9elo4e antigen o6 +I&3# @g436A. T:e ,IA is easy to
4er6or81 reB5ires a total ti8e o6 3 to " :o5rs and 4er6or8s <ell <it:
72
7lood collected on 6ilter 4a4er and on sali9a sa84les. An e!a84le is
gi9en in @ig. 7.3A.
g% 0?>
g% 0=>
% ??
%GG
%G0
g%E0
%@B
%@0
%=E
%0H
Control &'eru#*
g%@?
F-2. G.. HIV;$ '* HIV ; . W"s("r' /0)(
@T
T!A

0T Control le"els
=T
g%E0
%@0
HIVA0
%=E
%0H
g%@? HIVA=
F-2. G.= A ,)1/-'(-)' LIA
In ase of disordant results, final onfir#ation ,ill be done by follo, u% of the
indi"idual and #oleular tests li.e site direted serology, nulei aid
hybridisation and )C6 tehni+ues$
7)' 1rinci5le! of com=ined a!!a+! for HI0$% and HI0$" ; +ere t:e 4rinci4le o6
',ISA re8ains t:e sa8e e!ce4t t:at antigens o6 +I&31 and +I&3#
@reco87inant or 4e4tideA are 7o5nd to 8icro<ells so t:at anti7odies to
t:ese antigens @7ot: ty4esA can 7e detected. So8e co87ined assays
5se reco87inant antigens containing i885nodo8inant regions o6 t:e
en9 and gag gene 4rod5cts to 8a=e t:e test e!tre8ely sensiti9e. As a
res5lt o6 t:is sensiti9ity1 non s4eci6ic reactions 8ay 7e seen in so8e
sa84les. T:ere are co87ined assays <:ic: e84loy synt:etic 4e4tides1
<:ic: corres4ond to t:e :ig:ly antigenic and conser9ed seg8ents o6
t:e en9elo4e 4roteins to +I&31 and +I&3# so as to 8a=e t:e tests :ig:ly
s4eci6ic and o66ering t:e ad9antage o6 8ini8ising t:e incidence o6 non3
73
s4eci6ic reactions. or e!a84le1 in one ',ISA 6or8at1 se4arate <ells
are coated <it: synt:etic 4e4tide antigens corres4onding to
i885nodo8inant regions o6 t:e trans8e87rane o6 +I&31 @g4"1A or
+I&3# @g436A. T:e ',ISA is 4er6or8ed and t:e ratio o6 O.D.s 7et<een
+I&3# and +I&31 <ells is deter8ined. I6 t:e ratio is O0.# t:e s4eci8en
is considered to 7e +I&3# reacti9e. I6 t:e ratio is P0.$1 t:e s4eci8en is
considered to 7e +I&31 reacti9e. Sa84les t:at are 4ositi9e 7y t:e
co87ination test s:o5ld 7e tested 6or con6ir8ation 7y .2 assays <:ic:
co5ld detect 7ot: +I&31 and +I&3#. ,ine I885noassays containing
reco87inant 4roteins and 4e4tides 6ro8 +I&31 and a 4e4tide 6ro8 +I&3
# are also a9aila7le no<adays 6or di66erential detection o6 +I&31 or +I&3
#. T:e res5lts o6 t:e tests s:o5ld 7e inter4reted on t:e 7asis o6
instr5ctions o6 t:e 8an56act5rer as <ell as acce4ted standard nor8s o6
inter4retation. It 8ay 7e necessary to 4er6or8 so4:isticated tests e.g.
PC( 6or con6ir8ation. No<adays1 t:ird generation co87ination assays
:a9e 7een de9elo4ed in 7ot: 8icrotitre and 7ead 6or8ats. T:ese
assays 5tilise reco87inant and;or synt:etic 4e4tide antigens to ca4t5re
anti7odies to eit:er +I&31 or +I&3# <it: la7elled +I&31 and +I&3#
antigens 7eing incor4orated to 7ind to anti7odies. T:ese antigens :a9e
t:e ad9antage o6 7eing a7le to detect 7ot: Ig) and Ig% si85ltaneo5sly1
and t:ere6ore 8ay o66er an increased sensiti9ity. Dot 7lot co87ination
assays are also a9aila7le t:at :a9e se4arate dots containing antigens o6
+I&31 and +I&3#. .it: t:ese tests1 di66erentiation o6 +I&31 and +I&3#
in6ections can 7e acco84lis:ed in one go. T:is sa9es ti8e and 8oney.
Co87ination tests are 8ore e!4ensi9e t:an single detection tests1 75t
are less e!4ensi9e t:an 5sing t<o se4arate screening tests 6or +I&31
and +I&3#1 res4ecti9ely.
&ario5s co88ercial co84anies are 8ar=eting +I&31;+I&3#
co87ination assays. As <it: +I&31 assays a 9ariety o6 6or8ats are
a9aila7le 6or +I&3# incl5ding 7eads1 8icrotitre <ells and dot 7lots.
T:ere are si8ilarities and di66erences 7et<een t:e co87ination assays.
3 )ost incor4orate reco87inant and;or synt:etic 4e4tide antigens in
order to o7tain adeB5ate s4eci6icity.
3 So8e 5se a co87ination o6 t<o synt:etic 4e4tides and ot:ers as 8any
as 6o5r.
3 So8e 5se 7ot: reco87inant and synt:etic 4e4tides si85ltaneo5sly1 or
9iral lysates and synt:etic 4e4tides si85ltaneo5sly.
7)? La=orator+ dia#no!i! of HI0$" infection ; )ost o6 t:e 4rinci4les a44lied to
+I&31 diagnosis @screening and con6ir8atory assaysA also a44ly to +I&3
# diagnosis. Acc5rate diagnosis o6 +I&3# in6ection in t:e la7oratory is
essential to =no< t:e e4ide8iology1 nat5ral :istory and 4at:ogenesis o6
+I&3# in6ection. T:e si8ilarity o6 +I&3# to +I&31 on genetic and
antigenic le9els :as necessitated t:e de9elo48ent and i84le8entation
o6 ty4e s4eci6ic diagnostic assays. Since 8ost +I&31 =its cross react
<it: +I&3# anti7odies1 8ore so4:isticated tec:niB5es li=e synt:etic
4e4tide i885noassays and Poly8erase C:ain (eaction @PC(A :a9e
7een relia7ly 5sed 6or t:e diagnosis o6 +I&3# in6ection. Patients
in6ected <it: +I&3# 6reB5ently gi9e 4ositi9e res5lts <it: +I&31 ',ISA
=its1 75t re8ain indeter8inant or negati9e in +I&31 .2. It is t:ere6ore
stressed t:at +I&31 .2 and +I&3# .2 s:o5ld 7e done 6or con6ir8ation
o6 +I& 3# in6ection <:en 6acilities li=e dot 7lot :y7ridisation and PC(
are not a9aila7le.
Alt:o5g: t:ere are si8ilarities in t:e antigens o6 +I&31 and +I&3#1 t:eir
8olec5lar <eig:t 9ary slig:tly. So8e tests incor4orate certain strains
o6 +I&3# t:at contain en9 antigens o6 8olec5lar <eig:ts si8ilar to
74
t:ose o6 +I&311 75t antigenically distinct. )ost 8an56act5rers o6 test
=its :a9e recently 5sed certain strains o6 +I&3# and de9elo4ed 8et:ods
to 4rod5ce so8e consistency in antigen no8enclat5re.
7)?)% Te!tin# for anti $ HI0$" anti=odie! ; Sa84les s:o<ing grey Jones and
indeter8inate res5lts 7y +I&31 tests s:o5ld 7e s57jected to +I&3#
testing. Cross reactions occ5r <it: t:e core 4roteins @e.g. 4#";#6 and
4$$A and 4ol antigens <:ile t:e en9 antigens do not 4rod5ce cross
reactions. Hits incor4orating en9 antigens are 8ore s4eci6ic 6or
detecting anti7odies to t:is 9ir5s.
7)?)%)% Screenin# a!!a+! ; Screening tests 6or t:e detection o6 anti7odies
to +I&3# are identical to t:ose o6 +I&311 <it: one e!ce4tion t:at t:e
antigens o6 +I&3# are 5sed. T:e screening tests 6or +I&31 in6ection
:a9e already 7een descri7ed in c:a4ters 6. (a4id assays i.e dot 7lot1
co87 assays are also a9aila7le and 5sed in t:e sa8e <ay as 6or +I&31.
7)?)%)" HI0$" Confirmation a!!a+! ; T:e 6ollo<ing +I&3#
con6ir8atory;s544le8ental assays are a9aila7le.

iA +I&3# .estern 2lot @.2A : Se9eral 9iral lysate +I&3# .2 assays are
a9aila7le. +o<e9er1 concens5s con6ir8atory criteria 6or +I&3#
4ositi9ity :a9e not 7een de9elo4ed. S4eci8ens 6ro8 early
serocon9erters o6 +I&31 4atients can dis4lay indeter8inate +I&31 .2
4attern. T:e sera 6ro8 early serocon9erters can 4rod5ce si8ilar
res5lts in +I&3# .2 assay. T:is ill5strates t:e 4ro7le8 o6 5sing 9iral
lysate antigens to serologically di66erentiate 7et<een +I&31 and +I&3#
in6ections. T:e general r5le is t:at cross reactions <ill not occ5r <it:
t:e en9 antigens1 at least not to a signi6icant le9el. T:ere6ore1 <:en
+I&31 .2 4ro6ile e!:i7it reactions to gag and;or 4ol <it:o5t reacti9ity
to en91 +I&3# in6ections s:o5ld 7e considered and an +I&3# ',ISA;or
.2 4er6or8ed @ig 7."A.
iiA +I&3# s4eci6ic ',ISA;(a4id;Si84le assay 5sing di66erent antigens and
di66erent syste8s. @as in strategy II and strategy III o6 National +I&
Testing PolicyA.
iiiA ,ine i885noassays @,IAsA.
i9A (adioi885no4reci4itation assays @(IPAA.
9A Poly8erase C:ain (eaction @PC(A.
75

%@0
%=E
'eru# ontrol

g%@?
F-2. G.> A T96-,0 HIV;. S"r)6)s-(-!" S"r+1
,ist o6 assays a9aila7le co88ercially is gi9en in t:e anne!5re @.+O
a44ro9edA.
76
@) DI*ECT E0IDENCE OF HI0 INFECTION
5.$ I'(r)*+,(-)' : The diagnosis of infetious diseases an be #ade ,ith ertainty
by the diret de#onstration of the %resene of infeting organis#s in the linial
s%ei#ens olleted fro# the %atient$ 5hile isolation of the infeting organis# is
%ossible in #any infetious diseases, the diagnosis #ay also be #ade by
detetion of nulear #aterial or antigens s%eifi for the infeting organis#s in
the linial s%ei#ens$
HIV infetion is diagnosed by the %resene of antiAHIV antibodies in the blood$
Ho,e"er, there are situations ,here the serology is negati"e although there is
definite e"idene of e/%osure to HIV as seen in %atients ,ho are in the ,indo,
%eriod and in health are ,or.ers follo,ing aidental e/%osure to onta#inated
blood et$ 2lso hildren born to HIV infeted #others %resent dile##a as
antibody %ositi"ity seen in the infants u%to 01 #onths, #ay be due to the
#aternal antibodies in irulation$ The diagnosis in these situations #ay be
#ade by:
i* Detetion of )=E antigen &disussed in details in Cha%ter 0=*
ii* Detetion of HIVAs%eifi D82 by )oly#erase Chain 6eation &)C6*
iii* Isolation of HIV$
HIVA0 s%eifi D82 a#%lified in )C6 #ay also be used in Heterodu%le/ Mobility
2ssay &HM2* for deter#ining to ,hih subty%e the infeting "irus belongs$
Detetion of %=E antigen has been disussed in ha%ter 0= J hene the
disussion in this setion ,ill restrit to HIV isolation and HIVA)C6 inluding HM2
for subty%e deter#ination$
@)" 1ol+mera!e Cain *eaction 21C*4 for HI0 ;Serological assays 4ro9ide a
sensiti9e 4roced5re to screen 7lood 6or t:e 4resence o6 anti7odies to
+I&. Sa84les 6o5nd to 7e 4ositi9e 6or +I& anti7odies 7y screening
';(;S tests can 7e tested 7y one or t<o di66erent ';(;S tests
@s544le8entalA 7ased on t:e 4re9alence in t:e 4o45lation or 7y t:e
.estern 2lot test 6or con6ir8ation. In certain sit5ations :o<e9er1 direct
detection o6 +I& <o5ld 7e needed as seen in t:e 6ollo<ing settings:
Dia#no!i! ;
iA Detection o6 +I& in6ection in ne<7orns
iiA Indeter8inate .estern 2lot res5lts
iiiA To 8onitor 9iral load le9els in +I& in6ected indi9id5als d5ring anti3
retro9iral t:era4y
i9A To deter8ine +I& stat5s d5ring <indo< 4eriod
*e!earc ;
iA S57ty4ing o6 +I& 9ir5s
iiA or cloning genes 6or 5se in generating reagents 6or diagnostic 45r4ose
or 6or 4re4aring 9accines
iiiA or generating c:i8eric 9ir5ses
i9A or st5dying 65nctional c:anges ind5ced 7y site directed 85tagenesis
2eca5se o6 t:e sensiti9ity o6 t:e PC( test1 it is i84ortant t:at care is
ta=en to a9oid cross conta8ination o6 sa84les or carry o9er o6
a84li6ied 4rod5cts t:at can res5lt in 6alse 4ositi9e res5lts. Since PC(
can detect s8all n587er o6 8olec5les in t:e sa84le tested1 a9oidance
77
o6 conta8ination o6 4rocessed DNA 7y e9en 8in5te B5antities o6 target
8olec5les is i84ortant to o7tain relia7le res5lts.
@)")% 1rinci5le of 1ol+mera!e Cain *eaction ; PC( :el4s to 6ind a s4eci6ic gene
seB5ence a8ongst an a75ndance o6 ot:er DNA. S4eci6ic
oligon5cleotides are c:e8ically synt:esised @called 4ri8ersA <:ic: are
co84li8entary to DNA 6lan=ing t:e seB5ence o6 interest. %eno8ic DNA
is arranged in t:e 6or8 o6 a do57le stranded :eli! :eld toget:er 7y
:ydrogen 7onds. On :eating1 t:e do57le stranded DNA gets denat5red
to single stranded DNA. On cooling t:e DNA sol5tion1 t:e 4ri8ers 7ind
to t:e single strands o6 DNA. One 4ri8er is designed to anneal to t:e
sense strand and t:e ot:er 4ri8er to t:e anti3sense strand1 <it: t:eir 3V
ends 4ointing to<ards eac: ot:er.
Pri8ers are 8i!ed <it: a 7566ered sol5tion o6 dNTPs1 8agnesi581
t:er8osta7le 4oly8erase enJy8e and t:e te84late DNA. T:e 8i!t5re is
t:en co9ered <it: a layer o6 8ineral oil to 4re9ent e9a4oration and
4laced in a 4rogra88a7le :eating 7loc= @t:er8al cyclerA. T:e do57le
stranded DNA te84late is t:en denat5red 7y :eating to a te84erat5re
a7o9e its 8elting 4oint. T:e te84erat5re is t:en lo<ered s566iciently 6or
:y7ridiJation 7et<een 4ri8ers and te84late to occ5r. T:e te84erat5re
is :o<e9er1 =e4t :ig: eno5g: to 4re9ent 8is8atc: :y7ridisation o6 t:e
4ri8ers to si8ilar seB5ences else<:ere in t:e geno8e. +ig:
concentration o6 t:e 4ri8ers is 8aintained to ens5re t:at t:is reaction
is 6a9o5red o9er re3annealing o6 t:e co84li8entary te84late DNA
strands.
T:e te84erat5re is t:en raised to<ards t:e o4ti858 6or t:e
t:er8osta7le 4oly8erase enJy8e @7# deg CA1 <:ic: :as attac:ed itsel6
to t:e end o6 t:e 4ri8er3te84late d54le!. Synt:esis 4roceeds 6ro8 t:e
3V end o6 eac: 4ri8er 5ntil t:e reaction is sto44ed 7y :eating to t:e
8elting 4oint 6or t:e second ti8e. T:e 4rod5ct o6 t:is reaction is o6
inde6inite lengt: and is =no<n as long PC( 4rod5ct. T:is co84letes t:e
6irst cycle o6 PC(.

T:e second PC( cycle co88ences <it: t:e 8elting ste41 6ollo<ed 7y
4ri8er annealing. +ere1 t:e 4ri8ers anneal not only to t:e original DNA
75t also to t:e ne<ly synt:esised strands 6ro8 t:e 6irst cycle o6 PC(.
T:e second cycle re4eats t:e 6irst cycle <it: res4ect to t:e original
DNA1 75t synt:esis on t:e ne< strands can 4roceed only as 6ar as t:e
end o6 t:e 8olec5le <:ic: corres4onds to t:e $V end o6 t:e o44osite
4ri8er.
A6ter t:e t:ird cycle o6 PC(1 synt:esis directed 7y t:e PC( 4rod5cts o6
t:e 6irst t<o cycles <ill 7e 7o5nd 7y 7ot: ends o6 t:e 4ri8er seB5ences
and t:is PC( 4rod5ct <ill acc585late e!4onentially <it: s57seB5ent
cycles o6 a84li6ication. T:e 8ajor 4rod5ct o6 PC( <ill t:5s 7e o6 de6ined
lengt:1 consisting o6 t:e seB5ence 7et<een t:e 4ri8ers and incl5des
t:e 4ri8er seB5ences t:e8sel9es.
In t:e +I& PC( reaction1 o6ten a nested PC( reaction is 5sed. +ere1 an
aliB5ot o6 t:e PC( 4rod5ct can 7e re3a84li6ied <it: a second set o6
4ri8ers <:ic: are located at 3V or internal to t:e original set o6 4ri8ers
5sed 6or t:e 6irst ro5nd o6 PC( a84li6ication.
@)")" 1C* 1roced6re ;
@)")")% Sam5le 1re5aration for 5roviral DNA 1C* ; $ to 10 8l. o6 7lood is collected
7y 9ene45nct5re in an 'DTA or acid citrate de!trose containing
78
9ac5tainer t57e. 3 8l. o6 icoll3+y4aB5e is 4i4etted into a conical
centri65ge t57e. T:e <:ole 7lood collected is care65lly layered on to4 o6
t:is icoll3+y4aB5e sol5tion. T:e t57es are t:en centri65ged at #000
r48 6or 30 8in5tes at roo8 te84erat5re in a s<ing o5t 75c=et rotor.
T:e ly84:ocyte;8onocyte 6raction a44earing 7et<een icoll3+y4aB5e
layer and 4las8a layer is trans6erred to anot:er t57e and <as:ed t<ice
<it: eit:er 4:os4:ate 7566ered saline @P2S 4+ 7.# 3 7."AA or <it: (P)I
16"0. T:e cells are 4elleted 7y centri65ging at 1$00 r48 6or 10 8in5tes.
T:e 4elleted cells can 7e 5sed 6or DNA e!traction or can 7e
res5s4ended in D0Q 6oetal cal6 ser58 @CSA and 10Q D)SO and stored
at 370
o
C or in liB5id nitrogen.
@)")")" DNA EHtraction ; DNA can 7e e!tracted 6ro8 t:e se4arated ly84:ocytes
5sing ready8ade =its a9aila7le in t:e 8ar=et @e.g. K5iagen1 IsoB5ic=A.
DNA e!tracted 6ro8 a7o5t 3 8illion cells is res5s4ended in 100 l o6
<ater 4ro9ided in t:e =it. T:e DNA can 7e stored at eit:er 370
o
C 6or
longter8 5se or at "
o
C i6 t:e PC( 4roced5re is to 7e 4er6or8ed soon
a6ter e!traction.
@)")")< 1rimer! 6!ed for 1C* reaction 2de!i#ned for envelo5e #ene4 ;
Fir!t ro6nd of 1C*
Pri8er 1
$V TTA %%C ATC TCC TAT %%C A%% AA% AA% C%% 3V
Pri8er #
$V TCT T%C CT% %A% CT% TTT %AT %CC CCA %AC 3V
Second ro6nd of 1C*
Pri8er 1
$V T%T AAA AC% AC% %CC A%T CT% TTA AAT %%C A%T CTA %C 3V
Pri8er #
CA% %AA ACA %CT AT% ACC CAC TTC TCC AAT T%T CCC TCA 3V
@)")")( 1C* *eaction ; T:e nested PC( reaction 6or detection o6 +I& is carried
o5t in 0.$ 8l PC( reaction t57es 6or eac: sa84le1 5tilising t:e rele9ant
4ri8ers and t:e 4atientVs DNA sa84le. In t:e nested PC( reaction1 t:e
6irst ro5nd o6 PC( is carried o5t 5sing t:e 4atient0s DNA sa84le <:ile
t:e second ro5nd o6 PC( is done 5sing $ l o6 t:e 6irst ro5nd PC(
4rod5ct. PC( is also done 6or t:e re6erence 4las8ids @o7tained 6ro8
t:e NI+1 2et:esda1 CSA reagent 7an=A i6 :eterod54le! 8a44ing assay is
to 7e done 6or +I&31 s57ty4ing. 'ac: ro5nd o6 PC( reB5ires 4re4aration
o6 a 8aster 8i! co84osed o6 t:e 6ollo<ing:
3 10S PC( 7566er @10 l 6or 100 l reactionA
3 10 8) )gCl# @1#.$ l 6or 100l reactionA
3 DNTP 8i!t5re @#00 ) o6 eac: dNTPA
3 Pri8er 1 @#0 48ole 6or 100 l reactionA
3 Pri8er # @#0 48ole 6or 100 l reactionA
3 TaB 4oly8erase enJy8e @#.$ 5nits 6or 100 l reactionA
3 dd <ater @ to 8a=e 54 t:e 9ol58e to 100 lA
T:e reB5ired 9ol58e o6 t:e 8aster 8i! is t:en distri75ted into t:e PC(
reaction t57es and o9erlaid <it: 8ineral oil. T:e DNA sa84le is t:en
added to t:e PC( reaction t57e 5sing 7arrier ti4s to a9oid cross
conta8ination. T:e t57es are t:en 4laced in t:e t:er8al cycler and t:e
79
PC( reaction carried o5t. T:e t:er8al cycling conditions reB5ired are as
6ollo<s:
3 cycles o6 t:e 6ollo<ing:
3 D"
o
C 6or 1 8in
3 $$
o
C 6or 1 8in
3 7#
o
C 6or 1 8in5te
6ollo<ed 7y 3# cycles o6 t:e 6ollo<ing:
3 D"
o
C 6or 1$ sec
3 $$
o
C 6or "$ sec
3 7#
o
C 6or 1 8in.
A6ter t:is1 t:e PC( reaction is 8aintained at 7#
o
C 6or $ 8in5tes and
t:en cooled to "
o
C. A6ter co84letion1 t:e a84li6ied PC( 4rod5cts are
r5n on agarose gel to detect t:e 4resence o6 t:e PC( 4rod5cts. 10l o6
eac: second ro5nd PC( 4rod5ct o6 a nested PC( reaction is 8i!ed <it:
# l o6 a 6S 7ro8o4:enol 7l5e;!ylene cyanol dye 8i! and r5n on a 1Q
agarose gel containing et:idi58 7ro8ide @30 l o6 a 10 8g;8l sol5tion
added to $0 8l o6 1Q agarose sol5tion 8ade in 1S TA' 7566erA.
Si85ltaneo5sly1 a DNA 8ar=er s:o5ld 7e r5n <it: e9ery gel to detect
t:e siJe o6 t:e PC( 4rod5ct @1 H7 DNA ladder co5ld 7e 5sedA. A6ter
r5nning t:e gel on a :oriJontal gel electro4:oresis a44arat5s1 t:e gel is
trans6erred onto a C& transill58inator and a 4ict5re o6 t:e gel ta=en
5sing a s5ita7le gel doc58entation syste8. Presence o6 t:e PC(
4rod5ct o6 t:e reB5ired siJe @as s:o<n 7y co84arison o6 t:e PC(
4rod5ct <it: t:e DNA 7ands seen in t:e 8olec5lar <eig:t 8ar=er laneA
indicates a 4ositi9e PC( reaction. .it: t:e a7o9e 8entioned 4ri8ers1 a
PC( 4rod5ct o6 700 7ase 4air siJe s:o5ld 7e detected.
@)")")' 1reca6tion! to =e follo.ed for 1C* reaction ;
iA All 7lood sa84les 85st 7e :andled care65lly to a9oid 4ossi7ility o6
trans8ission o6 +I&. ,a7 coats and glo9es s:o5ld 7e 5sed <:ile
:andling t:e sa84les and PC( 4rod5cts. 2iosa6ety la8inar 6lo< :oods
s:o5ld 7e 5sed to 4re9ent cross conta8ination as <ell as 6or 7iosa6ety
4reca5tion.
iiA Cse o6 a dedicated area t:at is 6ree o6 PC( 4rod5ct 6or t:e 4re4aration
o6 t:e PC( reaction. A84li6ied 4rod5cts are to 7e =e4t a<ay 6ro8 t:e
sa84le 4rocessing area as 6ar as 4ossi7le.
iiiA Cse o6 7arrier ti4s or 4ositi9e dis4lace8ent 4i4ettes 6or dis4ensing o6
sa84les.
i9A (eagents s:o5ld 7e stored as s8all aliB5ots <:ere 4ossi7le
9A Pre4aration o6 8aster 8i!es to 8ini8iJe :andling o6 reagents
ig. E.1 de4icts t:e 4rinci4le o6 PC( 4roced5re and ig. E.# s:o<s t:e
7ands o7tained a6ter gel electro4:oresis o6 en9 PC( 4rod5cts @700 74A
r5n <it: ;+ind III 8ar=er
@)")<) *ea#ent! reC6ired for 1C*
Sr.No Na8e o6
c:e8ical
Cat.No. K5antity Price
80
01. (P)I 16"0 (6$0" 10 , W 13.E0
0#. D)SO D#6$0 100 8l W 76.60
03. +isto4aB5e +EEED $00 8l W 76.$0
0". etal 2o9ine
Ser58
1600030"" $00 8l W#10.00
0$. KIAa84 7lood =it %S#670KN 1 No. (s. 7##6;3
06. TaB. Poly8erase ,2D 1000 Cnits (s. E000;3
07. dNTP set %S$3"1)2 1 set (s. 7373;3
0E. ,a87da +ind III
digest
%S$#$1)2 1 9ial (s. #3D6;3
0D. 1 H7 DNA ladder %S$#"D)2 1 9ial (s. "7D#;3
10. Agarose 1$$1030#7 $00 g8 W 36".00
11. 't:idi58
2ro8ide
'1$10 10 8l W 30.10
1#. TriJ8a 7ase T1$03 $ Hg W #67.30
13. 'DTA '$13" $00 g8 W 66.E0
1". %lacial acetic
acid
1100$ $00 8l (s. 10";3
1$. 2ro8o4:enol
7l5e
2$$#$ $ g8 W 13.$0
16. Iso4ro4anol 13E#$ $00 8l (s. 100;3
17. 't:yl Alco:ol 10107 $00 8l (s $7$;3
1E. Sylene cyanol S"1#6 10 g8 W ##.10
T:e reagents listed a7o9e are t:ose 7eing 5sed at National AIDS
(esearc: Instit5te @NA(IA1 P5ne. T:e co84any0s na8e :as 7een
deli7erately <it::eld to a9oid 7ias. )olec5lar 7iology standard
reagents can 7e 45rc:ased 6ro8 any co84any dealing in s5c: c:e8icals
and reagents.
81
@)")( EC6i5ment reC6ired for 1C*
Sr.No Na8e o6 'B5i48ent
01. +oriJontal gel electro4:oresis
a44arat5s
0#. Po<er s544ly )odel #$0 @6or PC(A
)odel $00 @6or
+)AA
03. &ertical %el 'lectro4:oresis
a44arat5s @6or +)AA
0". C& Transill58inator
0$. %el doc58entation syste8
06. PC( t57es 1.$ 8l
0.$ 8l
07. )icro4i4ettes P #0
P #00
P 1000
0E. ,a8inar 6lo< @#V!#V!#V 6or
reagentsA
@"V!#V!#V 6or
sa84lesA
0D. +eat 7loc=
10. '44endor6 centri65ge
11. Progra88a7le T:er8al Controller
@PC( 8ac:ineA
1#. CPS @1 H&AA <it: " 7atteries
13. &orte! 8i!er
1". .ater 7at:
1$. )icro<a9e o9en
16. Pi4ette aid <it: c:arger
17. (e6rigerater
@)< Heterod65leH Mo=ilit+ A!!a+ 2Hma4 ; I6 t:e PC( 4rod5cts are to 7e 5sed 6or
s57ty4ing o6 t:e +I&31 9ir5s in t:e sa84le 6or e4ide8iological reasons1
t:e :eterod54le! 8o7ility assay co5ld 7e carried o5t. +eterod54le!es
are 6or8ed 7y 8elting a84li6ied PC( 4rod5ct 6ro8 5n=no<n 4atient0s
sa84le <it: a84li6ied PC( 4rod5ct 6ro8 re6erence strains and analysed
on 4olyacryla8ide gel electro4:oresis. +eterod54le!es 6or8ed 7et<een
t:e 5n=no<n sa84le and t:e 8ost closely related seB5ences e!:i7it t:e
6astest 8o7ilities on 4olyacryla8ide gel electro4:oresis. $l o6 t:e
second ro5nd PC( 4rod5ct o6 4atient0s sa84le is 8i!ed <it: $l o6 PC(
4rod5ct o6 di66erent re6erence 4las8ids in di66erent t57es toget:er <it:
1.1 l o6 annealing 7566er in eac: t57e and :eated in a t:er8al cycler
6or # 8in5tes at D"
o
C and t:en 4laced on <et ice. In one o6 t:e t57es1
82
10l o6 t:e 4atient0s PC( 4rod5ct is ta=en toget:er <it: annealing
7566er <it:o5t t:e addition o6 re6erence 4las8id PC( 4rod5ct. T:e
:eterod54le! reaction is 8i!ed <it: 3l o6 $S loading dye and t:en
loaded onto a $Q non3denat5ring 4olyacryla8ide gel and r5n at
constant c5rrent. A6ter r5nning t:e sa84les 6or adeB5ate d5ration
@de4ending on t:e siJe o6 t:e PC( 4rod5ctA1 t:e electro4:oresis is
ter8inated and t:e acryla8ide gel is 4laced in et:idi58 7ro8ide
containing 7566er 6or a7o5t an :o5r. S57seB5ently1 t:e acryla8ide gel
is 4laced on a C& transill58inator and 4:otogra4:ed 5sing s5ita7le
6ilter lenses. De4ending on t:e 4attern o6 t:e 7ands o7ser9ed1 t:e
s57ty4e o6 +I&31 in t:e DNA sa84le can 7e deter8ined.
@)<)% *ea#ent! reC6ired for Hma ;
S.N
o
Na8e o6
c:e8ical
)a=e Cat.No. K5anti
ty
Price
1. Acryla8ide @6or
+)AA
%i7co
2(,
1$$1#3
0#3
$00
g8
W
103.00
#. 2is3acryla8ide
@6or +)AA
%i7co
2(,
1$$163
0#"
100
g8
W 6".00
3. 2oric acid @6or
+)AA
Sig8
a
27660 1 Hg W "D.D0
". A88oni58 4er
s5l6ate @6or
+)AA
Sig8
a
AD16" #$ g8 W 13.70
$. T')'D @6or
+)AA
Sig8
a
TE133 $0 8l W 10."0

@)( I!olation of HI0; +I& 8ay 7e isolated 6ro8 t:e 7lood and ot:er 7ody
6l5ids. Isolation o6 +I& is a s4ecialiJed 4roced5re reB5iring at least P#R
contain8ent 6acility and :ig: degree o6 e!4ertise. +I& isolation :as
t:ere6ore re8ained 8ostly as a tool 6or researc:. +I& is isolated 6ro8
t:e 4eri4:eral 7lood 8onon5clear cells @ P2)C A or 4las8a and ot:er
7ody 6l5ids. T:e acti9ated CD"R cells are s5sce4ti7le 6or +I& in6ection.
+ence a5tologo5s or :eterologo5s P2)Cs acti9ated <it: 8itogen
P:yto:ae8aggl5tinin @P+AA are c5lt5red <it: t:e in6ectio5s 8aterial.
T:e c5lt5res are 8aintained at 37
0
C in $Q CO# at8os4:ere 6or 54 to #E
days. T:e c5lt5res are 6ed <it: 6res: acti9ated P2)Cs at reg5lar
inter9als. T:e 4resence o6 9ir5s in t:e c5lt5re s54ernatant is detected
eit:er 7y de8onstration o6 t:e 4resence o6 4#" antigen or enJy8e
re9erse transcri4tase. T:e in6ected cells 8ay also de8onstrate syncitia
in c5lt5re. &ir5s in6ected cells 8ay also 7e detected 7y +I&3s4eci6ic
i885no6l5orescent assay.
T:e t<o 8et:ods nor8ally 5sed 6or 9ir5s isolation are direct 8et:od or
co3c5lt5re 8et:od. In t:e direct 8et:od1 P2)Cs 6ro8 t:e 4atient are
c5lt5red in 9itro in 4resence o6 P+A. In t:e co3c5lt5re 8et:od P2)Cs
6ro8 :eterologo5s +I& 5nin6ected donor are sti85lated <it: P+A1 and
a6ter "E37# :o5rs t:e sti85lated cells are c5lt5red along <it: t:e
P2)Cs 6ro8 t:e 4atient. In o5r e!4erience1 co3c5lt5re 8et:od gi9es
greater s5ccess in 9ir5s isolation.
83
4. GUIDELINES FOR HIV TESTING
4.$ I'(r)*+,(-)' : The HIV!2ID' e%ide#i has raised a nu#ber of issues
%artiularly in the health are syste# and one of the "ery i#%ortant
issues has been the HIV test itself$ HIV testing has generated interest
not only in the sientifi o##unity but also a#ongst the general %ubli$
The "arious tehnial, ethial and legal issues that in"ariably ao#%any
the HIV testing ha"e led the ountries the ,orld o"er to de"elo% their
o,n HIV testing %oliies and guidelines$ The de"elo%#ent of tests to
detet infetion ,ith HIV has #ade it %ossible to deter#ine the
%re"alene of HIV infetion and to #onitor trends of the infetion in
%o%ulations$ Though this infor#ation is of great "alue in designing,
i#%le#enting and #onitoring %ubli health %rogra#s for the %re"ention
and ontrol, ho,e"er, testing of any %o%ulation for HIV re+uires areful
onsideration of a nu#ber of %oints relating to logistis, laboratory,
o%erational, legal and ethial$ These as%ets should be .e%t in #ind
,hile %resribing any HIV test$
4.. O/#",(-!"s )3 HIV ("s(-'2 : The early detetion of HIV infetion differs
fro# si#ilar detetion of #ost other infetious diseases on aount of the
follo,ing reasons : A
A Due to %rolonged asy#%to#ati stage of infetion, one re#ains fully
ati"e and de#ands an a%%ro%riate inter"ention ,hih #aintains the
lifestyle and dignity of the indi"idual$
A HIV infetion is belie"ed to be in"ariably fatal irres%eti"e of the best
%ossible treat#ent$
A HIV infetion and 2ID' are still assoiated ,ith high degree of
disri#ination and stig#atisation$
A The i#%liations of a %ositi"e test go ,ell beyond those related to
%hysial and #ental health of the indi"idual being tested$
Thus, any HIV testing that is done has any of the four follo,ing ob7eti"es
:
i* To #onitor the trends of HIV infetion in a %o%ulation or subgrou% for
failitation of inter"ention &sur"eillane : unlin.ed and anony#ous*$
ii* To test donated blood or donors of organs or tissues for ensuring safety
of the rei%ients &transfusion safety and donation safety*$
iii* To identify an indi"idual ,ith HIV infetion for diagnosis &,ith 2ID'
indiator diseases* or "oluntary testing %ur%oses &asy#%to#ati or
2ID' ases*$
i"* 6esearh$
4.= HIV ("s(-'2 s(r("29 : (ften the se%arate ob7eti"es #entioned abo"e
annot be #et by a single testing strategy$ Different ob7eti"es re+uire
se%arate testing %roedures and the hoie of tests &disussed in detail in
ha%ters ?*$ Based on rele"ant onsideration &ha%ter ?* different
%roedures and strategies of testing are ado%ted$
The different %roedures for testing are :
4.=.$ U'0-'B"* ')'91)+s ("s(-'2 : 'uh ty%e of sreening or testing is not
84
direted to the indi"idual, but has as its ob7eti"e, the %ubli health
sur"eillane of HIV infetion$ It is an e%ide#iologial #ethod for
#easuring HIV %re"alene in a seleted %o%ulation ,ith the #ini#u# of
%artii%ation bias$ By #ini#ising %artii%ation bias, unlin.ed anony#ous
sreening offers a distint e%ide#iologial ad"antage o"er #andatory or
"oluntary testing$ <nlin.ed anony#ous testing in"ol"es use of blood
already olleted for other %ur%osesJ therefore, the effet of seletion
bias ,ill re#ain and ,ill de%end u%on ti#e, loation and other details of
blood olletion$
4.=.. V)0+'(r9 ,)'3-*"'(-0 ("s(-'2 : Testing is often done for diagnosti
%ur%oses$ Here it is i#%ortant that the issues related to ,)'3-*"'(-0-(9
reei"e great attention$ 'ine this #ethod is based on "oluntary HIV
testing or testing for diagnosis of HIV!2ID' ases, it is i#%erati"e to
res%et the indi"idualCs need to #aintain onfidentiality$ By #aintaining
onfidentiality, it ,ill not only instill faith in the indi"idual about the
health are syste# in the o##unity but also enourage #ore and #ore
%eo%le %ratiing ris. beha"iour to o#e for,ard for an HIV test$
4.=.= M'*()r9 ("s(-'2 : 5hen testing is done ,ithout the onsent of the
%atient and data ould be lin.ed to identify the %erson it is alled
#andatory testing$ Mandatory testing is reo##ended only for
sreening donors of se#en, organs or tissues in order to %re"ent
trans#ission of HIV to the rei%ient of the biologial %roduts$
The hoie of tests is also based on the different ob7eti"e of HIV testing$
The test that are ado%ted are the ;LI'2, 6a%id or 'i#%le, lubbed
together as K;!6!'C$ (ne ;!6!' denotes test done on one single antigen
%re%arationJ t,o is ,hen all %ositi"e sa#%les on first antigen test is
re%eated on a seond antigen %re%aration and three is ,hen this test is
re%eated for a third ti#e using a different antigen syste#$ -or
transfusion safety %ur%oses one ;!6!' is usedJ for sur"eillane and
diagnosis of full blo,n 2ID' ases t,o ;!6!' are used and for
asy#%to#ati indi"iduals three ;!6!' are used$
4.> G"'"r0 6r-',-60"s )3 HIV ("s(-'2 : Testing %oliy in general should
onsider the follo,ing %oints :
i* It should be %art of the o"erall o#%rehensi"e and %re"enti"e %rogra#$
ii* Testing should be tehnially sound and a%%ro%riate$
iii* Test %roedure #ust be a%%ro%riate to the field situation$
i"* Testing %roedure #ust be ost effeti"e$
"* Laboratory %roedure #ust be #onitored for ensuring +uality$
4.? HIV ("s(-'2 -' H"0(h ,r" s"((-'2s : The fear and a%%rehension that
e/ists a#ong health are ,or.ers in #anaging HIV infeted indi"iduals
and 2ID' %atients is largely due to the #ini#al ris. that e/ists of HIV
trans#ission due to a needle sti. or other shar% in7ury$ Thus the
de#and for #andatory HIV testing of %atients ad#itted in hos%itals or
undergoing surgery et$ This de#and is neither rational nor a%%ro%riate$
2 #andatory HIV test is no substitute for U'-!"rs0 %r",+(-)'s that
need to be ado%ted for e"ery %atient in a hos%ital or any other health
are setting$ (n the other hand testing ,ithout "A60-,-( ,)'s"'( of the
%atient has been %ro"ed to be ounter%roduti"e in the long run$ In the
ontrol of the HIV e%ide#i suh testing an dri"e the target %eo%le
85
underground and #a.e it #ore diffiult for launhing inter"entions$
The '(-)'0 ("s(-'2 6)0-,9 reiterates the follo,ing :
A 8o indi"idual should be #ade to undergo a #andatory testing for HIV$
A 8o #andatory HIV testing should be i#%osed as a %reondition for
e#%loy#ent or for %ro"iding health are ser"ies and failities$
A 2ny HIV testing #ust be ao#%anied by a %retest and %ost test
ounselling ser"ies$
,ist o6 9ol5ntary 7lood testing centers and re6erences centers
@NACOA are gi9en in Anne!5re D.
86
Ch6("r 4 ; A''"A+r" I
HIV testing failities ha"e been lassified into Donal blood testing entres
&IBTCs* ,hih sreen the donated units of blood, sur"eillane entres
,hih arry out HIV testing for sur"eillane, "oluntary blood testing
entres ,here HIV testing for diagnosis and identifiation of the indi"idual
is underta.en ,ith %re and %ost test ounselling and national referene
entres ,hih underta.e all the abo"e ati"ities along ,ith the 7obs
defined under the entres$ List of 8ational 6eferene Centes and the
Voluntary and 'ur"eillane Testing 'ites is %laed belo, for ready
referene$
L-s( )3 HIV R"3"r"'," C"'(r"s
0$ 8ational Institute of Co##uniable Diseases, Delhi$
=$ 2ll India Institute of Medial 'ienes, 8e, Delhi$
@$ Indian Institute of I##unohae#atology, Bo#bay$
E$ 8ational Institute of Cholera and ;nteri Diseases, Calutta$
G$ 'hool of Tro%ial Mediine, Calutta$
?$ Madras Medial College, Chennai$
H$ 8aional 2ID' 6esearh Institute &826I*, )une$
1$ 6egional Medial College, I#%hal$
B$ Christian Medial College, Vellore$
The referene entres are entrusted ,ith the res%onsibility of arrying out
onfir#atory test$ They are also res%onsible for diagnosis, +uality ontrol
of HIV .its, guidelines for HIV testing, training in HIV testing and any
other ati"ity ,hih #ay be neessary for standardiDation of HIV testing$
These entres also underta.e "oluntary blood testing ,ith %re and %ost
test ounselling$ 2lso %ro"ide training for %ratie of Biosafety
)reautions and Quality 2ssurane 'yste#s$
L-s( )3 s+r!"-00'," '* !)0+'(r9 /0))* ("s(-'2 ,"'(r"s
s',(-)'"* +6() $445.
S',(-)'"* -' $445 EA-s(-'2
$. ANDHRA %RADESH
i 2ndhra Medial College,
Visha.ha%atna#$
/ De%tt$ of Mirobiology, (s#ania
Medial College, Hyderabad$
ii 6angaraya Medial College,
:a.inada$
/i De%tt$ of Mirobiology, '$V$
Medial College Tiru%ati$
iii 3untur Medial College, 3untur$ /ii De%tt$ of Mirobiology, 2ndhra
Medial College Visha.ha%atna#$
i" 'iddhastha Medial College,
Vi7aya,ada$
/iii 'ur"eillane Centre, Instt$ of
)re"$ Mediine, Hyderabad
" (s#ania Medial College,
Hyderabad$
/i" 'ur"eillane Centre, Indian 8a"al
'hi% Hos%ital, :alyani,
Visha.ha%atna#$
87
"i 3andhi Medial College,
'eunderabad$
"ii :a.tiya Medial College,
5arrangal$
"iii :urnool Medial College, :urnool$
i/ '$V$Medial College, Tiru%ati
.. ASSAM
i 3u,ahati Medial Cellege,
3u,ahati$
i" De%tt$ of Mirobiology, 3u,ahati
Medial College 3u,ahati
ii 'ilhar Medial College, 'ilhar$
iii 2ssa# Medial College, Dibrugarh$
=. BIHAR
i Darbhanga Medial College,
Leharia 'arai$
/ 6a7endra Me#orial 6esearh
Institute, )atna
ii '$:$ Medial College, MuDDafar%ur$
iii )atna Medial College, )atna
i" 6a7endra Medial College, 6anhi$
" M$3$M$ Medial College,
9a#shed%ur$
"i )atli%utra Medial College,
Dhanbad$
"ii Medial College, Bhagal%ur$
"iii Magadh Medial College, 3aya$
i/ 8alanda Medial College, )atna$
>. DELHI
i 2$I$I$M$'$, 8e, Delhi$ " De%tt$ of Mirobiology, <ni"ersity
College of Medial 'ienes,
'hahdara, Delhi$
ii M$2$M$C$, 8e, Delhi$ "i De%tt$ of Mirobiology, Maulana
2Dad Medial College, 8e, Delhi
iii <$C$M$'$, 8e, Delhi$
i" Lady Hardinge Medial College,
8e, Delhi$
"ii 'ur"eillane Centre, 2r#ed
-ores Co##and Hos%ital, Delhi
Cantt$
?. GOA
i 3oa Medial College, 3oa$ ii De%tt$ of Mirobiology, 3oa
Medial College, )ana7i
F. GUKARAT
i B$9$ Medial College, 2h#edabad$ "i De%tt$ of Mirobiology, B$9$
Medial College, 2h#edabad
88
ii Munii%al Medial College,
2h#edabad$
iii Medial College, Baroda$
i" M$)$ 'hah Medial College,
9a#nagar$
" 3o"t$ Medial College, 'urat$
G. HARYANA
i 3o"t$ Medial College, 6ohta.$ ii De%tt$ of Mirobiology, Medial
College 6ohta.$
5. HIMACHAL %RADESH
i I$3$M$C$ 'hi#la$ ii De%tt$ of Mirobiology, Indira
3andhi Medial College 'hi#la$
4. KAMMU '* KASHMIR
i 3o"t$ Medial College, 'rinagar$ i" De%art#ent of
I##uno%athology, 'herAeA
:ash#ir Instt$ of Medial
'ienes, 'rinagar$
ii 3o"t$ Medial College, 9a##u$ " De%tt$ of Mirobiology, 3o"t$
Medial College 9a##u$
iii 'herAeA:ash#ir Institute of Medial
'ienes, 'rinagar$
$I
.
KARNATAKA
i Mysore Medial College, Mysore$ "i De%tt$ of Mirobiology,
Bangalore Medial College,
Bangalore$
ii Bangalore Medial College,
Bangalore
"ii De%tt$ of Mirobiology, :asturba
Medial College, Mani%ur$
iii :arnata.a Medial College, Hubli$
i" Medial College, Bellary$
" Medial College, Mangalore$
$$
.
KERALA
i Medial College, Tri"andru#$ "i De%tt$ of Mirobiology, Medial
College, Tri"andru#$
ii T$D$ Medial College, 2lle%ey$ "ii 'ur"eillane Centre, Indian
8a"al 'hi% Hos%ital, Cohin$
iii Medial College,Caliut$
i" Medial College, Trihur$
" Medial College, :ottaya#$
89
$.
.
MADHYA %RADESH
i 3o"t$ Medial College, 9abal%ur$ "ii De%tt$ of )athology, 3andhi
Medial College, Bho%al$
ii 3$ 6$ Medial College, 3,alior$ "iii 6egional Medial 6esearh
Centre for Tribal Health,
9abal%ur$
iii M$3$M$ Medial College, Indore$ i/ Choitra# Hos%ital and 6esearh
entre, Indore$
i" 3andhi Medial College, Bho%al$
" '$'$ Medial College, 6e,a
"i )t$ 9$L$8$ Medial College, 6ai%ur$
$=
.
MAHARASHTRA
i 3rant Medial College, Mu#bai$ /i" De%tt$ of Mirobiology, 'eth 3$'$
Medial College, Mu#bai$
ii 'eth$ 3$'$ Medial College,
Mu#bai$
/" De%tt$ of Mirobiology, 9$9$
Hos%ital, Mu#bai
iii T$ 8$ Medial College, Mu#bai$ /"i 'ion Hos%ital, Mu#bai
i" L$T$M$ Medial College, Mu#bai$ /"ii B$L$8$ 8air Hos%ita, Mu#bai
" B$9$ Medial College, )une /"iii 6a7abari Hos%ital, 3hat.o%ar,
Mu#bair
"i 2$-$M$C$, )une$ /i/ De%tt$ of Mirobiology, B$9$
Medial College, )une$
"ii Mira7 Medial College, 'hola%ur$ // De%tt$ of Mirobiology, 3o"t$
Medial College, 8ag%ur$
"iii Dr$ V$ M$ Medial College, 'hola%ur$ //i 'ur"eillane Centre, Ci"il
Hos%ital, :olha%ur
i/ 3o"t$ Medial College, 2urangabad$ //ii 'ur"eillane Centre, Distrit
Hos%ital, Chandra%ur$
/ '$6$T$6$ Medial College,
2#ba7ogari$
//iii De%tt$ of Mirobiology, 3o"t$
Medial College, Mira7$
/i Medial College, 8ag%ur$ //i" 'ur"eillane Centre, Indian
8a"al 'hi% Hos%ital, 2sh,ini,
Mu#bai$
/ii Indira 3andhi Medial College,
8ag%ur$
//" De%tt$ of Mirobiology, 2r#ed
-ores Medial College, )une$
/iii 3o"t$ Medial College, 8anded$
$>
.
ORISSA
i '$C$B$ Medial College, Cutta.$ i" De%tt$ of Mirobiology, '$C$B$
Medial College, Cutta.
ii V$'$'$ Medial College, Buria$ " 'ur"eillane Centre, 6egional
Medial 6esearh Centre,
Bhubnesh,ar$
iii M$:$C$3$ Medial College,
Berha#%ur$
$?
.
%ONDICHERY
i 9$I$)$M$;$6$ ii 'ur"eillane Centre, 3o"t$
3eneral Hos%ital, )ondiherry$
iii De%tt$ of Mirobiology, 9I)M;6,
90
)ondiherry$
$F
.
%UNKAB
i Medial College, 2#ritsar$ i" De%tt$ of Mirobiology, 3o"t$
Medial College, 2#ritsar$
ii Medial College, )atiala$
iii Medial College, -arid.ot$
$G
.
RAKASTHAN
i '$M$'$ Medial College, 9ai%ur$ "i De%tt$ of Mirobiology, '$M$'$
Hos%ital, 9ai%ur$
ii '$)$ Medial College, Bi.aner$
iii 6$8$T$ Medial College, <dai%ur$
i" Dr$ '$8$ Medial College, <dai%ur$
" 9$L$8$ Medial College, 27#er$
$5
.
TAMIL NADU
i 3o"t$ Medial College, Chennai / De%tt$ of Mirobiology, Instt$ of
Child Health and Hos%ital for
Children, Madras$
ii 'tanley Medial College, Chennai$ /i De%tt$ of Mirobiology, Madurai
Medial College, Madurai$
iii :il%au. Medial College, Chennai$ /ii 'ur"eillane Centre, Medial
College, Chennai$
i" 'ri 6a#ahandra Medial College
and 6esearh Institute, Chennai$
" Than7abur Medial College,
Than7abur$
"i Medial College, Coi#batore$
"ii Madurai Medial College, Madurai
"iii Tirune"eli Medial College,
Tirune"eli$
i/ Medial College, Chingle%ut$
$4
.
UTTAR %RADESH
i '$8$ Medial College, 2gra$ i/ De%tt$ of Mirobiology, :$3$
Medial College, Lu.no,$
ii M$L$8$ Medial College, 2llahabad$ / 'ur"eillane Centre, Central
92LM2 Instt$ for Le%rosy, 2gra
iii 9$8$ Medial College, 2ligarh$ /i De%tt$ of Mirobiology, Instt$ of
Medial 'ienes, Varanasi$
i" Institute of Medial 'iene, BH<,
Varanasi$
/ii De%tt$ of Mirobiology, 9$L$8$
Medial College, 2ligarh$
" 3$'$V$M$ Medial College, :an%ur$ /iii 'ur"eillane Centre, :a#la
8ehru College, 2llahabad$
"i M$I$B$ Medial College, Lu.no,$
"ii :$3$ Medial College, Lu.no,$
"iii B$6$D$ Medial College, 3ora.h%ur$
.I
.
WEST BENGAL
i 3o"t$ Medial College, Calutta$ "ii 'ur"eillane Centre, 8ational
Institute of Hygiene and )ubli
Health, Calutta$
ii 6$3$ :ar Medial College, Calutta$
91
iii 8$6$'$ Medial College, Calutta$
i" 8ational Medial College, Calutta$
" B$'$ Medial College, Ban.ura$
"i 8orth Bengal Medial College,
'iliguri$
.$
.
CHANDIGARH
i )$3$I$ of Medial ;duation and
6esearh, Chandigarh$
ii De%tt$ of I##uno%athology,
)3I, Chandigarh$
..
.
MANI%UR
i 6egional Medial College, I#%hal$ ii 'ur"eillane Centre, 9$8$
Hos%ital, I#%hal$
.=
.
MEGHALAYA
i 'ur"eillane Centre, Ci"il
Hos%ital, 'hillong$
.>
.
MILORAM
i 'ur"eillane Centre, Ci"il
Hos%ital, 2iD,al$
.?
.
NAGALAND
i 'ur"eillane Centre, 8aga
Hos%ital, :ohi#a$
ii 'ur"eillane Centre, Distt$
Hos%ital, Di#a%ur$
.F
.
ARUNACHAL %RADESH
i 'ur"eillane Centre, Distt$
Hos%ital, Itanagar$
.G
.
SIKKIM
i 'ur"eillane Centre, '$T$8$M$
Hos%ital, 3angto.$
.5
.
TRI%URA
i 'ur"eillane Centre, Distt$
Hos%ital, 2gartala$
92
.4
.
ANDAMAN AND NICOBAR
ISLANDS
i 'ur"eillane Centre, 3$B$
Hos%ital, )ort Blair$
=I
.
LAKSHADWEE%
i 'ur"eillane Centre,3o"t$
Hos%ital, :a"aratti$
=$
.
DADRA AND NAGAR HAVELI
=.
.
DAMAN AND DIU
93
$I. QUALITY ASSURANCE %ROGRAMME
$I.$ N","ss-(9 '* -16)r('," : The diagnosti tests to detet antibodies to
HIV ha"e sensiti"ity and s%eifiity ,hih are not absolute$ In all these
tests ,e ha"e false negati"e results as ,ell as false %ositi"e results ,hih
are inherent and annot be a"oided$ The %erentage of false %ositi"e
results ,ill inrease as the %re"alene rate of %ersons ,ith HIV antibodies
in a %o%ulation dereases$ Te!e t.o 5ro=lem! are com5o6nded =+ te
fact tat d6rin# !creenin# 5ro#ramme!, la=oratorie! .ill rarel+ =e a=le to
5erform to te level of acc6rac+ tat te te!t! are tecnicall+ ca5a=le of
acievin#) Thus, the "alidity of diagnosti test results is de%endent to a
"ery large e/tent on the +uality of the tehnial onditions under ,hih
the tests are %erfor#ed$ Meaning thereby, onsistent %rodution of
reliable results re+uires a stringent o"erall assurane %rogra##e ,hih
,ould ontrol tehnial onditions before, during and after eah assay$
The Quality 2ssurane )rogra##e ensures that the final results
re%orted by the laboratory are orret, reliable and aurate as far as
%ossible$ This %rogra##e o"ersees re%orting results in a ti#ely #anner
and to the a%%ro%riate indi"idual$ It also ensures use of the #ost reliable
tests for the diagnosis of HIV infetion$ It is de%endent on a good
Q+0-(9 C)'(r)0 %r)2r11" and its effiay #ay be "erified by a good
Q+0-(9 Ass"ss1"'( %r)2r11".
i* Q+0-(9 ,)'(r)0 6r)2r11" : This %rogra##e inludes #easures ,hih
are introdued during eah assay to "erify the "alidity and reliability of
the test$ Ho,e"er, this does not indiate that the results generated are
aurate &,hih, indeed, is the harateristi of the test*$ It also does
not indiate that the results ha"e been re%orted ti#ely, %ro%erly and
orretly$
ii* EA("r'0 C+0-(9 ss"ss1"'( 6r)2r11" : This is a #eans to
deter#ine the +uality of results$ It is an e/ternal e"aluation of a
laboratory %erfor#ane by inor%orating %rofiieny %anels as the #eans
to e"aluation$ F)r 0/)r()r9 () /" ,)'s-*"r"* r"s6",("*
("s(-'2 3,-0-(9, -( 1+s( /" 0/)r()r9 (h( ,' 089s 6r)*+,"
,,+r(", r"0-/0" '* r"6r)*+,-/0" r"s+0(s.
$I.. G+-*"0-'"s () -16r)!" C+0-(9 )3 ("s(-'2
i* C)'*-(-)' )3 (h" s6",-1"'s
2ll s%ei#ens #ust be ins%eted at the ti#e of reei"ing and also before
testing to ensure that they are suitable$ <se of li%ae#i, hae#olysed and
onta#inated sera should be a"oided$ If they ha"e to be used, the
re%orting offier should #ention on the re%ort that the result of the test
#ay not be "alid beause of the ondition of the seru# sa#%les$ 2 ne,
s%ei#en should be re+uested for re%eat testing$
2ll s%ei#ens should be %ro%erly labelled before ae%tane$ The
label should inlude follo,ing infor#ation : a* na#e of the %atient, b*
olletion date, * %atientCs identifiation nu#ber if a%%liable$ ;ah
s%ei#en #ust be ao#%anied ,ith a test re+uest for# ,hih should
inlude age and se/ of the %atient, na#e of the %hysiian re+uesting the
in"estigation, ris. grou% of the %atient, reason for the in"estigation in
addition to the infor#ation gi"en on the label$ In ase of unlin.ed
anony#ous testing, only ode nu#ber and olletion date #ay be
#entioned on the label as ,ell as on the re+uest for#$
94
If the seru# or %las#a sa#%le is froDen before testing, it is
essential that after tha,ing at roo# te#%erature!@H
>
C, the sa#%le
should be ins%eted for any lot or floating %artiles$ The sa#%le should
be larified before testing$ The sa#%le should be ,ellA#i/ed before
testing$
)resently, all the test .its distributed by the 82C(, are #eant for
testing seru# or %las#a only$ Therefore, these .its #ay not reliably
detet %resene of antibodies in other body fluids$
ii* Q+0-(9 )3 B-(s '* "C+-61"'(s : The test .its #ust be used ,ithin the
e/%iration date stated on the .it to ensure "alid results$
;"ery bath of the .it should be tested and ertified for its effiay
&sensiti"ity and s%eifiity* before distribution &see ha%ter 00*$
;LI'2 reader, ,ashers, inubators, %i%ettes should be he.ed
regularly for their o%ti#al %erfor#ane$ They need to be alibrated at
least annually$
iii* C)'(r)0s +s"* -' (h" ("s(s : ;ah test run re+uires a set of
ontrols to "alidate the results$ These ontrols #ust be treated in the
sa#e #anner as un.no,n sa#%les$ They are run si#ultaneously and
under the sa#e onditions as the un.no,n sa#%les$ <%on o#%letion of
the test, the results of the ontrols and the sa#%les are e/a#ined using
the sa#e riteria for inter%retation$ The assay is "alid and the results are
reliable ,hen the ontrols %rodue ae%table results$
There are t,o ty%es of ontrols ,hih #ust be inluded in eah run :
) I'("r'0 ,)'(r)0s : These ontrols &%ositi"e and negati"e* are inluded
in eah HIVAtest .it by the #anufaturer and are to be inluded in eah
test run$ These are essential for +uality ontrol #easures for eah run$
They are intended for use ,ith the sa#e lot nu#ber of the .it in ,hih
they ha"e been %a.ed$ Internal .it ontrols are generally ad7usted by
the #anufaturer so that an e/%eted range of "alues are obtained ,ith
eah lot of .its$ To a"oid onsiderable flutuations in the (D "alues due
to "ariable oating of the antigen on the solid surfae during
#anufaturing, the #anufaturer, artifiially stabiliDes .it ontrols to gi"e
the sa#e "alues as the %re"ious lot$
/) EA("r'0 ,)'(r)0s : These should be inluded ,ith eah testArun to
#onitor onsistent %erfor#ane and lot to lot "ariation ,hih annot be
deteted using internal ontrols for the reasons #entioned abo"e$ These
ontrols are #ade fro# %ooled test .it ontrols or #ade fro# %ooled sera
fro# HIVA%ositi"e or negati"e indi"iduals in eah laboratory &inAhouse
ontrols*$ Ho,e"er, for #onitoring the %erfor#ane of other laboratories,
the seru# sa#%les for e/ternal ontrols are dra,n in suffiient +uantities
to last for at least 0= #onths &ha%ter 00*$
The #ost i#%ortant e/ternal ontrol to inlude is a borderline
reator$ This ontrol ,ould indiate any #inor hange s%eially around
ut off "alue beause suh hanges ,ould effet the results of un.no,n
sa#%les ,ith (D "alues near the ut off$ It #ay ta.e se"eral #onths to
establish ranges for e/ternal ontrols$ 8onetheless, this ty%e of ontrol
syste# an be "ery effeti"e in hel%ing to identify %otential %roble#s and
inauraies in the laboratory setting$
,) S('*r*-E-()' )3 (h" C+0-(9 )3 "A("r'0 ,)'(r)0s : 6e%roduibility
95
and +uality of internal and e/ternal ontrols #ust be standardiDed by
-'(r;r+' r"6r)*+,-/-0-(9 and -'("r;r+' r"6r)*+,-/-0-(9$ The ontrol
sa#%les &either internal or e/ternal* are tested at least three ti#es on
the sa#e testArun$ This ,ill indiate intraArun re%roduibility$ This
e/erise is then arried out on testAruns on three onseuti"e days to
deter#ine interArun re%roduibility$ By either of these #ethods "ariations
should not e/eed 0>F$ These seru# sa#%les #ay also be e"aluated at
the 8ational 6eferene Centers$
i"* I'("r6r"((-)' )3 *( : The test .it inserts arry instrutions to
establish range of internal ontrols and to define the outliers$ 'ine the
e/ternal ontrols are inluded in eah test run to #onitor onsistent
%erfor#ane of the test .it, it is neessary to deter#ine the li#it of
ae%tability statistially by alulating arith#eti #ean, standard
de"iation and error, oeffiient of "ariation et$ Ho,e"er, the "alues for
the internal ontrols, e/ternal ontrols and the ut off an be #onitored
easily by +uality ontrol gra%hs$ This is beause the gra%hi %resentation
of the ontrol "alues o"er ti#e ,ill #a.e subtle hanges in ontrols #ore
easily disernible$
2n aurate #ethod of gra%hially re%resenting the "alues of
ontrol of eah test run is to %lot (D!utAoff &C(* ratio on LAa/is$ In this
#ethod the ontrol "alues are e/%ressed relati"e to the utAoff "alue$
This is i#%ortant as the (D as ,ell as C( "alues ,ill hange slightly
bet,een testAruns$ Therefore, the ontrols should be o#%ared ,ith the
res%eti"e alulated C( "alue$
T,o ty%es of hanges an be obser"ed in the +uality ontrol gra%hs$
These are$
) Sh-3(s : 5hen ontrol "alues of si/ onseuti"e test runs fall on one side
of the #ean, it is alled shift$ This indiates a #a7or hange in the testA
%erfor#ane due to i* s,ithing to a ne, lot of .its, ii* ne, reagents, iii*
hanges in inubation te#%erature, i"* hange of %i%ettes, "* a ne,
tehniian, et$
/) Tr"'*s : 5hen ontrol "alues of si/ onseuti"e tests are distributed in
one general diretion, it is alled trend$ This is generally due to i*
deterioration of reagents, and ii* a routinely used %i%ette slo,ly losing its
alibration$
!) C0,+0(-)' )3 2r"9;E)'" r",()rs : During routine testing, #any
sa#%les #ay ha"e slightly ele"ated (D "alues 7ust abo"e C( "alue ,hih
#ay suggest %resene of lo, antibody ati"ity$ 'uh sa#%les are alled
2r"9;E)'" r",()rs )r /)r*"r0-'" r",()rs$ This reati"ity #ay be
due to a* early seroon"ersionJ b* "ery lo, antibody reati"ity %resent in
the seru#J e* false %ositi"e reati"ity$ Therefore, it is "ery essential to
re%eat any HIV reati"e result ha"ing (D "alues greater than or e+ual to
the utAoff "alue$ 2ny re%eatedly %ositi"e result is then "alidated by a
#ore s%eifi su%%le#ental test li.e 5estern blot$
'o#eti#es, due to a tehnial error a seru# ,ith lo, antibody
reati"ity #ay gi"e an (D "alue 7ust belo, the utAoff "alue$ Therefore,
#any laboratories ad"oate a re%eat HIV test on all the sa#%les ,hih
gi"e an (D "alue of 0>F belo, the utAoff$ 2lthough this is "ery
arbitrary, this a%%roah inreases the hane of finding so#e early HIV
seroon"erison$
!-) A,,+r,9 )3 E'E91" I11+')ss9 (EIA) : 'ine no assay an be
96
0>>F sensiti"e and 0>>F s%eifi, it is li.ely to gi"e false negati"e as
,ell as false %ositi"e results$
2ny ;I2 #ay be 30s" '"2(-!" under follo,ing onditions :
) B-)0)2-,0 C)'*-(-)'s :
i$ The test #ay be false negati"e during early stages of HIV infetion$ This
%eriod generally "aries fro# =A=E ,ee.s but #ay be as long as E=
#onths$ the false negati"e rate "aries fro# 0?F to G>F during this
%eriod$ &-arDadegan et al, 9$Clin$ Mirobiol =H: 011=, 0B1B*
ii$ In so#e %atients there is an early %rodution of antibodies follo,ed by a
essation of antibody %rodution and disa%%earane of antibodies fro#
blood$ This #ay be due to se+uestration of the "irus$
/) T",h'-,0 ,)'*-(-)'s :
i$ The test .it generally has sensiti"ity ranging fro# BB$EF to nearly 0>>F$
Ho,e"er, if %erfor#ed under less o%ti#al onditions the sensiti"ity #ay
dro% onsiderably$
The ;I2 #ay be 30s" 6)s-(-!" under follo,ing onditions :
) B-)0)2-,0 ,)'*-(-)'s :
i$ Certain onditions inrease false %ositi"ity rates of ;LI'2$ These are
hroni aloholis#, %arenteral drug abuse, hae#odialysis, #ulti%le
%regnanies and #ulti%le blood transfusions$
ii$ -alse %ositi"ity rises if the %re"alene of HIV antibodies in the %o%ulation
dereases$
/) T",h'-,0 ,)'*-(-)'s :
i* -alse %ositi"e rates "ary fro# one ;LI'2 .it to another$ ;"en the sa#e
ty%e of .it #ay sho, a signifiant "ariation in s%eifiity fro# bath to
bath$
ii* Hu#an error, e$g$, s%ei#en #i/ing #ay aount for false %ositi"ity$
iii* Laboratories ,ith less than o%ti#u# %erfor#ane also aount for higher
false %ositi"ity rates$
!--) R"6)r(-'2 )3 r"s+0(s : <nli.e any other test re%ort, the HIVAtest re%orts
#ust be handled ,ith are$ 'ine HIV infeted indi"iduals #ay fae
soial stig#a, i#%ro%er re%orting #ay so#eti#es lead to e#otional
brea.do,n of an indi"idual or e"en suiides$ Therefore, before re%orting
the result of HIVAantibody test it #ust be borne in #ind that:
a* The results of any sreening test &;6'* are only %resu#%ti"e and should
not be re%orted$ These #ust be "alidated by a su%%le#ental test$
b* In bloodAban.ing only donated blood is sreened by a single ;6' as %er
reo##endations of 3o"t$ of India$ Therefore, these results should not
be used to identify the indi"iudals$
* In serosur"eillane studies sine the %ositi"e results of first ;6' are
"alidated by a seond ;6' and not by a su%%le#ental test, these results
#ay not be used to identify the asy#%to#ati indi"iduals$
d* -or #a.ing a diagnosis of HIVAinfetion, the reo##endations of the
3o"t$ of India &ha%ter B* #ust be follo,ed$
e* HIV test results should be re%orted to the %hysiian ,ho has re+uested
the test$ 2ll HIV testing, in ,hih an indi"idual is identified, #ust be
%reeeded and follo,ed by %reAtest and %ostAtest ounselling res%eti"ely$
97
f* 2ll test results #ust be .e%t onfidential and should ne"er be disussed in
%ubli$ The test results should ne"er be o##uniated on tele%hone$
$I.= %r)3-,-"',9 ("s(-'2 )3 0/)r()r9 : )rofiieny testing is synony#ous
,ith ;/ternal Quality 2ssess#ent &;Q2*$ 6eently, 8ational 2ID' Control
(rganisation has for#ulated a Quality 2ssess#ent )rogra##e$ <nder
this %rogra##e oded %anels of .no,n HIV antibody %ositi"e and
negati"e sera are %ro"ided to the %artii%ating laboratories for HIV
%rofiieny testing on a regular basis$ These %anels are %roessed and
tested ,ithin a s%eified %eriod so that results fro# all the %artii%ating
laboratories #ay be analysed together$ The %artii%ating laboratories are
re+uested to handle these oded %anels in e/atly the sa#e #anner as
the the routine sa#%les for HIV testing in ;LI'2$ This is beause the
%ur%ose of suh a %rogra##e is to identify the %roble#s in HIV testing$
5hile analyDing the results and %re%aring a o#%rehensi"e re%ort,
are is ta.en not to identify any of the %artii%ating laboratory either by
na#e or by loation$ These re%orts are then trans#itted to the
%artii%ating laboratories ,here the laboratory inAharge #ay assess the
ade+uay of +uality assurane and +uality ontrol #easures in his o,n
laboratory and o#%are the %erfor#ane of his laboratory ,ith other
laboratories$ In ase there is a %roble# he #ay in"estigate, identify and
ta.e a%%ro%riate ti#ely ation to retify the %roble#$
'ine e"ery laboratory as%ires to be a res%eted testing faility it
#ust initiate a good and effeti"e +uality assurane!+uality ontrol
&Q2!QC* %rogra##e$ ;ffeti"eness of suh a %rogra##e #ay be
assessed by an e/ternal +uality assess#ent$ Ho,e"er, a good Q2!QC
%rogra##e annot be substituted by +uality assess#ent %rogra##e, but
a good Q2!QC %rogra##e #ay substitute for +uality assess#ent$
Many a ti#e false negati"e and false %ositi"e results #ay be due
to tehnial errors es%eially if the tests are %erfor#ed under less o%ti#al
onditions$ Therefore, it is neessary to #onitor the %erfor#ane of the
laboratory ,or.ers %eriodially by inor%orating the sa#%les ,ith .no,n
results ,ith the routine sa#%les ,ithout the .no,ledge of the laboratory
,or.ers$ This is done to ensure the auray, reliability and
re%roduibility of the test results re%orted by the laboratory$
$I.> R",)r* B""6-'2 :
a* 2 laboratory #ust #aintain a log boo. for reording of the laboratory
s%ei#ens$ The infor#ation ontained in the log boo. should be .e%t
onfidential$
b* 2 ,or. sheet &see anne/ure* ontaining the identifiation nu#bers of
sera to be tested #ust be %re%ared eah ti#e before the test run is
%erfor#ed$
* Daily reords of te#%erature of ,ater baths, inubators, refrigerators and
freeDers should be #aintained$
d* Miro%i%ettes should be alibrated atleast annually$
e* ;LI'2 readers should be alibrated to ensure auray of their readings$
f* The laboratory should #aintain a file ,here all %roedures!%a.age
inserts are .e%t for ready referene$
g* 8u#bers of ;LI'2 %ositi"e sa#%les being re%orted by eah laboratory
fro# "arious high ris. grou%, #oderate ris. grou% and lo, ris. grou%
should be #aintained$
h* 8u#ber of ;LI'2 %ositi"e sa#%les ,hih ha"e been found negati"e by
5estern blot assay should be #aintained$
i* 8u#ber of ti#es an ;LI'2 %ositi"e seru# sa#%le ,as rerun before
98
sub7eting it to su%%le#ental test should be #aintained$
These data ,ill reflet u%on the tehnial %roble#s if any e/isting in a
laboratory$
$I.? L/)r()r9 s6",(s :
$I.?.$S6",-1"' ,)00",(-)' '* 6r),"ss-'2 : -i"e #l blood is olleted by
"ene%unture using either a dis%osable %lasti syringe or an autola"ed
glass syringe$ 2fter olleton of the sa#%le the dis%osable syringe is
%ro%erly disarded, ,hereas the glass syringe is .e%t in a bea.er
ontaining li+uid bleah disinfetant for 0> to 0G #in before
resteriliDation$ 'ine li%ae#i, hae#olysed and onta#inated seru#
sa#%les do not yield reliable results, are should be ta.en to ollet
blood, fasting or after a light brea.fast, in a dry sterile ontainer$
Hae#olysis an be a"oided by a* not lea"ing the tourni+uet on the ar#
for #ore than 0 #inJ b* not forefully transferring the blood through a
needle to a tube$ <ni"ersal %reautions inluding ,earing glo"es #ust be
follo,ed ,hile dra,ing the blood$ It is "ery i#%ortant to %re"ent infetion
being %assed fro# an indi"idual to the laboraroty ,or.er and "ieA"ersa$
Labelling of the sa#%le for %ro%er identifiation of the indi"idual is
e/tre#ely i#%ortant to a"oid inorret re%orting of HIV status$
Therefore, the tube #ust be labelled %ro%erly i##ediately u%on olletion
of blood sa#%le$ The sa#%le is allo,ed to sit at @H
>
C for @> #in and
then E
>
C for lot retration$
Thereafter, the sa#%le is s%un at 0G>> r%# at E
>
C and the seru#
is olleted ase%tially in a sterile ontainer$ 8o %reser"ati"e is added$ In
ase it is not tested for HIV antibodies, it #ay be stored at E
>
C for 0A0>
days or at A @>
>
C for long ter# storage$ 6e%eated freeDing and tha,ing
should be a"oided$
$I.?..%r"6r(-)' )3 (h" "A("r'0 ,)'(r)0s :
%)s-(-!" '* '"2(-!" ,)'(r)0s : (ne unit of blood fro# eah donor,
tested negati"e for he%atitis B surfae antigen, but tested &i* %ositi"e for
HIV antibody by ;LI'2 and 5estern blot, or &ii* negati"e for HIV antibody
&out dated blood sa#%le*, or &iii* %ositi"e for HIV antibody by ;LI'2 but
negati"e by 5estern blot assay &-alse %ositi"e result* is olleted fro#
the blood ban.$ The %las#a is se%arated, heat inati"ated at G?
>
C for @>
#in and then realified$ It is then inubated at @H
>
C for lotting to
our$ The lot is s%un do,n and the seru# is se%arated ase%tially$ It is
filtered through a #illi%ore filter &>$= u# or =>>n# %ore siDe* to re#o"e
any baterial onta#ination$ 8o %reser"ati"e is added$ It is ali+uoted,
labelled %ro%erly and stored at AE>
>
C$ (ne tha,ed, the ali+uot is stored
at =
>
C to 1
>
C$ It is disarded after using it one$ HIV %ositi"e seru#
sa#%les should not be disarded$ These should be sent to national
referene entres for long ter# storage in seru# ban. at AE>
>
C$
$I.?.=B)r*"r0-'" )r 0)8 6)s-(-!" ,)'(r)0 : Ten fold serial dilutions &0>
A0
to
0>
A?
* of a .no,n %ositi"e seru# are %re%ared in the seru# fro# HIV
negati"e donor ,hih is used as a diluent in %lae of nor#al saline or
buffered salt solution to .ee% the %roteins in a natural en"iron#ent$ ;ah
dilution is assayed in se/tu%liates for HIV antibodies$ The #ean (D of
eah dilution and 'D are alulated$ 2 gra%h ,ith (D of eah 0> fold
dilution is then %lotted$ The highest dilution of the seru# gi"ing %ositi"e
result in ;LI'2 is ta.en as an end %oint$ Three dilutions, "iD$, end %oint
dilution T0 'D, end %ont dilution and end %oint dilution A 0 'D are tested
by ;LI'2 in se/tu%liates and also by 5B assay$ If these dilutions gi"e
99
re%roduible results &intraArun "ariability*, they are tested on si/
onseuti"e days for interArun "ariability$
Care #ust be ta.en not to selet a sa#%le ,ith so lo, antibody
titer that the (D flutuates abo"e and belo, the ut off due to nor#al
"ariations$ 2lso the sa#%le ,ith a "ery high antibody titer should not be
seleted$ It ,ill be of li#ited use for borderline #onitoring$
$I.?.>R",0,-3-,(-)' )3 60s1 : Ideal #aterial for +uality ontrol and
+uality assess#ent is seru#$ This is beause %las#a tends to be
unstable on long storage and #ay lot s%ontaneously during trans%ort$
'ine under #any iru#stanes blood olleted fro# a blood ban. #ay
be the only soure for HIV %ositi"e and HIV negati"e sa#%les, %las#a
#ust be realified to %rodue seru#$ Ho,e"er, realifiation should be
done ,ith are beause e/essi"e realifiation #ay affet so#e assays
li.e gelatin %artile agglutination, ad"ersely$ (nly itrated blood should
be realified$
a* Ma.e a $IM R",0,-3-,(-)' s)0+(-)' &CaC0=$?H=( GGg, MgCl=$ ?H=(
0?g, in 0>> #l distilled ,aterJ autola"e at 0=0
>
C for => #in*$
b* 2dd 0$G #l of realifiation solution to 0 unit of blood or =G> #l of
%las#a ,hih has been brought to roo# te#%erature$
* Inubate at @H
>
C for @>A?> #in &until lot for#ation*$
d* :ee% at E
>
C o"ernight$
e* '%in at 0G>> r%# for => #in$
f* 'e%arate the seru# ase%tially$
g* Label and store in s#all ali+uots at A E>
>
C$
h* Test s#all ali+uot fro# eah sa#%le for HIV antibodies by ;6' tests and
5estern blot assay$
$I.?.?%r"s"r!(-!"s 3)r s"r+1 s160"s : 2lthough 'eitD filters #ay be
used to re#o"e bateria and fungi fro# the seru# sa#%les, filtration of
realified seru# is often diffiult as filters tend to log due to fibrin lots$
Centrifugation ,ould re#o"e lots by and large before filtration$ <se of
%refilters ,ill also hel% in larifying the seru# sa#%les$
;"en though one #ay start ,ith sterile seru# sa#%le, subse+uent
use and #ani%ulation in the laboratory #ay easily onta#inate it$
Therefore, addition of Bronido/ L &GAbro#oAGnitroA0,@Adio/ane in
%ro%ylene glyol, Hen.el Che#ials* to the seru# sa#%le to a final
onentration of >$>GF #ay hel% in %re"enting gro,th of onta#inants$
Thio#ersal &#eruri hloride* is generaly used in a final onentration of
>$>0F but it is effeti"e only for a fe, ,ee.s$ This is beause it loses its
ati"ity es%eially ,hen e/%osed to light$ 'odiu# aDide should not be
used as it inati"ates %ero/idase on7ugate$
$I.?.FS()r2" )3 s"r+1 : 'eru# sto.s are best stored at A E>
>
C or belo, in
s#all ali+uots of GA0> #l$ 6e%eated freeDing and tha,ing should be
a"oided$ (ne tha,ed, it should be further ali+uoted in s#aller siDe and
.e%t at E
>
C until used$
$I.?.GUs" )3 3r""E";*r-"* ,)'(r)0s : -reeDe drying of a seru# sa#%le is an
effeti"e #ethod for storage of seru# sa#%le at =A1
>
C for a "ery long
ti#e$ In addition, the shi%#ent of freeDe dried sa#%le is "ery si#%le and
straight for,ard$ Ho,e"er, during the %roess of freeDe drying, antibody
ati"ity es%eially of the borderline sa#%les #ay be lost$
$I.?.5%r"6r(-)' )3 6r)3-,-"',9 6'"0s : 2 %rofiieny %anel onsisting of
t,o strong %ositi"es, t,o negati"e and t,o borderline %ositi"es sera
100
sa#%les is %re%ared fro# the ali+uots of e/ternal ontrols stored at AE>
>
C
and oded$ If %ossible, re%resentati"e seru# sa#%les fro# different
areas of the ountry #ay be inluded in the %anel$
$I.?.4Tr,B-'2 (h" 6"r3)r1'," )3 (h" B-( +'*"r 3-"0* ,)'*-(-)'s : ;ah
laboratory %erfor#ing ;LI'2 should #aintain the follo,ing infor#ation to
ensure good %erfor#ane of the .it$
i* (D "alues of internal and e/ternal %ositi"e ontrols of eah test run
ii* (D "alues of internal and e/ternal negati"e ontrols of eah test run
iii* (D "alues of e/ternal borderline %ositi"e ontrols of eah test run
i"* Cut off "alue of eah test run
"* 8a#e, bath and lot nu#ber of the .it used$
"i* ;/%iry date of the .it used$
"ii* Date of the test run$
This infor#ation ,ill hel% to .ee% a tra. of the %erfor#ane of the .it
under the field onditions$
$I.?.$I Q+0-(9 ,)'(r)0 )3 HIV '(-/)*9 ("s( B-(s :
-) M)'-()r-'2 )3 *-33"r"'( 0)(s )3 HIV K-( : 'o#eti#es a #anufaturer
%rodues a reagentAlot that %asses +uality ontrol re+uire#ent in his unit
but fails to %erfor# ade+uately in the field$ This ould be due to se"eral
fators inluding artifiial stabiliDation of the ontrols by the
#anufaturer, substandard storage and shi%%ing onditions et$
Therefore, it is i#%erati"e that these lots of reagents #ust be identified
+ui.ly before distribution to the sur"eillane entres$ This is done by a
tehni+ue alled )arallel testing in ,hih %erfor#ane of ne, lots of .it
,ith the %re"ious lots "ia a o##on ontrol #aterial &e/ternal ontrols
and the ontrols fro# %re"ious lots* is o#%ared$ If all ontrols %rodue
e/%eted results, the ne, lot has %assed the %arallel tests and #ay be
used for routine testing$
--) M)'-()r-'2 )3 *-33"r"'( ELISA K-(s : There are four #ain ob7eti"es of
HIVAantibody sreening %rogra##e, na#ely :
a* 'afe blood ban.ing
b* 'ero sur"eillane
* 'ero diagnosis
d* 6esearh
-or safe blood ban.ing, it is i#%erati"e that the ;LI'2 or any other test
.it should orretly identify all antibody %ositi"e sera$ In other ,ords the
test .it should ha"e 0>>F sensti"ity$ The sensiti"tiy is generally
e/%ressed in ter#s of %erentage ,hih is a +ualitati"e #easure$ 2
+uantitati"e e/%ression of sensiti"ity in ter#s of %ositi"e delta "alue
&delta T* is better and #ore reliable than %erentage "alue$ This hel%s in
seleting a#ong ;LI'2 .its ,ith e+ual sensiti"ity$ The higher the
deltaT"alue, the higher is the %robability that this test ,ill orretly
identify antibody %ositi"e sera$
-or seroAdiagnosis and seroAsur"eillane, an ;LI'2 .it ,ith a high
sensiti"ity and high s%eifiity is needed$
2 negati"e delta "alue &deltaA* is +uantitati"e e/%ression of
s%eifiity$ The greater the negati"e delta &deltaA* "alue, the higher is
the %ossibility that this assay ,ill orretly identify the true negati"e sera$
101
---) C0,+0(-)' )3 D"0( V0+"s :
a* To alulate the delta "alues, G> onfir#ed %ositi"e and G> onfir#ed
negati"e sera are tested$
b* Cut off "alue is alulated as suggested by the #anufaturer$
* Mean (D and statandard de"iation of %ositi"e and negati"e sa#%les is
alulated$
d* Mean (D!C( ratio of %ositi"e and negati"e sa#%les is alulate by the
follo,ing for#ula
#ean (D of the sa#%les
(D!C( ratio R AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA
#ean C( "alue
Delta "alue is alulated by the follo,ing for#ula :
Mean (D!C( ratio of %ositi"e sa#%les &log 0>*
Delta T R
AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA
'tandard de"iation of %ositi"e sa#%les
Mean (D!C( ratio of negati"e sa#%les &log 0>*
Delta A R
AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA
'tandard de"iation of %ositi"e sa#%les
$I.F E!0+(-)' )3 I'*-r",( I11+')30+)r"s"'," Ass9 (IFA) s
s+660"1"'(0 ("s( : 2lthough the 5estern Blot &5B* 2ssay has the
ad"antage of being able to indiate atual antibodies that reat ,ith the
s%eifi HIV antigens, it is "ery e/%ensi"e and ti#e onsu#ing$
'o#eti#es the results are diffiult to inter%ret$ In addition, it is "ery
#uh tehni+ue de%endent$
(n the other hand, the I-2 offers se"eral ad"antages o"er the 5B assay$
It is easy to %erfor#, #uh less e/%ensi"e than 5B and an be
o#%leted in a short %eriod of ti#e$ The test is easy to read but re+uires
an e/%ertise to do that$
Therefore, I-2 #ay be used as a su%%le#ental test in %arallel ,ith 5B
assay$ I-2 .its are o##erially a"ailable$
102
Ch6("r $I ; A''"A+r" $
STRUCTURE OF THE QUALITY ASSESSMENT %ROGRAMME
6egional 6eferene Center &5H(*
8ational HIV!2ID' 6eferene Centers
6egional and or 'tate HIV!2ID'
6eferene Center
Voluntary blood
Testing Laboratory
HIV 'reening
Laboratory
&Blood Ban.s*
HIV 'reening
Laboratory
&'ur"eillane Centers*
103
Ch6("r $I ; A''"A+r" .
QUALITY ASSESSMENT %ROGRAMME
L/)r()r9 E'r)001"'( F)r1
0$ 8a#e of the Laboratory
=$ Mailing 2ddress
@$ Tele%hone 8u#ber ;/tn$
E$ Tele/ 8u#ber
G$ -a/ 8u#ber
?$ Laboratory Inharge &8a#e*
H$ Laboratory Inharge &Title*
1$ I ,ish to enrol in the +uality assess#ent %rogra##e of the 8ational
6eferene Centre!6egional 6eferene Centre under 82C($
Y"s & N)
'ignature of the Inharge
)lease #ail the o#%leted for# to the onerned 8ational and ! or 6egional
6eferene Centre$
104
Ch6("r $I ; A''"A+r" =
%r)3)r1 3)r s"'*-'2 '(-;HIV ("s( r"s+0(s /9 6r(-,-6(-'2 0/)r()r9 +'*"r
QA%
ENJ:ME IMMUNOASSA: *ESULTS
Please record t:e a7sor7ance 9al5es @col58n AA and t:e a44ro4riate c5to66 9al5e
@col58n 2A and calc5late t:e sa84le;c5to66 ratios @A2A 6or eac: 4anel sa84le.
Also record <:et:er t:e ser58 is regarded as 4ositi9e or negati9e 7y t:e assay.
5lea!e circle one; SC*EENING SU11LEMENTAL
Sa84le
code
A7sor7ance
9al5es
@AA
C5to66
9al5e
@2A
@AA @2A
3 deci8al
4laces
eg. #.3"$10.$67
Test
inter4retatio
n
eg. negati9e
1
#
3
"
$
6
7
E
D
10
Co88ercial Na8e o6 Assay:
Na8e o6 )an56act5rer :
Assay Hit )aster ,ot No. : '!4iry Date:
Assay Conditions : Te84erat5re @
0
CA Ti8e @:r: 8inA
Sa84le Inc57ation:
Conj5gate Inc57ation:
S57strate Inc57ation :
Circle t:e inc57ation condition t:at a44lies. T:e 8icro4late <as inc57ated:3
1. In a <ater 7at:. #. In an inc57ator. 3. On a 7enc:. ". In a dedicated 8ac:ine.
Ot:er @gi9e detailsA
Na8e o6 O4erator : Date :
Comment! ;
105
Ch6("r $I ;A''"A+r" >
%r)3)r1 3)r s"'*-'2 '(-;HIV ("s( r"s+0(s /9 6r(-,-6(-'2 0/)r()r9 +'*"r QA%
RA%ID ASSAY (%ARTICLE AGGLUTINATION&S%OT&DOT BLOT&COMB) RESULTS
Please record i6 a reaction is o7ser9ed <it: 5nsensitised 4articles 6or eac: ser58
@see XControlY col58nA. I6 yo5r la7oratory titrates 4ositi9e sera 4lease record t:e
titre. Note1 :o<e9er t:at t:is is not essential1 75t o4tional. or8at 6or aggl5tination
assay is gi9en.
5lea!e circle one; SC*EENING SU11LEMENTAL
Sa84le
code
Cnsensitised
4articles
@ControlA
Sensitised
4articles
@TestA
Titre
@O4tionalA
Positi9e (e8ar=
1
#
3
"
$
6
7
E
D
10
Co88ercial Na8e o6 Assay:
Na8e o6 )an56act5rer:
Assay Hit )aster ,ot No.: '!4iry Date
Assay Conditions: Te84erat5re @
0
CA Ti8e @:r: 8inA
Sa84le Inc57ation:
Circle t:e inc57ation condition t:at a44lies. T:e 8icro4late <as inc57ated:3
1. In a <ater 7at: #. In an inc57ator. 3. On a 7enc:
Ot:er @gi9e detailsA
Na8e o6 O4erator: Date:
Co88ents:
106
Ch6("r $I ; A''"A+r" ?
%r)3)r1 3)r s"'*-'2 s"r+1 6'"0 ("s( r"s+0(s ('(-;HIV) /9 r"3"r"'," ,"'(r" () (h"
6r(-,-6(-'2 0/)r()r9 +'*"r QA%
Table 0 : Details of sa#%les fro# %anel
Panel
n587e
r
Dil5tion Sa84le ty4e Sa84le origin Anti3+I&31
stat5s
+I& #0 Cndil5ted sa84le Plas8a1
lyo4:ilised
NICD @7lood
donorsA
I
+I& 31 Dil5ted 1:#0"E as a7o9e as a7o9e N
+I& "6 Dil5ted 1:1#E as a7o9e as a7o9e I
+I& ED Dil5ted 1:16 as a7o9e as a7o9e P
+I&
1#0
Dil5ted 1:E as a7o9e as a7o9e P
+I&
131
Dil5ted 1:" as a7o9e as a7o9e P
Table = : 6eferene 5B!LI2 and sreening assay test results
Sam5le
-e!tern =lot and or line imm6noa!!a+ re!6lt!
Blot8 *a5id
n6m=er 5%
@
5"
(
5<
%
5<
(
5<' 5(& #5(%
$('
5'< 5'
'
5?
@
LIA te!t ELISA
+I& #0 R R;
3
3 3 3 3 3 3 3 3 I N 0.70
+I& 31 R;
3
3 3 3 3 3 3 3 3 3 N N 0.6#
+I& "6 R;
3
R;
3
3 3 3 3 3 3 3 3 I ( 0.66
+I& ED R;
3
R;
3
3 3 3 3 R 3 3 R;
3
P ( 1.#E
+I&
1#0
R;
3
R 3 3 3 3 R 3 3 R P ( #.0D
+I&
131
R R 3 3 3 3 RR 3 3 R P ( 3.$#
5estern Blot
Line I##unoassay
HIV dot blot
Ca%illus &late/ agglutination*
I##unoo#b
) : %ositi"e
8 : negati"e
6 : reati"e
I : indeter#inate
107
%%) E0ALUATION OF HI0 IITS AND 1*E1A*ATION OF SE*UM 1ANEL
%%)% Introd6ction : Periodic e9al5ation o6 +I& test =its is a 4rereB5isite o6 a
good B5ality 8anage8ent. A large n587er o6 assays are a9aila7le
co88ercially eac: o66ering essential as <ell as attracti9e 4er6or8ance
c:aracteristics na8ely sensiti9ity1 s4eci6icity1 e66iciency and 4redicti9e
9al5es. +o<e9er1 t:e 4er6or8ance o6 t:e test <ill de4end 54on t:e
conditions in t:e la7oratory1 tec:nical e!4ertise1 4o45lation 7eing
tested etc. So1 <:ere9er 4ossi7le1 e9al5ation o6 =its s:o5ld 7e done
5nder t:e sa8e conditions and in t:e sa8e locale 5sing t:e local 4anel
o6 sera to assess t:e 4er6or8ance c:aracteristics and e66iciency o6 a
test =it. Ne< and i84ro9ed =its are also 7eing de9elo4ed contin5o5sly
and t:ese also need e9al5ation in t:e sa8e <ay as t:e esta7lis:ed =its.
(e4orts o6 e9al5ation o6 =its 4ro9ide a 6eed 7ac= to t:e 8an56act5rers
to i84ro9e t:eir 4rod5cts so t:at t:e =its 8ay 7e 6it 6or 5se 5nder
di66erent sit5ations and in di66erent geogra4:ical areas. To :a9e an idea
a7o5t t:e 4er6or8ance c:aracteristics o6 t:ese +I& =its1 4rior
e9al5ation 7y so8e noti6ied agency s:o5ld 7e 8ade and is 8andatory.
Sit5ations 5nder <:ic: t:ese =its 8ay :a9e to 7e e9al5ated can 7e as
6ollo<s :
3 Hits 5nder i84ort F e9al5ation <it: local ser58 4anel is 9ery i84ortant.
3 Ne< =its de9elo4ed;8an56act5red indigeno5slyF to assess e66icacy.
3 To resol9e contro9ersies regarding discordant res5lts or deterioration
o6 co84onents and reagents. T:is 8ay 7e d5e to 7rea= in cold c:ain
d5ring transit and;or storage o6 in35se =its
As 6ar as t:e 6irst category is concerned Dr5gs Controller %eneral o6
India s:o5ld as= t:e i84orter to get t:e re4resentati9e sa84le o6 =its
e9al5ated1 5sing a locally 8ade and <ell c:aracterised 4anel 7y t:e
Instit5tes identi6ied 6or t:is 45r4ose. I6 6o5nd s5ita7le1 only t:en t:e
=its s:o5ld 7e allo<ed to 7e i84orted 6or distri75tion1 7ot: in
%o9ern8ent as <ell as 4ri9ate sector.
In second case1 <:en t:e =its are 8an56act5red indigeno5sly1 t:e
4er6or8ance c:aracteristics o6 t:e =its s:o5ld 7e ascertained 7y
e9al5ating di66erent 7atc:es o6 =its @t:e 6inal 4rod5ctA <it: t:e
standard ser58 4anel 6or e9al5ation o6 =its at least at t<o to t:ree
di66erent geogra4:ical sites and consistent res5lts s:o5ld 7e o7tained.
Only t:e =its 4er6or8ing to t:e standards @sensiti9ity and s4eci6icity
etc.A as laid do<n s:o5ld 7e licensed 6or 8ar=eting.
In t:e t:ird case1 t:e licensed1 and;or a44ro9ed =it0s 4er6or8ance 8ay
7e B5estioned 4artic5larly <:en di66erent centres re4ort inconsistent
res5lts. T:is can :a44en d5e to 7rea= in cold c:ain d5ring trans4ort
and storage o6 +I& =its. To resol9e t:is 4ro7le8 t:e =its :a9e to 7e
e9al5ated 7y t:e identi6ied centres to ens5re o4ti8al 4er6or8ance.
T:is c:a4ter gi9es t:e 7asic g5idelines to 7e 6ollo<ed to ens5re
4ro4er e9al5ation o6 +I& test =its.
%%)" 1re5aration of 5anel ; A ser58 4anel is de6ined as a collection o6 <ell
c:aracterised ser58 sa84les 6or a s4eci6ic re6erence;45r4ose.
Ser58 4anel can 7e 4re4ared and 5sed 6or di66erent 45r4oses.
T:ese are :
3 Panel 6or e9al5ation o6 =its i.e. e9al5ation 4anel incl5ding
serocon9ersion 4anel
108
3 Panel 6or 4ro6iciency testing @e!ternal B5ality ass5ranceA
3 Panel 6or B5ality control in t:e la7oratory @internal B5ality ass5ranceA
T:ese 4anels are 5sed 6or =it e9al5ation1 e!ternal B5ality control
assess8ent and 6or internal B5ality control in +I& testing la7oratories.
%%)")% 0ol6me of 5anel !am5le! ; T:e 9ol58e o6 ser58 7eing 4rocessed to
4re4are 4anel s:o5ld 7e s566icient to 8eet t:e reB5ire8ents 6or at least
#33 years 6or t:e 45r4ose 6or <:ic: t:e 4anel is 4re4ared. or e!a84le
68l o6 ser58 <ill 7e s566icient to e9al5ate a44ro!i8ately 100 =its.
%%)")" Collection of !5ecimen for 5anel ; Ideally 8aterial 5sed 6or 4anel s:o5ld 7e
ser58 alt:o5g: 4las8a can also 7e 5sed. +o<e9er1 4las8a tends to 7e
5ns5ita7le as it clots s4ontaneo5sly on storage. T:ere6ore 4las8a
s:o5ld 7e de6i7rinised <it: t:ro87in or recalci6ied and t:en 4rocessed
6or 8a=ing t:e 4anel @re6er to c:a4ter 10 6or recalci6ication 4roced5reA.
%%)")< AliC6otin# of !er6m ; (e4eat 6reeJing and t:a<ing o6 s4eci8ens a66ects
t:e B5ality o6 t:e 4anel. To a9oid t:is sa84les s:o5ld 7e aliB5oted in
9ol58es t:at 8ay 8eet t:e 4artic5lar 4anel reB5ire8ent at one ti8e.
S5ita7le anti37acterial agents e.g. 2ronido! 0.0$Q @$32ro8o3$3Nitro3
113 Dio!ane1 Sig8a c:e8icalsA s:o5ld 7e added to 4anel sa84les to
4re9ent conta8ination. T:e sol9ent 6or 2ronido! is 4ro4ylene glycol.
%%)")( Caracteri!ation of !er6m 5anel ; Ser58 sa84les s:o5ld 7e <ell
c:aracterised so t:at t:ere s:o5ld not 7e any do57t a7o5t t:e 6inal
o5tco8e o6 t:e test res5lts. S4eci8ens s:o5ld 7e tested 5sing :ig:
B5ality +I& anti7ody ',ISA or any ot:er screening assay 6ollo<ed 7y
con6ir8atory .estern 2lot assay to deter8ine t:eir tr5e stat5s and
s:o5ld also disting5is: 7et<een +I&31 and +I&3# ty4es. As
c:aracterisation o6 eac: 5nit in t:e 4anel is B5ite e!4ensi9e1 5se o6
large 9ol58e o6 eac: s4eci8en <ill 7e 8ore cost e66ecti9e.
%%)")' Stora#e ;AliB5ots 85st 7e stored at or 7elo<370
0
C. %lass 9ials s:o5ld
not 7e 5sed as t:ey 8ay e9ent5ally crac= <it: long ter8 storage in
6roJen condition.
%%)")? Tran!5ort of 5anel ;To 4ro9ide sa6ety 6or la7oratory sta661 co5riers and
co885nity1 s4eci8ens s:o5ld 7e trans4orted 5nder International Air
Trans4ort Association @IATAA reg5lation1 6or t:is 45r4ose. S4eci8ens
s:o5ld 7e 4ac=ed1 la7eled and doc58ented as classed 6.# CN #E1" to
8ini8ise t:e ris= o6 in6ection to anyone d5ring transit.
%%)< Com5o!ition of !er6m 5anel ; T:ese are <ell c:aracterised ser58
sa84les1 t:ose <:ic: :a9e gi9en consistent res5lts <it: all t:e
a9aila7le +I& test =its. T:ese sera s:o5ld 7e 6ro8 indi9id5als in <:o8
t:e tr5e +I& stat5s @in6ected or not in6ectedA is =no<n <it: :ig: degree
o6 certainty. S5c: sa84les are called >4edigreed sera.? Panel 85st
incl5de reacti9e1 nonreacti9e1 <ea=ly reacti9e @grey JoneA and
indeter8inate classi6ied sera. 2ot: +I&31 and +I&3# reacti9e sera
s:o5ld 7e incl5ded in t:e 4anel. T:e 4anel s:o5ld 7e re4resentati9e o6
t:e 4o45lation;area <:ere t:e =its are going to 7e 5sed.
To 4re4are a local 4anel1 sera s:o5ld 7e tested <it: all t:e a9aila7le
=its.
3 At least $0 s4eci8ens s:o5ld 7e incl5ded in eac: 4anel o6 reacti9e1
<ea=ly reacti9e1 indeter8inate and non reacti9e. T:e 4anel siJe can 7e
109
increased as 8ore t:e n587er o6 sera in eac: 4anel1 8ore statistically
9alid is t:e co84arison.
3 Non reacti9e 4anel s:o5ld also incl5de re4resentati9e sera 6ro8
indi9id5als s566ering 6ro8 local ende8ic diseases @e.g. 8alaria1
t57erc5losis1 :y4erga88aglo75linae8ia etc.A.
3 Panel s4eci8ens s:o5ld 7e a44ro4riately 4rocessed;c:aracterised and
stored as aliB5ots in 6roJen condition. All s4eci8ens s:o5ld 7e in t:e
sa8e condition as 6ar as storage is concerned <:en test 6or e9al5ation
o6 =its is 4er6or8ed.
3 S4eci8ens s:o5ld 7e non:ae8olysed1 nonli4ae8ic and nont5r7id etc.
%%)( *eference te!t ; A <ell esta7lis:ed1 re6erence gold standard test 85st 7e
a9aila7le to t:e la7oratory 6or co84arison1 t:e res5lts o6 <:ic: s:o5ld
correlate <it: t:e act5al stat5s o6 t:e 4atient. C5rrently1 s5c: a test in
serologic testing 6or +I& in6ections is considered to 7e .estern 7lot
@.2A test. +o<e9er1 so8eti8es .2 8ay gi9e indeter8inate res5lts in
<:ic: case 8ore so4:isticated tests li=e synt:etic 4e4tide 7ased ,ine
I885noassay1 4oly8erase c:ain reaction @PC(A and 9ir5s c5lt5re can
7e 5nderta=en. T:e tests are e!4ensi9e1 la7o5r intensi9e and reB5ire
:ig:er le9el o6 tec:nical e!4ertise. T:5s 6or t:e 8o8ent .2 5s5ally is
regarded as t:e (e6erene;%old standard test. .:en a n587er o6 =its
are 7eing e9al5ated t:e tie 7rea=er 6or 6alse 4ositi9e;6alse negati9e is
5s5ally a .2 test1 so8eti8es s544orted 7y 8ore so4:isticated tests as
8entioned a7o9e.
%%)' Te!tin# condition! ; All e9al5ations 85st 7e 4er6or8ed 5nder identical
conditions. T:e tests 85st 7e cond5cted in t:e sa8e la7oratory 5sing
sa8e eB5i48ents @4i4ettes1 instr58ents etc.A1 7y t:e sa8e tec:nician1
5sing =its <:ic: :a9e not e!4ired and :a9e 7een stored 4ro4erly and
5sing t:e e!act 4rotocols 4ro9ided in t:e 4ac=age insert.
T:e tec:nicians 85st not =no< t:e res5lts o7tained on t:e 4anel
<it: ot:er =its i.e. t:e e9al5ation testing is done 7lindly.
T:e 4ara8eters to 7e calc5lated to assess t:e 4er6or8ance
c:aracteristics o6 t:e =its incl5de :
3 Sensiti9ity
3 S4eci6icity
3 '66iciency
3 Delta 9al5es
3 Positi9e 4redicti9e 9al5e
3 Negati9e 4redicti9e 9al5e
%%)')% Sen!itivit+ : Sensiti9ity o6 a test is de6ined as its a7ility to detect tr5ly
in6ected indi9id5als as also its a7ility to detect 9ery s8all a8o5nts o6
analyte. Sensiti9ity can 7e calc5lated 7y t:e 6ollo<ing 6or85la :
Tr5e 4ositi9es
Sensiti9ity U 3333333333333333333333333333333333333333333 S 100
Tr5e 4ositi9es R alse negati9es
or
N587er o6 sa84les detected as 4ositi9es 7y t:e =it 5nder
e9al5ation
Sensiti9ity U
33333333333333333333333333333333333333333333333333333333333333333333333333333333333 S 100
Total n587er o6 4ositi9es 7y t:e re6erence test e84loyed
'!a84le :
110
Total sera tested U 100
+I& in6ected U 10
+I& nonin6ected U D0
(es5lts <it: t:e
re6erence test
(es5lts <it: =it 5nder e9al5ation : D o5t 10 +I& in6ected detected as 4ositi9e
D
Sen!itivit+ of te 3it 6nder eval6ation .ill eC6al to $$$$$$$$$$$$$ S 100 U D0Q
DR1
%%)')" S5ecificit+ : T:is is t:e a7ility o6 an assay to correctly identi6y all t:e
5nin6ected indi9id5als i.e. t:ere s:o5ld 7e no 6alse 4ositi9es @reacti9e res5lt in
t:e a7sence o6 diseaseA.
T:e s4eci6icity can 7e calc5lated 7y t:e 6ollo<ing 6or85la :
Tr5e negati9es
S4eci6icity U 333333333333333333333333333333333333 S 100
Tr5e negati9es R alse 4ositi9es
or
N587er detected as negati9es 7y t:e =it 5nder e9al5ation
S4eci6icity U
33333333333333333333333333333333333333333333333333333333333333333333333 S 100
Total n587er o6 negati9es detected 7y re6erence test e84loyed
'!a84le :
Total sera tested U 100
+I& in6ected U 10
+I& nonin6ected U D0
(es5lts o7tained
<it: re6erence test
T:e res5lts o7tained <it: =it 5nder e9al5ation1 D 4ositi9e o5t o6 10 +I&
in6ected and 1 4ositi9e o5t o6 D0 +I& nonin6ected.
ED
S5ecificit+ of te 3it 6nder eval6ation .ill eC6al to 33333333S 100 U DE.DQ
@a44ro!i8atelyA
EDR1
%%)')< Efficienc+ : It is t:e o9erall a7ility o6 a test to correctly identi6y all 4ositi9es
as 4ositi9e and all negati9es as negati9e. '66iciency is deter8ined as 7elo< :
Tr5e 4ositi9es R Tr5e negati9es
'66iciency U
333333333333333333333333333333333333333333333333333333333333333333333333333333 S 100
Tr5e 4ositi9es R alse negati9es R Tr5e negati9es R alse
4ositi9e
or
Total n587er detected as 4ositi9es R as negati9es 7y t:e =it
5nder e9al5ation
'66iciency U
33333333333333333333333333333333333333333333333333333333333333333333333333333333333 S 100
Total n587er o6 4ositi9es R negati9es detected 7y re6erence test
e84loyed
'!a84le :
Total sera tested U 100
111
+I& in6ected U 10
+I& nonin6ected U D0
(es5lts o7tained
<it: re6erence test
T:e =it 7eing e9al5ated detected D o5t o6 10 +I& in6ected as 4ositi9e
and one o5t D0 nonin6ected also as 4ositi9e.
DR ED
So efficienc+ of te 3it .ill eC6al to 33333333333333333 S 100 U DEQ
DR1REDR1
%%)')( Delta val6e ; T:is is a statistical 8eas5re o6 sensiti9ity and s4eci6icity and is
calc5lated 6ro8 t:e act5al OD 9al5es generated 7y t:e test. T:is also :el4s to
deter8ine :o< close t:e O.D. 9al5e o6 6alse 4ositi9e and or 6alse negati9e is to
t:e real 4ositi9e and or real negati9e 9al5e1 res4ecti9ely. I6 t:ese 9al5es are
too close t:e test is 6la<ed.
Delta 9al5es are de6ined as t:e distance o6 t:e 8ean O.D. ratio o6 t:e
sa84le 4o45lation 6ro8 t:e c5t o66 @ C.O.A 8eas5red in standard de9iation
5nits.
)ean O.D.; C5t o66 ratio @log 10A
Delta 9al5es U 33333333333333333333333333333333333333333333333333 S 100
Standard de9iation
Positi9e delta 9al5e is calc5lated on 8ean o6 ODs o6 4ositi9e sa84les
and negati9e delta 9al5e is calc5lated on 8ean ODs o6 negati9e ser58
sa84les. .:en t<o tests are 7eing co84ared1 t:e test t:at :as a :ig:er
4ositi9e delta 9al5e <o5ld c:aracterise 4ositi9e sa84les 8ore acc5ratelyF
si8ilarly t:e test <it: :ig:er negati9e delta 9al5e <ill 7etter classi6y t:e
negati9e sa84les.
'!a84le :
Positi9e sa84les Negati9e sa84les
O.D.;C.O. ".#0 .03
".60 .0#
$."0 .0"
)ean O.D.;C.O. ".73 .03
Standard de9iation 0.61 .01
,og10 o6 t:e 8ean
O.D.;C.O.
0.67 31.$#
Delta
9al5es
Positi9es delta 9al5e
R1.10
Negati9e delta 9al5e
31$#
%%)')' 1redictive val6e! ; T:is is t:e 8eas5re o6 9al5e o6 a test in relation to t:e
4re9alence o6 t:e disease in t:e 4o45lation. T:e 9al5e o6 a test in addition to
t:e 4ara8eters descri7ed a7o9e de4ends on t:e 4o45lation 7eing tested.
Positi9e 4redicti9e 9al5e @PP&A is t:e a7ility o6 t:e test to identi6y
act5ally in6ected indi9id5als a8ong all 4ersons gi9ing a 4ositi9e res5lt <it:
t:e =it 7eing e9al5ated.
Tr5e 4ositi9es
PP& U 33333333333333333333333333333333333333333333333333 S 100
Tr5e 4ositi9es R alse 4ositi9es
112
Negati9e 4redicti9e 9al5e @NP&A is t:e a7ility o6 t:e test to correctly
identi6y t:e really non in6ected indi9id5als 6ro8 a8ong all t:e 4ersons gi9ing
negati9e res5lt.
Tr5e negati9es
NP& U 33333333333333333333333333333333333333333333333333 S 100
Tr5e negati9es R alse negati9es
I6 t:e 4re9alence is :ig: t:e PP& o6 a test <ill 7e :ig: i.e. an indi9id5al
testing 4ositi9e <ill 8ean real in6ection. .:ereas1 in lo< 4re9alence area t:e
c:ance t:at an indi9id5al testing 4ositi9e is really in6ected is lo<er.
'!a84le : ,o< 4re9alence area 1Q
Total sera tested U
1000
+I& in6ected U 10
+I& nonin6ected U
DD0
<it: t:e re6erence test
(es5lts o7tained <it: =it 5nder e9al5ation : Positi9e res5lts o7tained in
E indi9id5als1 t<o 6ro8 +I& in6ected gro54 and 6 6alse 4ositi9es 6ro8 +I&
nonin6ected gro54.
Negati9e res5lts o7tained in DD# indi9id5als1 DE" 6ro8 t:e nonin6ected gro54
and 6 6alse negati9es 6ro8 +I& in6ected gro54.
#
T:e PP& U 3333333 S 100 U #$Q
E
DE"
T:e NP& U 33333333333 S 100 U DD.3Q
DD#
'!a84le : +ig: Pre9alence area 10Q
Total sera tested U 1000
+I& in6ected U 100 <it: t:e
+I& nonin6ected U D00 re6erence test
(es5lts o7tained <it: +I& =it 5nder e9al5ation F 10$ tested 4ositi9e1
D$ 6ro8 +I& in6ected gro54 and 10 6ro8 +I& non in6ected gro54 i.e. ga9e 6alse
4ositi9e res5lt. Negati9e res5lt <as o7tained in a total o6 D00 cases. ED#
o6 D00 <ere 6ro8 +I& nonin6ected gro54 and E <ere 6ro8 +I& in6ected gro54.
D$
PP& U 333333333333333 S 100 U D0.$Q @a44ro!A
10$
ED#
NP& U 333333333333333 S 100 U DD.1#Q
D00
All t:e a7o9e 4ara8eters are ascertained 7y t:e la7oratory carrying o5t
t:e e9al5ation o6 =its. T:e 4ara8eter0s 9al5es s:o5ld 7e <it:in t:ose
reco88ended 7y t:e tec:nical e!4ert co88ittee1 %.O.I. on t:is s57ject. T:e
6inal a44ro9al o6 t:e test =it is gi9en 7y DC%I on t:e 7asis o6 t:e e9al5ation
re4orts s578itted 7y t:e identi6ied e9al5ation centres.
113
$.. LABORATORY TESTS FOR MONITORING STAGE AND %ROGRESSION OF HIV
INFECTION
%")% Introd6ction ; In6ection <it: t:e :58an i885node6iciency 9ir5s @+I&A
8ay de9elo4 to acB5ired i885ne de6iciency syndro8e @AIDSA at
di66erent rates in di66erent indi9id5als1 <it: a s4ectr58 9arying 6ro8
ra4id 4rogression to long ter8 non34rogression. T:is 9aria7ility
8a=es it essential to :a9e tests <:ic: can acc5rately assess t:e
stage o6 in6ection in an indi9id5al1 as <ell as 4redict its co5rse and
8onitor its 4rogression. T:ese la7oratory tests are 9ery 9al5a7le
d5ring t:e 4eriod o6 clinical latency1 and1 s57seB5ently1 s544le8ent
9ario5s clinical 4ara8eters1 <:ic: are also e!tre8ely i84ortant in
categoriJing t:e in6ection ; disease stage. (es4onse to anti3retro9iral
t:era4y is also 8onitored 5sing t:ese 4rognostic tests.
$.... L/)r()r9 ("s(s : The laboratory tests used for #onitoring stage and
%rogression of HIV infetion an be lassified into:
Viral #ar.ers
A )las#a HIV 682 load
A Viral antigenae#ia e$g$ %=E antigen
'urrogate #ar.ers
A Virus s%eifi #ar.ers e$g$ antibodies to HIV antigens &e$g$ %=E, %0H,
nef, et$*
8onAs%eifi #ar.ers
A Cellular #ar.ers e$g$ CDE ell ount, %henoty%i #ar.ers of
ly#%hoyte ati"ation$
A 'oluble #ar.ers of i##une ati"ation e$g$ neo%terin, beta =
#iroglobulin and so#e yto.ines and their ree%tors in
seru#$
The basis of #any of these laboratory tests is that HIV indues i##une
syste# ati"ation, beginning in the early stages, ,hih %rogresses throughout
the ourse of HIV infetion$ This an be deteted by hanges both in seru#
le"els of soluble i##une ati"ation %roduts and in se"eral %henoty%i
#ar.ers of ly#%hoyte ati"ation$ 2lthough suh i##une ati"ation is usually
benefiial to the host in other infetions, it an be deleterious in infetion ,ith
HIV$ Ly#%hoyte ati"ation triggers HIV re%liation, releases "irus fro#
ellular reser"oirs and ontributes to disease %athogenesis$ The inreased rate
of HIV re%liation is refleted in an inrease in %las#a "iral 682 load, ,hih is
%resently onsidered the #ost re%resentati"e and sensiti"e laboratory test for
#onitoring %rogression of HIV infetion$ Inreased HIV re%liation leads to a
de%letion of CDE ells, ,hih are its #ain target$ CDE ell ount, a "ery useful
surrogate #ar.er, ,as one of the earliest to be desribed and is still used for
staging infetion and #onitoring its %rogression$
114
$.. =. V-r0 1rB"rs :
$..=.$. %0s1 HIV RNA 0)* : )las#a "iral load &HIV 682* +uantifiation is
%resently onsidered the best #ethod for #onitoring %rogression and
res%onse to antiAretro"iral thera%y$ 2ti"e re%liation of "irus ours in all
linial stages of infetion$ It is %ossible to detet and +uantify "irus
throughout the ourse of HIV infetion$ The "iral load usually ranges bet,een
0>
=
and 0>
H
HIV 682 o%ies!#L in untreated indi"iduals though it #ay be
lo,er in those on treat#ent$
The tehni+ues a"ailable for +uantifying "iral 682 are:

A Quantitati"e 682A)C6
A Branhed D82 assay
A 8ulei aid se+uene based a#%lifiation
M"(h)*s '* 6r-',-60"s
$..=.$.$.Q+'(-((-!" RNA %CR (A160-,)r HIV;$ M)'-()r T"s(, R),h"): This
test is based on %arallel a#%lifiation of HIV 682 &0E= b% se+uene fro# gag
gene* fro# the sa#%le and a .no,n a#ount of +uantitation standard &Q'*
682, using 6TA)C6 &re"erse transri%tase A %oly#erase hain reation* ,ith
the sa#e set of biotinAlabeled %ri#ers to gi"e different %roduts &a#%lions*$
These are deteted in se%arate ,ells of a #iro,ell %late by hybridiDation ,ith
i##obiliDed HIV and Q' s%eifi %robes$ The bound biotinylated a#%lion is
+uantified olori#etrially by #easuring the o%tial density &(D*, obtained by
using a"idinAhorse radish %ero/idase &H6)* on7ugate and an a%%ro%riate
substrate$ The HIV 682 o%y nu#ber an be then alulated by e/tra%olating
fro# the .no,n in%ut nu#ber of o%ies of Q' 682$ To o"er a ,hole range of
HIV 682 o%ies %ossible in %las#a of infeted %ersons, a nu#ber of dilutions
are %ut u% &-ig$0=$0*$
$..=.$...Br',h"* DNA ss9 (Q+'(-60"A /;DNA, Ch-r)') : The branhed
D82 assay is based on a#%lifiation of the signal instead of the target$ 2
target %robe o#%le#entary to both the target se+uene and the a%ture
%robe &i##obiliDed on the solid %hase* hybridiDes ,ith both of these$
2nother se+uene on the target %robe &no, attahed to the solid %hase
through the target* then hybridiDes ,ith an Ua#%lifierU branhed D82 &diretly
or "ia an inter#ediate e/tender %robe*$ 2n enDy#eAlabeled %robe, ,hih an
bind to the #ulti%le branhes of the a#%lifier branhed D82, is allo,ed to
hybridiDe ,ith it$ This a#%lified label is deteted by a lu#ino#eter and
+uantified by its ation on a he#ilu#inisent substrate$ The +uantity of the
target in the sa#%le an be deter#ined fro# this &-ig$0=$=*$
$..=.$.=.N+,0"-, ,-* s"C+"'," /s"* 160-3-,(-)' (NASBA, Or2')'
T"B'-B) : This tehni+ue uses @ enDy#es A re"erse transri%tase &2MVA6T*,
682se H and TH 682 %oly#erase A at a single te#%erature &@H
o
C*, to
generate #ulti%le o%ies of the target HIV 682, by a strategy si#ilar to that
used by retro"iruses for their re%liation$ 6T enDy#e and a %ri#er
&o#%le#entary to HIV 682 target* designed ,ith the TH 682 %oly#erase
%ro#oter se+uene at one end, are utiliDed for synthesis of an 682AD82
hybrid ontaining the TH %ro#oter fro# the target$ -ro# this, the original
682 target strand is re#o"ed by addition of 682se H and dsD82 is
generated by the D82 %oly#erase ati"ity of 6T and a seond %ri#er$ Multi%le
o%ies of 682 are then generated by TH 682 %oly#erase enDy#e$ This 682
enters another si#ilar yle and the a#ount of 682 #ulti%lies e/%onentially
,ith eah yle &-ig$0=$@*$ 5ith a%%ro%riate ontrols and standards it is
%ossible to +uantitate the original 682 target by this #ethod$
%r"s"'( s((+s )3 HIV RNA 0)* : )las#a HIV 682 le"els are #ore than
115
0>Afold lo,er in longAter# nonA%rogressors &#ean "alue: H>,>>> o%ies!#L*
than in indi"iduals ,ith %rogressi"e disease &#ean "alue: 0,>>>,>>>
o%ies!#L*$ 2 "iral load of P0>>,>>> o%y e+ui"alents!#L by bAD82 assay,
,ithin ? #onths of seroon"ersion inreases the ris. of %rogression to 2ID'
,ithin G years by #ore than 0>Afold$ )ersistently detetable "irae#ia and
high baseline le"els arry a %oor %rognosis, ,hile ris. of %rogression is lo, at
o%y nu#bers O 0>,>>>!#L$ 2%art fro# being an e/tre#ely useful %reditor
of %rogression, "iral load res%onds "ery ,ell to antiretro"iral thera%y and an
be used to #onitor it$
The "ariation bet,een different #ethods for HIV 682 load +uantitation ,as
bet,een >$>1 and >$=> log0> by o#%arati"e e"aluation in a #ultiAentri
study, ,ith a greater "ariation ,ith linial sa#%les than ,ith s%i.ed sa#%les$
The use of a o##on set of HIVA0 682 standards eli#inates differenes in
absolute HIVA0 682 o%y nu#ber$
The bAD82 assay had less %roble#s ,ith detetion of different HIVA0
subty%es as it assesses a larger %ortion of the HIV se+uene using #ulti%le
%robes$ The +uantitati"e 6TA)C6 .its gi"e #ore aurate results in the hands
of tehniians e/%eriened in )C6, as all the %reautions nor#ally ta.en for
any )C6 based test are essential to follo,$
It is reo##ended that blood sa#%les should be olleted in ;DT2 &rather
than 2CD or he%arin* and that %las#a is better than seru# &and usually
sho,s higher "alues*$ )las#a should %referably be se%arated on the sa#e
day and stored at MH>
o
C &%las#a is stable at E
o
C for u% to G days and at A=>
o
C
for u% to => ,ee.s*$ The "alue of "iral 682 load &as also #ost other #ar.ers*
in #onitoring %aediatri %atients needs further "alidation$
$..=... 6.> '(-2"'"1- : HIV gag gene enoded ore %rotein antigen, %=E, is
one of the earliest "iral antigens detetable in the blood after infetion$ It is
useful for diagnosis during the ,indo, %eriod during early infetion and in
the ne,born$ Le"els of free antigen deline after the a%%earane of antiA%=E
antibodies and for#ation of i##une o#%le/es$ 2n inrease in free %=E le"els
is a %reditor of %rogression$
M"(h)* '* 6r-',-60": ;LI'2 %=E antigen a%ture assay .its are a"ailable
for detetion of %=E antigen$ To inrease sensiti"ity, #ost .its %resently used
for diagnosis inlude i##une o#%le/ dissoiation &ICD* at aidi %H
&%retreat#ent ,ith dilute HCL or glyine, follo,ed by neutraliDation* as a %art
of the %rotool of %=E detetion, to inrease sensiti"ity$ Quantitation for
#onitoring %=E is %ossible by testing serial dilutions of the %las#a!seru#$
'ensiti"ity of detetion is bet,een 0> to G> %g!#L$

%r"s"'( s((+s )3 ("s(s 3)r 6.> '(-2"'"1- : <ntil the a"ailability of
tests for HIV 682 load, it ,as the #ain "iral load assay a"ailable$ The antigen
is %oorly +uantifiable, #ay not be detetable in #any indi"iduals and does not
hange "ery ,ell ,ith antiAretro"iral thera%y$ It has a li#ited role in diagnosis
&in s%eifi situations* and is not onsidered as useful a %rognosti #ar.er
no,$
$..>. S+rr)2(" 1rB"rs :
V-r+s s6",-3-, 1rB"rs : 2 "iral harateristi assoiated ,ith %rogression is
the on"ersion of strains fro# a nonAsynytiu# induing &8'I* %henoty%e to a
synytiu#Ainduing &'I* %henoty%e$ This hange of harater has no, been
assoiated ,ith the ability to infet ells ,ith ertain he#o.ine oree%tors$
HIV s%eifi antibodies ha"e also been used as #ar.ers of %rogression,
though they are not onsidered %artiularly sensiti"e$ Deline in or absene of
116
antibodies to "arious HIV antigens inluding %=E, %0H, g%0=>, g%E0 and nef
gene %rodut ha"e been used as surrogate #ar.ers in the %ast$
N)' HIV;s6",-3-, 1rB"rs : Infetion ,ith HIVA0 has a dra#ati effet on
the i##une syste#: the loss of CDE ells and the ati"ation of the i##une
syste#, both of these effets are thought to %lay a role in the de"elo%#ent
of linial 2ID'$ I##une sti#ulation is refleted by
hy%erga##aglobulinae#ia, e/%ansion of #aro%hage and inrease in
&ati"ated* CD1 ells$ It also leads to a hange in the le"els of soluble
#ar.ers of i##une ati"ation and so#e yto.ines and their ree%tors$ CDE
ell nu#bers are dereased in HIV infetion but CD@ &total T ell* nu#ber
re#ain onstant$
2 nu#ber of non HIVAs%eifi ellular #ar.ers ha"e been used for staging,
#onitoring %rogression of HIV infetion and assessing res%onse to thera%y$
(ne of the #ost useful and o##only used ellular #ar.ers is the absolute
ount &or %erentage* of CDET ly#%hoytes in the blood$ (ther ly#%hoyte
%henoty%i #ar.ers assoiated ,ith %rogression inlude an inrease in
indiators of i##une ati"ation on T ly#%hoytes li.e CD@1 &es%eially on
CD1T ly#%hoytes*, HL2AD6, ILA=6, CDEG6( and #ar.ers of a%o%tosis &e$g$
-as*$ 2 flo, yto#eter ,ith at least P= olour analysis faility is a neessary
re+uire#ent for these #ar.ers$
$..>.$. CD> ,)+'( : The #ost o##only used ellular #ar.er is the CDE
ly#%hoyte ount$ Its deline is the hall#ar. of HIV infetion and the rate of
loss in eah %erson is uni+ue$ CDE ell nu#ber hanges during HIV infetion
in the follo,ing stages :
A 6a%id deline for ?A01 #onths at the ti#e of seroon"ersion
A )lateau of gradual deline that an last at least se"eral years during the
asy#%to#ati %eriod
A 'tee%er deline for se"eral #onths 7ust before 2ID' de"elo%s
A Continued CDE ell deline until death$

CDE ell ount is e/tre#ely i#%ortant in the staging of HIV infetion and a
re"ised lassifiation of the Center for Disease Control, 2tlanta, <'2 &0BB@*
di"ides HIV %ositi"e %ersons into three CDE ount ategories : 0R PG>>!LJ
=R =>>AEBB!LJ @R O=>>!L &along ,ith @ %arallel linial stagesA 2, B and
C*$ 2 lo, CDE ount & less than 0>F*, a nu#ber less than 0>>!L and a lo,
CDE!CD1 ratio &O>$=* are highly %rediti"e for death fro# 2ID' Arelated
o#%liations$ It an su%%le#ent "iral load as a #ar.er of %rogression$ 2
%ersistently high "iral load is %rediti"e of a %oor %rognosis, es%eially ,hen
ao#%anied by a "ery lo, CDE ount$
Ho,e"er, the linial benefit of antiretro"iral drugs is not ,ell e/%lained by
CDE ounts &unli.e "iral 682 load* and it is less effeti"e in #onitoring
thera%y$ It is e"en less li.ely to be of use in e"aluation of i##unothera%y$
'ustained inreases in CDE ounts ,ith ne,er #ultidrug regi#ens are #ore
effeti"e in %rediting a %ositi"e res%onse to thera%y than the transient
inreases ahie"ed by earlier regi#ens$
M"(h)*s '* 6r-',-60"s :
$..>.$.$. F0)8 ,9()1"(r9 : 'ignals are generated by ells or %artiles in
sus%ension %assing through a light &usually laser* soure in the flo, ell, in a
single file &aligned by the sheath fluidi syste#*, and are analyDed
eletronially in a flo, yto#eter$ The %ara#eters #easurable inlude for,ard
satter & an indiator of ell siDe*, side satter &an indiator of granularity of
the ell* and signals fro# #ulti%le fluoresent dyes &e$g$ -ITC, %hyoerythrin*
117
tagged to ell surfae %henoty%i #ar.erAs%eifi antibodies$
The ,hole blood lysis #ethod is %referred for analysis of ly#%hoyte
%o%ulations$ Blood is ideally olleted in ;DT2 and %roessed on the sa#e day
&also for he#atologial tests*$
A 2 "olu#e of buffer &e$g$ about G>L of )B', at roo# te#%erature* is
dis%ensed into tubes$
A This is follo,ed by the addition of reo##ended "olu#es &usually 0> L for
#ost o##erial %re%arations* of eah fluoresent antibody &de%ending on
the o#bination to be used* and G> to 0>> L of ,hole fresh blood$

A 2fter gentle "orte/ing, the tubes are inubated at roo# te#%erature for 0>A
=> #inutes$
A This is follo,ed by lysis of 6BCs and fi/ation of ells &singleAste% or as
se%arate ste%s ,ith ,ashing and entrifugation in bet,een, de%ending on the
reagents used*$ If o##erial reagents are not used then a##oniu# hloride
solution an be used for lysis and %arafor#aldehyde as fi/ati"e$
2fter fi/ation the sa#%le should be analyDed i##ediately if %ossible, though it
re#ains stable for =E hours$ The alibration ontrol beads &o##erially
a"ailable* are run in the flo, yto#eter to he. the settings, %rior to
analyDing the sa#%le$

The ideal reo##ended %anel of antibodies for ly#%hoyte %henoty%ing by
t,oAolour analysis &using o#binations of = #onolonal antibodies to
different #ar.ers and ,ith different labels, %er tube* is as follo,s:
A CDEG &%an leu.oyte #ar.er*! CD0E &#onoyte #ar.er*
A CD@ &%an T ell* ! CDE
A CD@ ! CD1
A CD@ ! CD0? T CDG? &8: ell #ar.ers*
A CD0B &B ell #ar.er*
A Isoty%e ontrols &Ig30!Ig3=*
The gate for analysis #ay be set on the ly#%hoyte %o%ulation by -' "s$ ''
&ly#%hoytes se%arate out to the left of and belo, granuloytes as they are
s#aller in siDe and less granular, but #onoytes lie #uh loser to the#*$
CDEG!0E staining hel%s to further define the gate in ,hih the ells are to be
#easured and ensures ly#%hoyte %urity, e/luding #onoytes &at least 1GF
of ells in the -'!'' gate should be ly#%hoytes i$e$ CDEG
bright
,CD0E
A
*$ This
and the CD@!CDE o#bination is essential for CDE ounting$ The CD1 ells, B
ells and 8: ells #ay be ounted to onfir# if the totals add u%, for further
auray$ Isoty%e ontrols #ay not be neessary if the %ositi"e ell %o%ulation
is stained brightly and is distint fro# the negati"e %o%ulation$ (ther,ise
these ,ill be re+uired to set the le"el ,here autofluoresene and nonAs%eifi
fluoresene ends and s%eifi fluoresene begins$
$..>.$... M-,r)s6h"r" ss9 : 2 o##erial #iros%here assay for CDE ount is
a"ailable as the Manual CDE Count :it &Coulter, <'2*$ The ste%s are as
follo,s:
A 5hole ;DT2 blood &0>> L* is added to a tube$
A This is follo,ed by addition of 0> L s#all &>$? M* late/ #iros%heres
oated ,ith antiACD0E antibodies, to blo. #onoytes by binding to these$
118
A 'ubse+uently, 0> L large &= M* antiACDE oated beads are added$ The
#i/ing and binding is for = #inutes eah$
A 2fter attah#ent of the larger beads to CDE ells, 0> L of the #i/ture is
transferred to another tube ,ith 0>> L of a lysis and staining solution, and
#i/ed for 0>A0G seonds$

A This is used to harge the t,o ha#bers of a hae#oyto#eter, ,hih is
e/a#ined #iroso%ially$ (nly ells ,ith @ or #ore large beads attahed
&rosettes* are ounted, and not the ells ,ith s#all or both s#all and large
beads &,hih inlude #onoytes* $

A The total nu#ber of suh ells in both ha#bers is #ulti%lied by H$@ &a fator
aounting for all dilutions and on"ersions* to gi"e the absolute CDE ount
%er L$
$..>.$.=. E'E91" -11+')ss9 (EIA) : Ca%ellia CDE!CD1 test &'anofi )asteur,
-rane* is a oneAste% i##unoenDy#ati assay based on a%ture of T ells by
CD= antibodies bound to the ,ells of a #irotitre %late, instead of
solubiliDation, follo,ed by detetion ,ith antiACDE or CD1 %ero/idase
on7ugate in an ;I2 for#at$ The CDE and CD1 nu#ber is obtained by
on"ersion fro# %#ol!L "alues on a standard ur"e$
T62/ CDE test .it &T Cell Diagnostis, <'2* is a test based on solubiliDation of
CDE ells fro# ,hole blood by a lysis solution to release its antigen follo,ed
by its detetion in an ;LI'2 for#at$ The nu#ber of CDE #oleules %er ell is
about G>,>>> and it does not "ary #uh$ 2lso, the ontribution of CDE
#oleules fro# #onoytes is too s#all &the density of #oleules being a
tenth of that seen in CDE ly#%hoytes* and lies belo, the sensiti"ity li#it of
the test$ Therefore, #easure#ent of the nu#ber of solubiliDed CDE
#oleules by ;LI'2 an be used to alulate the nu#ber of CDE ells$
Gy885ne CD";CDE Cell )onitoring Test =it @ Intracel1 CSAA 5ses
non36lo< cyto8etry 8et:ods to 4ro9ide a7sol5te cell co5nts 6or 7ot:
CD" 4ositi9e and CDE 4ositi9e T ly84:ocytes in less t:an 3$ 8in5tes.
Its 4ro4rietary tec:nology 5ses a 8i!t5re o6 anti7ody3coated
8agnetic and 6l5orescent 7eads. T:e 8agnetic 7eads isolate t:e cells
o6 c:oice and t:e 6l5orescent 7eads 4ro9ide t:e signal to co5nt t:e
cells.
%r"s"'( s((+s )3 1"(h)*s 3)r CD> ,"00 ,)+'( : -lo, yto#etry is the
#ost aurate #ethod for deter#ining CDE ell ounts, though it is
tehnially de#anding and o#%arati"ely e/%ensi"e both in ter#s of
e+ui%#ent and reagents$ The use of a%%ro%riate ontrols in eah run and both
internal and e/ternal +uality ontrol &e$g$ through the e/ternal +uality
assurane %rogra##e of the International <nion of I##unologial 'oieties,
.no,n as Q2'I* are essential$
In single %latfor# tehnology &e$g$-2C'ount fro# Beton Di.inson, I##uno
VC' fro# Coulter*, total leu.oyte and differential ly#%hoyte ounts are
%ossible on the sa#e instru#ent as the one ,hih deter#ines the CDE
%erentage$ These #ethods are less %rone to errors in absolute CDE ounting$
'uh errors #ay our in other flo, yto#eters ,hih #easure the
%erentage of CDE ells, and re+uire se%arate auto#ated or #anual
#easure#ent of hae#atologial %ara#eters &i$e$ TLC and DLC* to generate
absolute ounts fro# CDE %erentage$ -2C'ount is, ho,e"er, a%able of only
li#ited analysis &e$g$ only CD@, E and 1*$ These syste#s are not e/tendable
to studies on other %henoty%i #ar.ers and are also losed in ter#s of
reagent usage$ Their #ain ad"antage is o#%arati"e ease of analysis and
119
auray ,ithout the need for highly s%eialiDed tehnial s.ills or a se%arate
hae#atologial ounter$
;I2Abased and #anual #ethods are easier to %erfor# and less e/%ensi"e
than flo, yto#etry, but are less aurate$ 2n e"aluation o#%aring the
Coulter #anual .it ,ith flo, yto#etry sho,ed a oeffiient of orrelation of
>$B0$ These .its ha"e a role in de"elo%ing ountries due to lo,er ost and
si#%liity$ 2nother ad"antage is that single sa#%les an be tested$
$..>... S)0+/0" MrB"rs : 2 large nu#ber of soluble #ar.ers of i##une ati"ation
ha"e been e"aluated as %rognosti indiators in HIV infetion$ These inlude
seru# ! %las#a le"els of neo%terin, beta =A#iroglobulin, tu#our nerosis
fator al%ha &T8-*, soluble CD1, soluble yto.ine ree%tors &sILA=6 for ILA=,
sT8- 6II for T8-* et$ 'oluble ree%tors are usually #ore stable than their
orres%onding yto.ines$
$..>...$. N")6("r-' : 8eo%terin &?ADAerythroAtrihydro/y%ro%yl%terin* is a %rodut
of guanosine tri%hos%hate atabolis# ,hih is deri"ed fro#
#aro%hages and B ells sti#ulated by interferon ga##a and reflets its
ati"ity, ,hile being stabler$ 8eo%terin is %resent in both urine and seru#$
<rine and seru# neo%terin le"els are ele"ated in %atients ,ith asy#%to#ati
HIV infetion as o#%ared to seronegati"e indi"iduals$ It is an early #ar.er of
HIV infetion$ The le"els rise further on %rogression fro# %reA2ID' to linial
2ID'$ 'ine neo%terin le"els are sti#ulated by HIV infetion, #easure#ent
of neo%terin le"el an be useful in #onitoring %rogression and e"aluating
anti"iral thera%y$
Ho,e"er, sine it is a nonAs%eifi #ar.er, besides HIV infetion, high
neo%terin le"els in urine and seru# #ay be found in nu#erous infetions
and infla##atory disorders, ollagen "asular diseases and in ad"aned
stages of ertain #alignanies$

M"(h)* '* 6r-',-60" : The earlier tests a"ailable for neo%terin assay in
seru# ,ere based on radioi##unoassay but no, on"enient .its based on
;LI'2 are a"ailable e$g$ ;LItest 8eo%terin &B62HM' Diagnostis, 3er#any*
and fro# IBL, Ha#burg, 3er#any$ The .its are based on o#%etiti"e ;LI'2 $
The %lates are oated ,ith %olylonal antiAneo%terin antibody raised in
ani#als e$g$ shee%! rabbit$ The neo%terin in the test sa#%le o#%etes ,ith
enDy#e&al.aline %hos%hatase or %ero/idase*Aon7ugated neo%terin$ The
enDy#e on7ugate is deteted by addition of the substrate and a lo,er (D
orres%onds to a higher le"el of neo%terin in the sa#%le$ 8eo%terin standards
,ith different le"els of neo%terin are run along ,ith sa#%les to generate a
standard ur"e for on"erting (D readings to le"els in n#ol!L$

$..>..... B"(;. 1-,r)20)/+0-' : BetaA= #iroglobulin &=A#*is a lo, #oleular
,eight &00,1>> .D* %oly%e%tide that for#s the light hain of lass 0
#a7or histoo#%atibility o#%le/ antigen %resent on the surfae of #ost
nuleated ells$ It is a nons%eifi but relati"ely sensiti"e #ar.er for
i##une ati"ation$ The results fro# se"eral longitudinal studies of highAris.
grou%s ha"e also de#onstrated that betaA= #iroglobulin is an early
#ar.er of HIV infetion$ 'eru# onentration abo"e ba.ground le"els are
deteted in #ost infeted indi"iduals ,ithin ? #onths of seroon"ersion$
High seru# betaA= #iroglobulin le"els, es%eially during the first year after
seroon"ersion to HIV %ositi"ity, ,ere assoiated ,ith %rogression to 2ID'$
Its #easure#ent #ay be useful in e"aluating antiA"iral thera%y$
High le"els of betaA= #iroglobulin ha"e also been re%orted in tuberulosis,
yto#egalo"irus infetion and ly#%ho#as, ,hih #ay be %resent in the late
120
stage of HIV infetion #a.ing it i#%ortant to inter%ret the data on seru#
betaA= #iroglobulin le"els in the linial onte/t$ ;le"ated le"els are also
seen in ertain HIV high ris. grou%s suh as hae#o%hilias and drug abusers$
M"(h)* '* 6r-',-60" : BetaA= #iroglobulin is %resent in #ost biologi
fluids at lo, onentrations and an be #easured in seru# or %las#a by
using o##erially a"ailable +uantitati"e o#%etiti"e ;LI'2 based tests$ The
o##erially a"ailable .its inlude Quanti.ine =A# &64D 'yste#s,<'2*and
=A#&(63enTe Diagnosti.a, 3er#any*$
%r"s"'( s((+s )3 s)0+/0" 1rB"rs )3 -11+'" ,(-!(-)' : Tests for
soluble #ar.ers of i##une ati"ation, inluding these t,o #ar.ers, as ,ell as
others li.e soluble ree%tors for T8- and ILA=, are a"ailable as o##erial
;LI'2s and are o#%ariti"ely easy to %erfor# and less e/%ensi"e$
6eent studies ha"e sho,n that sT8-A6II le"el ,as %rognostially si#ilar to
neo%terin and CDE le"els and better than sILA=6 le"el$ 2t CDE ounts O
=>>!L, CDE and indi"idual soluble #ar.ers ,ere found %rognostially
su%erior to e"en HIV 682 load, though "iral load ,as a better #ar.er at CDE
P =>>!L $ 2 o#bination of CDE ,ith sT8-A6II or neo%terin ,as also slightly
su%erior to "iral load aross the range of CDE "alues$ -urther#ore, these
soluble #ar.ers inrease signifiantly P ? #onths before deline in CDE le"el
and inrease in HIV 682 le"el$ This i#%lies that i##une ati"ation %reedes
the inrease in "iral load and is follo,ed by a ra%id and aelerated loss of
CDE ells$
$..?. C)s( 3,()r '* !-0/-0-(9 :
NOTE: M)s( )3 (h" ("s(s !-0/0" 3)r 6r)2')s(-, 3)00)8;+6 r"
6r)6r-"(r9 -("1s. Th" 0-s( )3 1'+3,(+r"rs '* +(h)r-E"* *"0"rs -'
I'*- -s ')( "Ah+s(-!" '* r"6r"s"'(s -'3)r1(-)' !-0/0" ( (h"
(-1" )3 8r-(-'2. I( *)"s ')( -1609 (h( (h" s6",-3-"* -("1s r" -' '9
89 r",)11"'*"* /9 (h" 6'"0 )3 "A6"r(s.
V-r0 1rB"rs : 6ohe 2#%lior HIV Monitor .it &only reagents* for =E tests
osts about 6s$H>,>>>$ It re+uires a ther#oyler and is read olori#etrially$
2ddress:
5heeon Instru#ents In$
BG, III 't$, :ar%aga# 2"enue,
6$2$)ura#, ChennaiA?>>>=1$
-a/ >EE EBE>H11

82'B2 &(rganon Te.ni.a* is yet to beo#e a"ailable$ (ther %roduts of this
o#%any are a"ailable through Infar &India* Ltd$
2ddress:
Infar &India* Ltd$
0=D, 9udges Court 6oad,
CaluttaAH>>>=H$
-a/ >@@ EH>?0HH
S+rr)2(" 1rB"rs : 2 flo, yto#eter an ost bet,een 6s$=G,>>,>>>A
6s$G>,>>,>>> de%ending on aessories needed$ ;ah antibody or
o#bination for =Aolour flo, yto#etry osts about 6s$0>,>>>A0=,>>> for G>
tests$ 2fter adding the osts of all reagents inluding lysis solution, #ini#u#
of = o#binations of antibodies, isoty%e ontrols and buffers et$, the reagent
ost ,or.s out to about 6s$H>>A1>> %er sa#%le$
121
The Manual CDE Count .it &Coulter* osts about 6s$G>> %er test$
The ;I2 based Ca%ellia CDE!CD1 ,hole blood .it &'anofi )asteur* osts
about 6s$=G,>>> for E@ tests$
6eagents for flo, yto#etry are a"ailable fro# Beton Di.inson &no, also
inludes )har#ingen* and Be.#an Coulter &inludes so#e I##unoteh
flo, %roduts also*$
2ddresses:
Coulter )roduts Di"ision
!o 2#ersha# )har#aia Bioteh 2sia )aifi Ltd$
E0, Co##unity Centre, :ailash Colony ;/tension,
Ia#rud%ur, 8e, DelhiA00>>E1$
-a/ >00 ?=0=00E
Coulter )roduts Di"ision
!o 2#ersha# )har#aia Bioteh 2sia )aifi Ltd$
01, =
nd
floor, Cunningha# 6oad,
BangaloreAG?>>G=$
-a/ >1> ==>E1>@
Beton Di.inson India )"t$Ltd$
Manor Hotel, HH, -riends Colony,
8e, DelhiA00>>?G$
-a/ >00 ?1@0H1@! ?1=E>G?
The ost of an ;LI'2 .it for B? tests for the soluble #ar.ers, inluding
yto.ines and their ree%tors, is about 6s$=>,>>>A==,>>>$ :its are a"ailable
fro# a nu#ber of #anufaturers, so#e of ,hih are listed belo,$
2ddresses:
-or 64D 'yste#s %roduts A
'%etral Diagnostis )"t$Ltd$
CA@=, :irti 8agar,
8e, DelhiA00>>0G$
Tel >00 GEG1>EB

-or 3enDy#e %roduts A
Bioteh India
BA=B!0>, 8andigra#, Lan.a,
VaranasiA==0>>G$
-a/ GE= @0@EHE
-or I##unoteh %roduts A
('B 2genies )"t$Ltd$
=!@HA2, 2nsari 6oad,
Daryagan7,
8e, DelhiA00>>>=$
-a/ ==0?H@?
$..F S+22"s("* r"*-'2:
i* CDC$ 0BBH 6e"ised guidelines for %erfor#ing CDET TAell deter#inations in
%ersons ,ith hu#an i##unodefiieny "irus &HIV*$ MM56 0BBHJ E? &8o$ 66A
122
=*:0A=B$
ii* CDC$ 0BB@ 6e"ised lassifiation syste# for HIV infetion and e/%anded
sur"eillane ase definition for 2ID' a#ong adolesents and adults$ MM56
0BB=J E0 &8o$66A0H*:0A@G$
iii* -ahey 9L, Taylor 9M3, Detels 6, et al$ The %rognosti "alue of ellular and
serologial #ar.ers in infetion ,ith hu#an i##unodefiieny "irus ty%e 0$ 8
;ngl 9 Med 0BB>J @==:0?=AH=$
i"* 3raha# 8MH$ The role of i##unologial and "iral #ar.ers in %rediting
linial outo#e in HIV infetion$ 2ID' 0BB?J 0> &su%%l G*:'=0A'=G$
"* 'alaDarA3onDaleD 9-, MartineDAMaDa (, 8ishanian ), et al$ Inreased i##une
ati"ation %reedes the infletion %oint of CDE T ells and the inreased seru#
"irus load in hu#an i##unodefiieny "irus infetion$ 9 Infet Dis 0BB1J
0H1:E=@A@>$
9iA Nic:olson IHA1 &elleca .)1 I57ert S1 et al. '9al5ation o6 alternati9e
CD" tec:nologies 6or t:e en58eration o6 CD" ly84:ocytes. I
I885nol )et:ods 1DD"F 177:"33$".
123
$=. TRANSFUSION TRANSMITTED INFECTIONS OTHER THAN
HIV INFECTION
$=.$ I'(r)*+,(-)' : Blood transfusion is usually life sa"ing but an trans#it
diseases if blood is not tested %ro%erly for the %resene of #iroAorganis#s
before transfusion$ The #ain diseases trans#itted through blood are
he%atitis, hu#an i##unodefiieny "iruses &HIV*, sy%hilis and #alaria Table
0@$0 gi"es the list of #ain diseases trans#itted through blood$
T/0" $=.$ L-s( )3 *-s"s"s (r's1-(("* (hr)+2h /0))*
VIRAL BACTERIAL %ARASITIC
A He%atitis B "irus &HBV* A 'y%hilis A Malaria
A He%atitis C "irus &HCV* A Bruella A To/o%las#a
A He%atitis 3 "irus &H3V* A Mirofilaria
A Hu#an i##unodefiieny "irus
&HIV*
A ;%stein Barr "irus &;BV*
A Cyto#egalo"irus &CMV*
A Hu#an T ly#%hotro%i "iruses
& HTLVA0 and HTLVAII*

$=.. Tr's3+s-)' ss),-("* h"6(-(-s : Transfusion assoiated
he%atitis &T2H* de"elo%ing as a onse+uene of transfusion of infeted
blood or blood o#%onents affets e"ery s%here of #ediine$ The illness
initially often re#ains sublinial but ould later result in hroni li"er
disease and e"en %ri#ary he%atoellular arino#a$ It is i#%ortant that
dotors ,or.ing in transfusion #ediine, general #ediine, #irobiology,
laboratory #ediine, %athology, soial and %re"enti"e #ediine should be
a,are of the s%etru# and onse+uenes of T2H$
Viral he%atitis being one of the serious o#%liations of blood transfusion,
an be due to different infetious agents li.e he%atitis 2,B,C,D,;, and 3
"isuses &H2V, HBV, HCV, HDV, H;V, H3V* besides yto#egalo"irus &CMV*
and ;%stein Barr "irus &;BV*$ )ost transfusion he%atitis is #ainly due to
he%atitis B "irus or he%atitis C "irus$ There are about three hundred
#illion he%atitis B "irus arriers and about one hundred #illion he%atitis C
"irus arriers in the ,orld$ <nfortunately #ost of the arriers of he%atitis
"iruses are not e"en a,are of their arrier status$
$=...$H"6(-(-s A !-r+s : He%atitis 2 "irus &H2V* ,as identified by -einstone
in 0BH@ and is %ri#arily res%onsible for infeti"e he%atitis$ Transfusion of
He%atitis 2 "irus by blood transfusion is rare beause of t,o #ain reasons
:
i )resene of "irus in blood for a short ti#e$
ii 6are arrier state$
He%atitis 2 "irus irulates in blood only during the initial aute %hase of
infetion and during this %eriod the indi"idual is not the andidate for
donation of blood$ Ho,e"er, in rare ases "irae#ia #aybe %resent before the
de"elo%#ent of sy#%to#s and blood donated during this %eriod ould be an
124
effeti"e "ehile for trans#ission of H2V$
$=....H"6(-(-s B !-r+s (HBV) : He%atitis B "irus is the #ost i#%ortant ausati"e
agent of T2H$ Hu#ans are the only reser"oir of He%atitis B "irus &HBV*$
He%atitis B "irus ,as diso"ered by Blu#berg et al in 0B?G in the seru# of an
2ustralian aborigine and thus its antigen is also alled as 2ustralia antigen$
The "irus %artile &"irion* is a s#all o#%le/ double shelled struture ha"ing
an e/ternal dia#eter of E= n# ,ith a nuleoa%sid ore and li%o%rotein oat$
This %artile ,as first diso"ered by Dane and his olleagues in 0BH> and thus
is alled Dane %artile$ It re%resents the o#%lete HBV$ The nuleoa%sid
ore is =H n# in dia#eter$ It re%liates in the nulei of infeted he%atoyts
and %ossesses a distint antigen alled he%atitis ore antigen &HB2g*$ The
"irion ore antigen ontains D82 %oly#erase and double stranded D82
#oleule -ig$ 0@$0$
The #a7or strutural %rotein is li%o%rotein "iral oat and is alled he%atitis B
surfae antigen &HBs2g*$ This %rotein is synthesiDed in the yto%las# of
infeted he%atoytes in e/ess +uantities than that needed for "iral asse#bly$
The e/ess #aterial in the for# of s#all %artiles, either ==n# s%herial
%artiles or s#aller tubular %artiles of "arying lengths are released in the
blood strea#$ 'ine these s#aller %artiles ontain only the #a7or "iral
%rotein HBs2g they are onsidered nonAinfetious but at as #ar.ers of
infetion$ (asionally o#%lete "irus ours in the blood strea#$ The "irion
has an other #ar.er i$e$ he%atitis B e antigen &HBe2g* ,hih is losely
related to ore antigen and is onsidered as a #ar.er of infeti"ity$
S+/(96"s )3 HBV : HBV has an antigeni heterogeniity$ It onsists of at
least three different antigeni o#%onents: a grou% s%eifi antigen and t,o
%airs of ty%e s%eifi antigens d3y and <3r. (nly one #e#ber of eah %air
being %resent at a ti#e$ HBV an thus be di"ided into four #a7or antigeni
subty%es ad<1 adr1 ay< and ayr$
Tr's1-ss-)' )3 HBV -'3",(-)' : The "irus is highly infetious and #inute
a#ounts of blood!body fluids as little as >$>0 l an trans#it the infetion$
Trans#ission of HBV is #ainly by the %arenteral route ,hih in"ol"es diret
ontat ,ith body fluids$ The #ost o##on routes of infetion are :
A Diret or indiret ontat ,ith infeted blood &needle sti.s, uts et$*$
A IVD< &sharing needles*$
A <n%roteted se/ual ontat$
A Transfusion of infeted blood or blood %roduts$
A Vertial trans#ission &%erinatal trans#ission*$
C0-'-,0 6r"s"'((-)' )3 HBV -'3",(-)' : The inubation %eriod of HBV
infetion is about G>A0G> days during ,hih the %atient has no signs and
sy#%to#s but the "irus #ay be deteted in the blood strea#$ There is a
%rodro#e of #alaise, ,ea.ness, loss of a%%etite, nausea follo,ed by
sy#%to#s suh as fe"er, rash, arthralgia and 7aundie de%ending u%on the
se"erity of infetion$ In #ild ases 7aundie does not our ho,e"er, #ore
se"ere ase an result in serious disease$ 2ntibodies to three #ain antigens
i$e$ antiA +2sA71 antiA +2cA7 and antiA +2eA7 a%%ear at different ti#es
#ainly during the ourse of disease and #ay %ersist for years, so#e ti#es for
life after reo"ery$
2ntiA+2sA7 is usually assoiated ,ith resistane to reinfetion
Anti3+2cA7 is not %roteti"e but is onsidered as a sensiti"e indiator of HBV
re%liation$ Its %resene in asy#%to#ati blood donor indiates either reent
HBV infetion or hroni arrier state$
125
Anti3+2eA7 is seen #ostly in healthy arriers of HBs2g$
In 0> to =>F of linially diagnosed HBV infetions, the infetion does not
resol"e but enters into hroni %hase, ,hih #ay last for 7ust si/ #onths and
then resol"e or %ersist for life$ In so#e hronially infeted %ersons, no ill
effets are seen but #ay de"elo% hroni li"er disease ,hih #ay lead to
irrhosis, he%atoellular arino#a and death$
S"r)0)2-,0 '* /-),h"1-,0 1rB"rs )3 HBV -'3",(-)' : The
de"elo%#ent of "arious serologial #ar.ers during the aute and hroni HBV
infetion are sho,n in fig 0@$= and 0@$@, res%eti"ely and the inter%retation of
"arious #ar.ers is gi"en in table 0@$=$
A66"r'," )3 HBsA2 -' 6(-"'(Ds s"r+1 : It is usually the first detetable
serologial #ar.er in aute HBV infetion and re#ains detetable fro# fe,
days to se"eral #onths de%ending u%on reo"ery of the %atient$ In so#e
ases HBs2g %ersists for years$ These are alled arriers$
A66"r'," )3 A'(-;HB, (HB,A/) : In #ost ases HB2b a%%ears in the
seru# during the aute illness &%reiteri %hase* and %ersists for #onths$

A66"r'," )3 HBV N"O '(-2"' '* -(s '(-/)*9 : He%atitis B e antigen
&HBe2g* a%%ears in the seru# al#ost at the sa#e ti#e as HBs2g but
disa%%ears in #ost ases ,ithin a fe, ,ee.s$ Its disa%%earane is follo,ed
by the a%%earane of anti HBe &HBe2b* in seru# ,hih lasts for se"eral
#onths$
A66"r'," )3 '(- HBs (HBs 2b* : 2ntibody to HBs 2g &HBs2b* a%%ears
after the disa%%earane of HBs2g and indiates reo"ery fro# aute illness$ 2t
ti#es the a%%earane of HBs2b #ay be delayed for ,ee.s after HBs2g
beo#es undetetable and during this %eriod alled ,)r" '(-/)*9 8-'*)8,
anti HBAIgM #ay be the only detetable #ar.er of reent HBV infetion$
126
T/0" $=.. I'("r6r"((-)' )3 !r-)+s HBV 1rB"rs
HBsA2 HB"A2 '(-;
HB,
'(-;HB" '(-;
HBs
I'("r6r"((-)'
A A A A A 8o e"idene of %resent
or %ast HBV infetion$
T A A A A Inubation %eriod of HBV$
T T T A A ;arly HBV infetion ,ith
high infeti"ity$
T A T T A Late aute HBV infetion
or hroni HBV infetion
,ith lo,er infeti"ity$
A A T T A 5indo, %eriod in late
aute HBV infetion$
A A T T T Con"alesent fro# HBV
infetion$
A A T A T Late %hase of
on"alesene &antiA HBe
has ,aned*$
A A A A T 6es%onse to HBV "aine
or reent ad#inistration
of hy%eri##e antiA HBs
i##unoglobulin$
%r"!0"'," )3 H"6(-(-s B -'3",(-)' : The %re"alene of HBs2g arrier rate
"aries fro# >$>0F to @>F fro# one ountry to another$ In general, ountries
,ith lo, soioeono#i struture ha"e higher HBs2g arrier rates and #ost of
the industrialised and de"elo%ed ountries ha"e HBs2g arrier rate less than
0F$ De%ending u%on the %re"alene of he%atitis B in the general %o%ulation,
the International Tas. -ore on he%atitis B has lassified geogra%hial areas
into lo, &O=F*, inter#ediate &= to H F* and high %re"alene &PHF* regions$
India falls in the inter#ediate %re"alene region$
S,r""'-'2 ("s(s 3)r h"6(-(-s;B !-r+s : In order to redue the ris. of %ost
transfusion he%atitisAB infetion, HBs2g sreening ,as first of all atte#%ted
by (.ohi et al at the <ni"ersity of To.yo hos%ital in Dee#ber, 0B?1$
Donated blood ,as sreened by i##unodiffusion tehni+ues for the %resene
of he%atitis B surfae antigen &HBs2g*$ 'ine then a nu#ber of test
%roedures ha"e been de"elo%ed to sreen the blood for the detetion of
HBs2g$
Currently the #ost sensiti"e and ,idely used tests for detetion of HBs2g are
based on ;I2!;LI'2 %rini%le$ In India, unfortunately, #any blood ban.s
es%eially those ,ith little or no blood in the +uarantine stage and ha"ing
,al. in donor %rogra##es are o#%elled by the iru#stanes to use ra%id
sreening tests for detetion of HBs2g$ 2s these test %roedures are based
on late/ agglutination
127
T/0" $=.= M"(h)*s 3)r (h" *"(",(-)' )3 HBsA2 -' )r*"r )3 s"'s-(-!-(9.
Test
'ensiti"ity
&#ini#u# le"el of HBs2g
deteted*
I##unodiffusion &I$D$* Lo,
I##unoeletroAos#o%heresis &I;()* &P 0>> ng!#l*

Late/ agglutination &L$2$*

6e"erse %assi"e hae#agglutination
&6)H2*
Moderate
&0>A0>> ng!#l*
I##uno hro#atogra%hy test &ICT*
;nDy#e I##unoassay &;I2!;LI'2* High
6adio I##unoassay &6I2* >$GA0> ng!#l$
or hro#atogra%hi %rini%les, ,hose sensiti"ity is lo,, a large nu#ber of
HBV blood units are leared for transfusion and thus the infetion gets
trans#itted to the unlu.y %atients$ Centers ha"ing a large blood olletion
use ;LI'2 #ethod for sreening of blood for %resene of HBs2g$
E'E91" I11+')ss9 (EIA&ELISA)
%r-',-60" : ;nDy#e i##unoassay is based on sand,ih %rini%le in"ol"ing
the use of solid %hase e$g$ the ,ell of #iro%late oated ,ith antibodies of
HBs2g &-ig 0@$E*$ Test sa#%le &seru#!%las#a* is added to the #iro,ell and
inubated$ 2t the end of inubation %eriod, the seru# is ,ashed off and
on7ugate i$e$ #onolonal Ig3 on7ugated to traer enDy#e is added and
inubated$ HBs2g if %resent in the sa#%le, allo,s the enDy#e on7ugate to
bind to solid %hase and build sand,ih '(-/)*9;'(-2"';'(-/)*9
o#%le/$ ;/ess on7ugate is ,ashed a,ay and hro#ogen substrate is
added for enDy#e ation to %rodue a olour$ The intensity of the olour is
diretly %ro%ortional to the enDy#e ati"ity ,hih in turn is %ro%ortional to the
HBs2g onentration in the sa#%le or ontrol$ The resulting olour an be
deteted either "isually or by ;LI'2 reader$
There are a nu#ber of o##erial .its &#iro ;LI'2 test* a"ailable for
detetion of HBs2g and it is reo##ended that the %roedure %ro"ided by the
#anufaturers in their .it is follo,ed$
R"!"rs" 6ss-!" h"1220+(-'(-)' ss9 (R%HA) : 6e"erse %assi"e
hae#agglutination assay also gi"es satisfatory result but its sensiti"ity is
lo,er than enDy#e i##unoassay so it is not in use %resently$
I11+') ,hr)1()2r6h9 ("s( (ICT) : It is a ra%id stri% test for the
detetion of HBs2g and is #ore sensiti"e than late/ agglutination test$
%r-',-60" : In hro#atogra%hi i##unoassay the #e#brane is %reoated
,ith anti HBs2g a%ture antibody on the test band region and goat antiA
#ouse antibody on the ontrol band region$ During testing the s%ei#en
&seru#!%las#a* is allo,ed to reat ,ith the olloidal gold %artiles ,hih ha"e
been oated ,ith anti HBs2g #onolonal antibody$ The #i/ture then #o"es
u%,ard hro#atogra%hially by a%illary ation$ In ase of %ositi"e result a
oloured band ,ill be for#ed in the test band region of the #e#brane ,ith
s%eifi anti HBs2g antibody olloidal gold %artile o#%le/$ 2bsene of
oloured band in the test band region indiates a negati"e result$ In order to
ha"e %roedural ontrol, a oloured band ,ill al,ays a%%ear at ontrol region
128
irres%eti"e of the %resene or absene of HBs2g$
L-1-((-)'s )3 HBsA2 s,r""'-'2 : ;"en after using the #ost sensiti"e
sreening test for the detetion of HBs2g on blood fro# "oluntary donors,
HBs2g negati"e units ha"e been found to trans#it the he%atitis B "irus
infetion$ The basi ai# of blood transfusion ser"ies should be to %ro"ide
Dero ris. blood for transfusion, ho,e"er in atual %ratie this is not
ahie"able beause of the follo,ing reasons:
A La. of test ,hih is 0>>F sensiti"e %robably due to HBs2g #utants$
A -ailure to detet HBs2g in so#e donors in the early inubation %eriod$
A Diffiulty in arrying out highly sensiti"e tests li.e D82 hybridiDation and )C6$
;/tensi"e studies ha"e sho,n that the e/lusion of blood units ,ith isolated
high titre of anti HB &e"en ,hen HBs2g is negati"e* are effeti"e in #ar.edly
reduing the hanes of %ost transfusion he%atitis B infetion$ This fat led
so#e of the blood ban.s to inor%orate the testing for anti HB in the donor
sreening %rotools$ This strategy is not ado%ted in our ountry and #ost of
our blood ban.s restrit sreening for he%atitis to HBs2g alone$
$=...=H"6(-(-s C !-r+s (HCV) : HCV is the #ost o##on ause of %ost
transfusion nonA2 nonAB he%atitis in the de"elo%ed ,orld$ The %re"alene of
HCV antibodies in blood donors in de"elo%ed ountries ranges fro# >$EA=F$
Ho,e"er in ;gy%t it is as high as 0EF$ The %re"alene of HCV in Indian blood
donors has not been studied e/tensi"ely but as %er so#e a"ailable re%orts the
%re"alene of anti HCV "aries fro# >$?F to G$=F in "oluntary!re%lae#ent
donors$ G>F of HCV infetion is re%orted to %rogress to hroni infetion of
,hih =>F #ay %rogress to irrhosis and he%atoellular arino#a$
HCV ,as identified by loning tehni+ues$ The "irus is an en"elo%ed single
stranded 682 "irus, G>A?> n# in dia#eter ,ith a geno#e of 0>,>>>
nuleotides and shares relationshi% ,ith -la"i"iruses$ The geno#e enodes
for three strutural %roteinsJ a%sid %rotein &CA=>:Da* and t,o en"elo%e
glyo%roeins &;lAg%@G and ;=Ag%H>* and se"eral nonstrutural %roteins &8'*
i$e$ 8'=, 8'@, 8'E2, 8'EB, 8'G2, 8'GB ha"e been identified &fig$ 0@$G*$
The detetion of antibodies to HCV is used for serodiagnosis of HCV infetion$
It ,as in 0BB> that the 6irst generation test for anti HCV ,as introdued
,hih deteted antibodies to C 0>>A@ and a %art of 8'E region$

129
'trutural )roteins 8onAstrutural )roteins
8'E 8'G
GC
AAAAA
8C6
C ;0 ;= 8'= 8'@ 2 B 2 B AAA&<*nA
@C
8C6
)=0 g%A
@G
g%A
H>
)rotea
se!
helia
se
8T)as
e
8'@
oA
fato
r
682
%oly#e
rase
;n"elo%e
F-2. $=.? G"')1" )3 HCV
The second generation assays ,ere introdued in 0BB= ,hih inluded
antigens fro# ore and 8'@ region &C== and C=>> fro# 8'@A8'E!@@ fron
8'@ alone* in addition to C0>>A@$ T:ird generation tests ha"e been
de"elo%ed ,hih in addition ontain antigen fro# 8'G region$ The %resently
a"ailable reo#binant i##unoblot &6IB2* for the detetion of antibodies to
HCV inludes bands GA0A0, C0>>A@ C==A@, @@ and 8'G %oly%e%tide$
6eently .its ha"e been de"elo%ed ,hih inor%orate syntheti %e%tides
instead of reo#binant %e%tides #ar.ing the test #ore s%eifi$ If antibodies
to = or #ore antigens are %resent, the sa#%le is onsidered as %ositi"e$ )rior
to the de"elo%#ent of s%eifi #ar.ers ertain S5rrogate 8ar=ers i$e$
deter#ination of seru# alanine a#inotransferase &2LT* ,as used to identify
the high ris. donors for trans#ission of HCV$ 2lthough the surrogate #ar.ers
%layed an i#%ortant role in the %re"ention of he%atitis C %rior to the
introdution of HCV s%eifi assays but %resently the "alue of these tests
re#ains inonlusi"e and ontro"ersial$
$=...>H"6(-(-s D : It is also .no,n as Delta he%atitis$ It is aused by a
defeti"e 682 "irus that is unable to %rodue its o,n %rotein oat and thus
oats itself ,ith HBV surfae antigen$ The delta "irus is de%endent on %reA
e/isting or ono#itant HBV infetion for %ro%agation$ CoAinfetion auses
aute "iral he%atitis ,hih often beo#es ful#inant and is usually fatal, ,hile
su%erinfetion auses aute he%atitis %rogressing to hroni ati"e he%atitis
and irrhosis in HGF of arriers$
Delta he%atitis has %arenteral #ode of trans#ission and usually follo,s HBV
infetion$ 2lthough tests for delta antigen and antibody e/ist but should not
be used in blood donor sreening as all infetions ,ith delta "irus are %ositi"e
for HBs2g$ 'o routine testing for HBs2g ,ill eli#inate the ris. of trans#ission
of Delta "irus$
$=...?H"6(-(-s G !-r+s (HGV) : He%atitis 3 "irus is an en"elo%ed 682 "irus and
belongs to the -la"i"iridae fa#ily$ The "irus ,as diso"ered in 0BBG$ The
"irus geno#e is si#ilar to that of HCV ,ith ,hih it shares a =GF ho#ology
at the nuleotide le"el$ The H3V a%%ears less "ariable although "ariants ha"e
been desribed$ H3V re%liates in %eri%heral blood ells but its re%liation in
the li"er is still +uestionable$ The #ode of s%read of H3V is al#ost sa#e as
HCV$ The H3V an be deteted in the seru# by nulei aid tehnology
&)C6*$ 6eently an i##unoAassay has been de"elo%ed to detet antibody to
H3V$
The %re"alene of H3V in blood donor %o%ulation by doing antibody tests in
the de"elo%ed ountries ranges bet,een 0 to GF but the detetion of
antibody generally indiates reo"ery fro# and %robably the i##unity to the
H3V agent$
130
%r"!"'(-)' )3 6)s( (r's3+s-)' h"6(-(-s : -ollo,ing %oints should be .e%t
in #ind for %re"ention of %ost transfusion he%atitis:A
i )ro#otion of "oluntary blood donation$
ii )ro%er donor inter"ie, and seletion$
iii <se of #ost sensiti"e and s%eifi test for HBs 2g detetion &@rd
generation tests*$
i" 'reening for antibody to he%atitis ore antigen &2ntiAHB* if %ossible$
" 'reening for antiA H3V to detet arrier of H3V is %referable$
"i 3ood #anufaturing %ratie &3M)* for blood frationation and
inor%oration of "iruidal tehnology &'ol"ent detergent tehni+ue* for
"iral inati"ation$
$=.= M0r- : Trans#ission of #alaria an our after transfusion of ,hole blood,
red ells or any blood o#%onent onta#inated ,ith red ells$ )las#a
frations do not trans#it the disease$ The %arasites of all s%eies re#ain
"iable for at least one ,ee. in stored blood at =A?
>
C$ Ho,e"er, )las#odiu#
fali%aru# an re#ain "iable for =A@ ,ee.s$ The inubation %eriod of
)las#odiu# "i"a/ and )las#odiu# fali%aru# is one ,ee. to one #onth
,hile that of )las#odiu# #alariae is #any #onths$ )ost transfusion #alaria
&)TM* is %artiularly dangerous in %regnant ,o#en and i##unodefiient
%atients$
$=.=.$D"(",(-)' )3 10r- -' ,rr-"rs
EA1-'(-)' )3 /0))* 3-01 : The e/a#ination of blood fil# is not suitable
for sreening large nu#ber of blood donations beause it is diffiult to find
%arasites in blood fil# in the short a"ailable ti#e es%eially if density of
%arasites is less than 0>> %er #irolitre of blood$ Ho,e"er, the e/a#ination
of blood fil#s for %arasites ,ithin red ells is onsidered best #ethod for
diagnosis in sus%eted ases$
T"s( 3)r 10r- '(-/)*9 : Tests for #alaria antibody either by indiret
fluoresent antibody &I-2* test or ;LI'2 are alright for nonende#i areas but
not ideal for #alaria ende#i areas eg$ India as there is a li.ely hane that
#any blood donors ,ill ha"e antibodies and ,ill be re7eted$
$=.> C9()1"20)!-r+s (CMV) -'3",(-)' : Cyto#egalo"irus #ay be trans#itted
by blood transfusion and an ause se"ere disease in seronegati"e %re#ature
babies of lo, birth ,eight and seronegati"e i##unosu%%ressed %atients e$g$
%atients of bone #arro, trans%lantation or %atients ,ith HIV infetion$
'ine #ore than G>A1>F of adults in de"elo%ed ountries and al#ost 0>>F
in so#e de"elo%ing ountries ha"e antibodies to CMV, so sreening of all
donors is not %ratially feasible$ In high ris. rei%ients it is ad"isable to use
leuoyte de%leted blood$
$=.? S96h-0-s : 'y%hilis is a disease aused by Tre%one#a %allidu#$ It is %ri#arily
a se/ually trans#itted disease but the organis# an gain entrane in to the
body through #inute lesions on s.in or #uous #e#branes$
Blood transfusion is a %otential route of infetion, %artiularly if fresh infeted
blood is transfused$ The inubation %eriod in transfusion 'y%hilis is on the
a"erage BA0> ,ee.s and rei%ient usually %resents ,ith seondary eru%tions$
The hanes of %ost transfusion 'y%hilis ha"e been greatly redued beause
of the follo,ing reasons :
A ;/lusion of high ris. donors$
131
A 'reening for 'y%hilis$
A 'torage of blood at =A?
>
C for H= hours %rior to transfusion$
&'%irohaetes are "ery sensiti"e to te#%erature and die at lo,er
te#%erature*$
$=.?.$S"r)0)2-,0 ("s( : There an be non s%eifi tests and s%eifi tests
N)' S6",-3-, ("s( : The non s%eifi tests inlude :
i VD6L &Veneral Diseases 6esearh Laboratory test*
ii 6)6 &6a%id )las#a 6eagin* test$
VDRL ("s( : VD6L test is a si#%le, ra%id and eono#ial test for sreening
sy%hiliti infetion, ho,e"er the test needs inati"ation of the s%ei#ens
&sa#%le* and the shelf life of the reonstituted antigen is %oor$
R%R ("s( : It is a #aroso%i nonAtre%one#al floulation test for
sreening sy%hiliti infetion$ The test is ra%id and does not need inati"ation
of sa#%le, the antigen used is a #odified VD6L antigen &ardioli%in* ,hih
ontains #iro%artiulate haroal!arbon to enhane the "isual
inter%retation$ The test is su%%lied in the for# of .its by #anufaturers and it
is ad"ised to follo, the instrutions %ro"ided ,ith the .it$
S6",-3-, ("s(s : '%eifi tests are #ainly used for onfir#ation and are not
suitable for routine sreening$
i Tre%one#a %allidu# hae#agglutination test &T)H2*$
ii -luoresent tre%one#al antibody test &-T2*
iii ;nDy#e i##uno assay &;I2*$
$=.F Cr"+(E3"0*( ; KB)/ D-s"s" (CKD) : CreutDfeldt A 9a.ob disease is one of
the series of trans#issible s%ongifor# ene%halo%athies &T';* and is thought
to be aused by a %roteinaeous %artile s#aller than a "irus$ This nuleotide
infetious %artile is alled %r-)' ,hih ,as one thought to be a slo, "irus$
2lthough trans#ission of C9D through blood and blood %roduts has been
s%eulated but no ase has been re%orted$
$=.G H+1' T;,"00 0916h)(r)6-, !-r+s"s : Hu#an TAell ly#%hotro%i "irus
ty%eAI &HTLVA0* ,as the first hu#an retro"irus isolated fro# a %atient ,ith
adult TAell leu.ae#ia!ly#%ho#a &2LT* by 6obert 3allo and his grou% in
0B1>$ The disease an %ersist for a life ti#e but usually does not ause #a7or
illness in #ost %eo%le ,ho are infeted$ In rare instanes HTLVA0 is
assoiated ,ith a hroni degenerati"e de#yelinating neurologial disease
alled tro%ial s%asti %ara%aresis &T')* but no, often alled HTLV assoiated
#yelo%athy &H2M*$ HTLVA0 is #ore o##on in south,estern 9a%an and
Caribbean islands$ The route of trans#ission for HTLVA0 is by se/ual ontat
&%redo#inantly #ale to fe#ale*, by %arenteral e/%osure to blood and #other
to hild through breast #il.$
The seond hu#an retro"irus HTLVAII ,as diso"ered se"eral years after
HTLVA0$ It is losely related to HTLVA0 and it is diffiult to differentiate
bet,een these t,o "iruses in the routine laboratory$ 2lthough no fir#
assoiation bet,een HTLVAII and any disease has been found but the "iurs
,as first found in a %atient ,ith hairy TAell leu.ae#ia and subse+uently in
other TAell #alignanies, "iral 682 e/%ression has not been found in
#alignant ells$ High %re"alene of HTLVAII has been seen in I$V$ drug
abusers and their se/ual ontats in <'2 and other ountries$ The %re"alene
of HTLVAI and HTLVAII in intra"enous drug abusers in 8e, Lor. is BF and
01F, res%eti"ely$ The %re"alene of HTLVAI and HTLVAII in India is not
132
a"ailable$
The "iruses are trans#itted by transfusion of ellular o#%onents of blood but
not by ell free o#%onents li.e %las#a$ <ni"ersal sreening of blood donors
for HTLVAI ,as introdued in Dee#ber, 0B11 in <'2 and in 9uly, 0BB0 in
-rane$ In south ,estern 9a%an ,here HTLV is, ende#i sreening of blood
#ar.edly redued the rate of seroon"ersion in rei%ients of blood$
'e"eral HTLV sreening assays are a"ailable li.e gelatin %artile agglutination
&)$2$*, enDy#e i##unoassay &;I2* and indiret i##unofloresene &I-*$ The
sensiti"ity and s%eifiity of ;LI'2 tests ranges fro# B1$EF to 0>>F$ 8one
of the sreening tests a"ailable is able to differentiate bet,een anti HTLVAI
and anti HTLVAII beause of ross reati"ity due to HGF ho#ology of a#ino
aid se+uenes bet,een the t,o "iruses$ Confir#atory assays li.e ,estern
blot, 6I)2, antibody a%ture radioi##uno assay and )C6 are also a"ailable$
133
$>. INFECTIONS TRANSMITTED BY TRANSFUSION OF BLOOD COM%ONENTS
AND BLOOD %RODUCTS
$>.$ I'(r)*+,(-)' : The transfusion of blood began ,ith the use of ,holeAblood
and ontinued for #any years$ Ho,e"er, no,adays a "ariety of blood
o#%onents and %roduts are a"ailable for their use in %atients of anae#ia,
thro#boyto%aenia, neutro%aenia, abnor#al oagulation and
i##unodefiieny, et$ 5hole blood is transfused #ainly in ases of ra%id
blood loss due to #assi"e unontrolled hae#orrhage et$ The ter# bloodA
o#%onent is used for those blood #aterials that are se%arated by #eans of
differential entrifugation follo,ed by e/%ression into satellite!seondary
%a.s attahed to the original donor blood %a. ,hereas blood %roduts are
those blood #aterials that ha"e been e/trated and %urified fro# rude
%las#a and %reser"ed in stable for#$ 'o#e of the i#%ortant blood %roduts
and blood o#%onents ha"e been su##ariDed in Table 0E$0$ These %roduts
are su%%osed to be life sa"ing drugs$ Ho,e"er, unfortunately these blood
%roduts!o#%onents are assoiated ,ith ris. of trans#ission of "iral,
%arasiti and baterial diseases$ ;tiologial agents of these "arious diseases,
listed in Table0E$=, ha"e got e/ellent o%%ortunity to enter in a ne, hu#an
host$ (ut of the total ad"erse effets of transfusion of
blood!%roduts!o#%onents a%%ro/i#ately E>F are due to trans#ission of
these etiologial agents$ The degree of %roble# aused by these #irobial
agents is diretly related to their ende#iity in a %artiular area$
T/0";$>.$ L-s( )3 !r-)+s (96"s )3 /0))* ,)16)'"'(s & 6r)*+,(s
C"00+0r ,)16)'"'(s B0))* ,)16)'"'(s B0))* 6r)*+,(s
A
A
A
A
A
A
6ed blood ells
Leuoyte redued red ells
5ashed red blood ells
-roDen! deglyerolysed
red ells
)latelet onentrate
3ranuloyte onentrate
A
A
A
'ingle donor %las#a
-resh froDen
%las#a
Cryo%rei%itated
antihae#o%hili fator
A
A
A
A
A
A
A
-ator VIII
onentrate
-ator IV
onentrate
2lbu#in
)las#a %rotein
fration
I##une seru#
globulin
6h D
i##unoglobulin
2ntithro#bin III
onentrate
Hy%eri##une
seru#
$>.. M-,r)/-0 2"'(s (rs1-(("* (hr)+2h (r's3+s-)' )3 /0))* 6r)*+,(s
'* ,)16)'"'(s :
$>...$V-r+s"s : Ma/i#u# %erentage of transfusion trans#itted diseases are "iral
follo,ed by %arasiti and baterial infetions$ Trans#ission of "iruses is "ery
high if the blood!o#%onents!%roduts are fro# large %ool of %las#a ta.en
fro# #any donors$ The transfusion trans#issible "iruses are listed in tables
0E$= and 0E$@$
$>...$.$ R"(r)!-r+s"s : 6etro"iruses ,ere .no,n to ause #alignany and
leu.ae#ia in ani#als and ,ere ne"er sus%eted to be a ause of hu#an
disease$ The ad"ane#ent in #oleular biology and reo#binant D82
tehnology led to the diso"ery of the first hu#an T ly#%hoyte "irusATy%e
134
HTLV I$ 'ubse+uently HTLV II ,as also diso"ered$ The .no,ledge about
these "iruses enouraged the ,or.ers to %ro"e the etiology of 2ID' "irus$
The 2ID' "irus ,as initially na#ed as L2V or HTLV III$ 6etro"iruses
res%onsible for 2ID' are %ri#arily of t,o ty%es HIVA0 and HIVA=, these ha"e
been disussed in ha%ters ? and H$
T/0";$>... M-,r)/-0 2"'(s (r's1-(("* (hr)+2h /0))* 6r)*+,(s
V-r+s"s %rs-("s B,("r- (,'
,)'(1-'(" (h"
6r)*+,( 3("r )r *+r-'2
6r),"ss-'2)
A
A
A
A
A
A
A
A
A
A
A
A
HIVA0 and HIVA=
He%atitits B "irus
Delta "irus
He%atitits C "irus
He%atitits 3 "irus
He%atitits 2 "irus
Cyto#egalo"irus
;%stein barr "irus
Hu#an %ar"o"irus BA0B
HTLVAI
HTLVAII
Transfusion trans#itted
"irus &TTV*
A
A
A
A
A
A
)las#odiu# s%%$
Mirofilaria
To/o%las#a gondii
Babesia #iroti
Leish#ania s%%$
Try%anoso#a ruDi$
A
A
A
A
A
A
A
A
A
A
A
A
2erobator
2hro#obater
2laligenes
Bruella s%%$
'ta%hyloous albus
'ta%hyloous
e%ider#idis
Hae#olyti ;sh$ oli
)seudo#onas
;nterobater
:lebsiella
Lersinia
)roteus
T/0";$>.= Tr's1-ss-)' )3 !-r+s"s /9 60s1& ,"00+0r ,)16)'"'(s
%0s1 C"00+0r ,)16)'"'(s
A
A
A
A
A
A
A
HIVA0 and HIVA=
He%atitits B "irus
He%atitits C "irus
He%atitis 3 "iruse
Delta "irus
He%atitits 2 &rarely*
)ar"o"irus
A
A
A
A
HIVA0 and HIVA=
Hu#an TAly#%hotro%i "irus HTLVAI
Hu#an TAly#%hotro%i "irus HTLVAI
;%stein Barr "irus
The first re%ort of trans#ission of 2ID' by fator VIII onentrate ,as
%ublished by CDC 2tlanta in <'2$ Blood transfusion is a "ery effiient #ode
of trans#ission for HIV, as #ore than B> %erent rei%ients of blood fro# HIV
sero%ositi"e donor, de"elo% infetion and 2ID'$ Transfusion indued infetion
#ay lead to 2ID' #ore ra%idly than the infetion trans#itted by other
routes!#odes$ -ortunately transfusion assoiated HIV infetion is dereasing
beause of the %rior sreening of blood %roduts$ There is a %ossibility of
trans#ission of HIV through blood and blood %roduts ta.en fro# donors
during ,indo, %eriod ,hen antibodies to HIV are not detetable$
HTLV I and HTLV II are onsidered to be ono"iruses$ These "iruses are
ly#%ho%roliferati"e and ause #alignany$ These "iruses are trans#itted
#uh in the sa#e ,ay as 2ID' "iruses$ The first detailed re%ort of
transfusion assoiated HTLV I trans#ission ,as fro# 9a%an in 0B1>s$ These
"iruses are trans#itted by transfusion of ellular ele#ents of blood and not by
ell free o#%onents!%roduts$ 'ero%ositi"ity is found ,ithin t,o #onths of
135
e/%osure$
Both HTLV I and HTLV II are single stranded 682 "iruses$ These are sensiti"e
to organi sol"ents, detergents and heat but resistant to <V light and VArays$
Its geno#e odes for "iral oat %rotein in the gagAregion, %oly#erase in the
%ol region and surfae en"elo%e %rotein in the en"elo%e region$ The "irus
binds the hel%er!induer T ells through CDE ree%tors$ It #ay re#ain latent
in the infeted ell for a long ti#e$
S,r""'-'2 ("s(s 3)r HTLV : 'e"eral HTLV assays are a"ailable li.e gelatin
%artile agglutination &)2*, enDy#e i##unoassay &;I2* and indiret
i##unofluoresene &I-*$ The sensiti"ity and s%eifiity of ;LI'2 ranges fro#
B1$EF to 0>>F, res%eti"ely$ 8one of the sreening tests a"ailable is able to
differentiate bet,een anti HTLVAI and anti HTLVAII beause of ross reati"ity
due to the HGF ho#ology of a#ino aid se+uenes bet,een the t,o "iruses$
Confir#atory assays li.e ,estern blot, 6I)2, antibody a%ture radioi##uno
assay and )C6 are also a"ailabe$
$>...$..C9()1"20)!-r+s (CMV) : CMV is one of the largest Her%es "irus
,ith double stranded D82$ This infetion is ,ide s%read and is trans#itted by
a nu#ber of ,ellAreogniDed routes$ The infetion an also be trans#itted by
organ trans%lantation and transfusion of blood!blood %roduts$ Trans#ission
of CMV by blood ,as first re%orted in 0B??$ It is %resu#ed that CMV is
trans#itted by ellular blood o#%onents$
2fter transfusion the "irus gets reati"ated$ The infetion an be se"ere in
i##unoo#%ro#ised %atients and %re#ature infants$ It #ay be fatal in ases
of trans%lantation of allogeni #arro,$ )atients of renal and heart
trans%lantation #ay de"elo% se"ere %ri#ary CMV infetion follo,ed by
transfusion of onta#inated blood ! ellular o#%onents$
Transfusion trans#itted CMV an be he.ed!ontrolled to a greater e/tent
only by de%leting the ,hite ells by filtration or entrifugation$ 2t %resent the
only #ethod to a"oid its trans#ission is by using the CMV negati"e donors$
IgM s%eifi antiACMV ;LI'2 an be used to sreen the %las#a %roduts$
$>...$.=%r!)!-r+s : )ar"o "irus BA0B, a single stranded non en"elo%ed D82 "irus
,as diso"ered in 0BHG in the seru# of nor#al blood donors$ It is a o##on
ause of a blood disease .no,n as erythe#a infetiosu# and #ay be
i#%liated in %lasti anae#ias$ This linial entity #ay be se"ere in %regnant
,o#en, 2ID' ases, and hae#o%hilias et$ -ortunately this "irus is not a
%roble# in India$ Diagnosti #ethods li.e hae#agglutination, gel test, D82
hybridisation and )C6 ha"e been de"elo%ed$
$>....%rs-("s : Most o##on %arasites ,hih #ay be trans#itted through
blood!blood %roduts are #alarial %arasite and To/o%las#a gondii$ 6arely,
others li.e Leish#ania s%%$ and Babesia #iroti et$ #ay also be trans#itted
through transfusion of blood!blood %roduts$
$>.....$M0r-0 6rs-("s : Malaria is a #a7or %ubli health %roble#
throughout the ,orld$ The infetion is aused by a %rotoDoan %arasite of the
genus )las#odiu#$ -our s%eies of )las#odiu# na#ely 1) vivaH, 1)
falci5aram, 1) malariae and 1) ovale are res%onsible for hu#an #alaria$
The %arasite an sur"i"e in the blood of host for #any years follo,ed by
rela%se or rerudesene$ 'uh ases as blood donors %ose %otential threat
of %ost transfusion #alaria$ Transfusion trans#itted #alaria an be lethal$
Quite a good nu#ber of deaths ha"e been re%orted due to transfusion
136
trans#itted #alaria$ To a"oid the transfusion trans#itted #alaria, %ro%er
srutiny and sreening of blood donors should be done$ I##unofluoresene
#ethod for detetion of #alaria has been found to be better than the routine
#iroso%y #ethod$ 8o, enDy#e based i##unodiagnosti assays are
a"ailable o##erially as .its for early diagnosis and sreening of blood$
$>......T)A)60s1 2)'*-- : To/o%las#osis is aused by an obligate intraellular
%rotoDoan %arasite .no,n as To/o%las#a gondii$ The disease is %re"alent
throughout the ,orld$ 3enerally it is trans#itted by ingestion of ooyst ! yst
of %arasite$ The infetion an also be trans#itted by tro%hoDoites in blood
through transfusion$ The infetions initiated by ooyst or yst are in"ariably
#ild but infetion indued by tro%hoDoites of blood stage is #ore se"ere and
in the aute for#$ That tro%hoDoite of %arasite an sur"i"e in blood stored at
E
>
C u%to G> days$ Leu.oyte transfusion has also been assoiated ,ith
trans#ission of To/o%las#osis in i##unoo#%ro#ised %atients$ Both these
obser"ations suggest that no, transfusion assoiated To/o%las#osis is %osing
a serious threat to 2ID' %atients$
$>...=B,("r- : <nli.e the "iruses the diret trans#ission of bateria fro# donor
to rei%ient is less o##on$ Tre%one#a %allidu# has been found to be
assoiated ,ith transfusion indued sy%hilis$ It is disussed in Cha%ter 0@$
$>...=.$B,("r-0 ,)'(1-'(-)' )3 /0))*&/0))* ,)16)'"'(s&/0))*
6r)*+,(s: Handling and storage of blood, its o#%onents and %roduts at
roo# te#%erature as ,ell as at E
>
C #ay %ro"ide suffiient ti#e and
o%%ortunity for baterial onta#ination$ The bateria that ha"e been found to
onta#inate the blood o#%onents and %roduts are 2erobater,
2hro#obater, 2laligenes, Bruella s%%$, 'ta%hyloous albus,
'ta%hyloous e%ider#idis, hae#olyti ;sh$ oli, )seudo#onas s%%$,
;nterobater s%%$, :lebsiella s%%$, Lersinia s%%$ and )roteus et$
Transfusion of blood onta#inated ,ith bateria has a ty%ial linial
syndro#e of "iolent hills during the transfusion or shortly after the
transfusion$ The sy#%to#s #ay be due to endoto/ins$ 'o#eti#es
transfusion of onta#inated blood #ay ause se%ti sho. syndro#e resulting
in death of the rei%ient$ Bateria o##only assoiated ,ith se%ti sho. are
;sh$oli, ;nterobater s%%$, )seudo#onas, :lebsiella s%%$ et$ 'e%tiae#ia
has been re%orted follo,ing the transfusion of %las#a and ryo%rei%itates
et$
In #ost of the ases, onta#ination of blood, its o#%onents!%roduts ours
due to the introdution of bateria at the ti#e of olletion, %roessing,
#anufaturing or storage$ 5ith the i#%ro"e#ent in olletion and storage
#ethods, the onta#ination of blood ,ith bateria has delined signifiantly$
The blood onta#inating organis#s are #ainly ubi+uitous 3ra# negati"e
bateria$ In blood they gro, "ery fast and unfortunately the onta#inated
blood #ay a%%ear nor#al e"en ,ith hea"y onta#ination, as these bateria
are not hae#olyti$ 3ra# %ositi"e bateria ,hih for# a %art of s.in flora
&'ta%hyloous and di%htheroids* are rarely in"ol"ed in transfusion
assoiated diseases$
2%art for# blood, ,hih is a good ulture #ediu# for #iroAorganis#s the
itrate, gluose and antioagulant %reser"ati"e solutions are ready #ade
energy soure for these bateria$ -ortunately, #aro%hages and leu.oytes
%resent in blood ha"e natural bateriidal a%aity$
Baterial onta#ination of blood!%roduts!o#%onents an be %re"ented to a
ertain e/tent by :
i <se of losed blood olletion syste#s
137
ii Transfusion of blood ,ithin ?A1 hrs$ of olletion or re#o"al fro#
refrigeration$
iii )re%aration of blood o#%onents in sterile onditions$
" )ro%er ins%etion of blood unit for hae#oloysis!disolouration of 6BCs$
$>.> %r"!"'(-)' )3 (r's3+s-)' ss),-("* *-s"s"s ,+s"* /9 /0))*
6r)*+,(s&,)16)'"'(s: Blood %roduts!o#%onents ha"e been reognised
as life sa"ing drugs by 5H($ 2 ,ide range of blood o#%onents and blood
%roduts are no, a"ailable that are being #anufatured throughout the
,orld$ Till 0B1>s the #a7or onern of transfusion indued diseases ,as only
he%atitis "iruses but the detetion of 2ID' "irus alerted the entire ,orld to
o#bat the #enae of transfusion assoiated infetions$
-or ontrolling the transfusion assoiated trans#ission of infetions +uality
ontrol!+uality assurane J good laboratory %raties &3L)*, good
#anufaturing %raties &3M)* inluding "iruidal tehnologies and biosafety
%roedures et$ should be i#%le#ented stritly$
$>.>.$G"'"r0 ,)'(r)0 1"s+r"s :
i Transfusion should be gi"en only ,hen there is a lear #edial
indiation$
ii 'reening of units of donated blood and %roduts for all the diseases
notified as already disussed in Cha%ters ?, H and 0@ &HIVJ
he%atitis, Malaria and 'y%hilis*$
iii 2utologous transfusion should be enouraged$
$>.>..G))* 0/)r()r9 6r,(-,"s&2))* 1'+3,("r-'2 6r,(-,"s : To ontrol
suh trans#ission follo,ing #easures should be ta.en$ Trans#ission of
transfusion assoiated infetions an be effeti"ely ontrolled by effeti"e
i#%le#entation of Quality Control!Quality 2ssurane %rogra##e$
i 'eletion of N)r10 H"0(h9 D)')rs$
ii I##unodiagnosti .its!reagents for diagnosing the transfusion assoiated
diseases #ust be "alidated by Quality Control 2uthority!6egional labs$
iii 'trit 3L)!3M) should be follo,ed$
i" 2ll the e+ui%#ents!instru#ents used for transfusion #ust be sterile$
" Transfusion o##ittee #ust be o%erational and should ha"e ,ritten +uality
%lan for transfusion safety and standard o%erati"e %roedures &'()s* for
"arious tests et$
"i )re"ention of baterial onta#ination during donation, olletion, storage and
%roessing of blood$
"ii ;ffeti"e "iral inati"ation of ertain %las#a %roduts!o#%onents$
"iii 9udiious use of blood!blood %roduts!o#%onents$
i/ Quality audit$
/ ;nfore#ent of orreti"e #easures$
/i Biologial safety abinet should be used for handling the biologial #aterials$
/ii )ro%er handling and dis%osal of dis%osables li.e needles, syringes and bags
should be ensured$
/iii ;ffeti"e deonta#inants should be used for leaning!deonta#ination of
,or. %lae$
/i" Blood should be entrifuged in o"ered ontainers$
/" ;+ui%#ents should be deonta#inated before!after re%air!transfer et$
/"i )roteti"e lothings &go,n, #as. et$* should be used$
/"ii Hands should be ,ashed %ro%erly$

$>.? Q+0-(9 C)'(r)0 :
$>.?.$Q+0-(9 ,)'(r)0 )3 "C+-61"'(s '* -11+')*-2')s(-, B-(s&1"(h)*s :
138
The instru#ents under use for storage, handling, %roessing, testing et$
#ust be %ro%erly #aintained and #onitored to ensure 3ood Laboratory
)raties and 3ood Manufaturing )raties &3L)!3M)*$ Biosafety abinet,
auto#ati ell ,ashers, ,ater bath et$ #ust be "alidated$
I##unodiagnosti .its!reagents and #ethods used for sreening of the donor
#ust be e"aluated for Q2!QC for assuring the %ro%er diagnosis of diseases$
$>.?..Q+0-(9 ,)'(r)0 )3 6"rs)''"0 : Quality ontrol of %ersonnel is one of the
#ost i#%ortant as%ets of +uality ontrol %rogra##e as it is entral %oint
around ,hih the entire %roess re"ol"es$ )ersonnel also regulate the entire
%roess of Q2!QC$
2ll %ersonnel should be :
i Trust,orthy$
ii )ro%erly +ualified!trained and able to ,ithstand stressful situations$
iii 2+uainted ,ith '()M$
i" Tested ,ith %rofiieny testing$
" 'hould be trained regularly through ontinuing #edial eduation
%rogra##es$
"i 'hould be gi"en suitable inenti"e!o##endation fro# ti#e to ti#e$
$>.?.=Q+0-(9 ,)'(r)0 )3 /0))* ,)16)'"'(s&C+0-(9 ,)'(r)0 )3 6r),"*+r"s :
Quality Control )rogra##e should be strengthened in 3L)!3M) ,hih onsist
of &i* olletion of blood &ii* testing &iii* se%aration for o#%onents &i"*
%re%aration of blood %roduts &"* "iral inati"ation &"i* %ro%er storage &"ii*
safe transfusion in %atients and &"iii* i#%at assess#ent$
There are si/ i#%ortant ste%s of +uality ontrol in transfusion #ediine
ser"ies$
i I##unohae#atologial in"estigations$
ii Mirobiologial in"estigations$
iii Blood olletion
a* Donor reruit#ent
b* DonorCs in"estigations
i" 'e%aration of blood o#%onents and %re%aration of blood %roduts$
" 'torage$
"i <se of assoiated e+ui%#ents$
139
$?. COUNSELLING AND HIV&AIDS TESTING
$?.$ D"3-'-(-)' : Counselling is a %roess ,here one %erson, e/%liitly and
%ur%osefully, gi"es his!her ti#e, attention and s.ills to assist a lient to
e/%lore their situation, identify and at u%on solutions ,ithin the li#itations of
their gi"en en"iron#ent$ In si#%le ter#s, ounselling is a %roess in ,hih
t,o %eo%le #eet and ha"e a dialogue to resol"e a risis, sol"e a %roble# or
#a.e deisions in"ol"ing highly %ersonal and inti#ate #atters and
beha"iours$
5orld Health (rganisation &5H(* defines HIV!2ID' ounselling as a dialogue
bet,een a lient and a are %ro"ider ai#ed at enabling the lient to o%e ,ith
stress and to ta.e %ersonal deisions relating to HIV!2ID'$ The ounselling
%roess inludes the e"aluation of %ersonal ris. of HIV trans#ission and the
failitation of %re"enti"e beha"iour$
$?.. D-33"r"'," /"(8""' ,)+s"00-'2 '* h"0(h "*+,(-)' : Counselling differs
fro# health eduation in #any ,ays and they are :
i Counselling is usually an Kone is to oneC %roess ,hereas, health eduation
addresses a grou% of %eo%le$
ii Counselling is useful for not only gi"ing infor#ation but also for hanging
attitudes and #oti"ating beha"ioural hange$ Health eduation is used #ostly
for infor#ation sharing$
iii Counselling sessions in"ol"e %ersonal %roble# sol"ing$ In health eduation,
general issues are disussed$
i" Counselling is #ore foussed, s%eifi and goal targeted, ,hereas health
eduation is #uh #ore generaliDed$
" Counselling e"o.es strong e#otions in both ounsellor and lient ,hereas
health eduation sessions are e#otionally neutral in nature$
$?.= O/#",(-!"s )3 HIV&AIDS ,)+'s"00-'2 : The ob7eti"es of HIV!2ID'
ounselling inlude :
i )re"ention of HIV infetion by hanging life styles and beha"iours$ &)ri#ary
%re"ention*$
ii )ro"iding %syhologial su%%ort to those already infeted$
iii Moti"ating for hange in high ris. beha"iour & 'eondary %re"ention*$
$?.> Wh9 -s HIV&AIDS ,)+'s"00-'2 '","ssr9 : HIV!2ID' ounselling is
#andatory for %ro"iding "oluntary HIV testing ser"ies$ This is so beause
diagnosis of HIV in an other,ise healthy indi"idual indues a series of
%syhologial reations li.e denial, anger, an/iety, de%ression, to finally
ae%tane$ 'o far, there is no suessful ure or "aine a"ailable for HIV
infetion$ In order for the indi"idual to be able to ae%t the infeti"e status,
arry on ,ith life, %lan future, %re"ent trans#ission and ontinue to funtion
as a useful #e#ber of o##unity, ounselling is a #ust$ Counselling indues
a %ositi"e attitude and high life fore in the indi"idual hel%ing hi# to arry on
as before ins%ite and irres%eti"e of the HIV infetion, HIV!2ID' ounselling is
#andatory &%re and %ost test* as laid do,n by 5H(!<82ID'$ 3o"ern#ent of
India is also ati"ely e#%hasising the neessity of HIV ounselling$
140
$?.? Wh) r"C+-r"s HIV&AIDS ,)+'s"00-'2P : HIV!2ID' ounselling is
reo##ended for the follo,ing grou%s of %eo%le :
i )ersons already identified as ha"ing 2ID' or being infeted ,ith HIV and their
fa#ilies$
ii Those being tested for HIV &%re and %ost test*$
iii Those see.ing hel% beause of %ast or urrent ris. beha"iour and %lanning
their future$
i" Those not see.ing hel% but %ratiing high ris. beha"iour$
$?.F Wh"r" ,' HIV&AIDS ,)+'s"00-'2 /" 6r)!-*"*P : HIV!2ID' ounselling
an be %ro"ided in any setting inluding hos%ital ,ards, 'TD linis, 28C
linis, fa#ily %lanning linis, blood donation enters, drug deaddition
enters, %risons, %ri#ary!seondary health enters and o##unity based
%rogra##es$
$?.G WHO ,' 6r)!-*" HIV&AIDS ,)+'s"00-'2P : HIV!2ID' ounselling an be
%ro"ided by anyone ,ho has a sy#%atheti ear, an gi"e ti#e to listen, has
.no,ledge of aurate sientifi fats about HIV!2ID' and undergoes
syste#ati and %eriodi training in ounselling$ In addition to dotors, nurses,
%syhologists, %syhothera%ists and soial ,or.ers, e"en teahers,
o##unity and %eer leaders, youth and self hel% grou%s an underta.e
%re"enti"e and su%%orti"e ounselling$
$?.5 %r";r"C+-s-("s )3 ,)+'s"00-'2 : 'o#e of the %reAre+uisites of ounselling
inlude :
i Ti#e and a"ailability of ounsellor
ii 2e%tane of the ati"ity by ounsellor
iii ;asy aessibility to ounselling
i" 2%titude for #aintenane of onfidentiality
" Corret .no,ledge and infor#ation about HIV!2ID' and %oliies et$
"i Consisteny in ounselling
$?.4 T96"s )3 ,)+'s"00-'2:
i )reAtest
ii )ost testA%ositi"e, negati"e and indeter#inate results$
iii -ollo,Au%
i" 3eneral
" -a#ily
$?.$I C)'("'(s )3 6r"!"'(-!" HIV ,)+'s"00-'2 :
i Deter#ine ,hether the beha"iour of an indi"idual or grou% of indi"iduals
in"ol"es a high ris. beha"iour ,hih an lead to HIV infetion$
ii 5or. ,ith indi"iduals to #a.e the# understand the ris.s and a.no,ledge
the ris.s assoiated ,ith their beha"iour$
iii Define and disuss ,ith the# ho, their life, attitude!"alues and self i#age is
lin.ed to these beha"iours$
i" Hel% indi"iduals to define their %otential for attitude shifts, beha"iour
#odifiation and hange$
" 5or. ,ith indi"iduals to introdue and sustain the #odified beha"iour$
$?.$$ C)+'s"00-'2 : HIV!2ID' is a life long disease ,hih to date, has no ure or
"aine$ In addition, la. of a healthy non 7udg#ental and non disri#inatory
en"iron#ent has resulted in isolation, "iti#isation and breah of
funda#ental rights inluding denial of basi #edial ser"ies to %eo%le ,ith
HIV infetion$ HIV!2ID' ounselling an hel% %eo%le in aessing orret
infor#ation, assessing ris.s, #a.ing a%%ro%riate beha"ioural hanges, leading
141
to %rotetion of self and others$ This ,ill hel% in de"elo%ing o%ing
#ehanis#s in %eo%le ,ith HIV infetion$ Counselling an lead to
e#%o,er#ent and raise indi"idual onsiousness and an #a.e indi"idual
ta.e res%onsibility for their o,n beha"iour$
142
%?) INF*AST*UCTU*E FO* HI0 TESTING LABO*ATO*IES
%?)% Introd6ction ; 'sta7lis:8ent o6 o4ti8al in6rastr5ct5re 6or +I& testing
la7oratories is one o6 t:e 8ost i84ortant co84onents to ens5re
B5ality testing. +o<e9er1 t:e s4eci6ications regarding t:e reB5ired
in6rastr5ct5re cannot 7e 8ade e!cl5si9e and rigid in a de9elo4ing
co5ntry li=e India <it: li8ited reso5rces. or instance1 t:e sa8e
',ISA reader and <as:ing syste8 8ay 7e 5tilised 6or screening 7ot:
+I& and +2& <it:in t:e co88on s4ace and 7y t:e co88on
8an4o<er @e.g. tec:nicianA. T:e o4ti8al in6rastr5ct5re 6or di66erent
categories o6 +I& testing la7oratories is descri7ed :ere.
%?)" Cate#orie! of HI0 te!tin# la=oratorie!8centre! ; +I& testing la7oratories
:a9e 7een categorised into 6ollo<ing 8ajor gro54s 7y National AIDS
Control Organisation1 %OI.
%?)")% Screenin# la=oratorie! in =lood =an3! ; T:e 8ain 65nction o6 t:ese
la7oratories is to ens5re donated 7lood sa6ety. T:5s t:e la7oratories
in 7lood 7an=s in addition to ot:er testings e.g. gro54ing1 cross
8atc:ing 85st screen t:e donated 5nits o6 7lood 6or trans65sion
trans8itted in6ections @TTIA i.e. +I&1 +2& @+2sAgA1 sy4:ilis and
8alaria as 4er t:e e!isting g5idelines 6ro8 %OI. T:ese la7oratories
are also called >Gonal 2lood Testing Centres? @G2TCsA.
%?)")" Sentinel !6rveillance la=oratorie! and vol6ntar+ te!tin# la=oratorie! ; T:ese
categories o6 la7oratories are 65nctioning in 9ario5s 8edical colleges1
:os4itals and instit5tes. T:ese la7oratoires carry o5t 9ol5ntary
testing @<it: 4re and 4ost test co5nsellingA and testing 6or
s5r9eillance. 2eing located in 8edical colleges1 :os4itals and
instit5tes1 t:e necessary in6rastr5ct5res 6or +I& testing in t:ese
la7oratories are li=ely to 7e in 4lace.
%?)")< *eference centre! ; Presently t:ere are D re6erral centres distri75ted
all o9er India t:at are entr5sted <it: t:e 6ollo<ing 7road
res4onsi7ilities:
3 Pro9iding re6erral +I& diagnostic ser9ices in t:e region.
3 '9al5ation o6 +I& test =its.
3 +I& screening o6 7lood 4rod5cts 6or :58an 5se.
3 Carrying o5t e!ternal B5ality ass5rance o6 ot:er la7oratories
and tro57le s:ooting.
3 Pro9iding training in +I& testing and 7iosa6ety 4roced5res.
3 Pro9iding tec:nical s544ort 6or 6or85lating g5idelines 6or
B5ality ass5rance and ot:er +I& testing related iss5es.
3 &ol5ntary testing <it: co5nselling.
3 Pro9iding +I& testing ser9ices 6or 9ario5s gro54s as directed
7y NACO1 %.O.I. 6ro8 ti8e to ti8e.
3 Pro9iding diagnostic ser9ices @incl5ding trainingA 6or
o44ort5nistic in6ections.
In India1 t:e a7o9e t:ree categories are already 65nctioning1 75t <it:
9arying degrees o6 in6rastr5ct5re. T:e 9ario5s le9els o6 in6rastr5ct5re
necessary 6or t:e 9ario5s categories o6 +I& testing
la7oratories;centers are 8entioned in Ta7le 16.
143
Ta=le No)%? Nece!!ar+ infra!tr6ct6re for vario6! cate#orie! of HI0 te!tin# la=oratorie!8centre!
Cate#or+ of HI0
te!tin#
la=oratorie!8centre
S5aceK Man5o.er EC6i5ment! and !655lie! Comment
1. 2lood 2an= T<o roo8s
@a44ro!i8ately 1#S1#
6t.A
(oo8 1 6or
3 45tting 54 t:e test
3 storage o6 sa84les
3 4lace8ent o6
eB5i48ents
3 4rocessing o6
sa84les
3 record =ee4ing
(oo8 # 6or
3 deconta8ination and
dis4osal o6 la7oratory
<astes.
S54er9ising )edical
O66icer 31
Tec:nician;,a7oratory
assistant 3 1
,a7oratory attendant
c583cleaner 3 1
Bio!afet+;
,a7. coats;go<ns @6ront closedA
C:a44als;sli44ers @to4 co9eredA
%lo9es
Sodi58
:y4oc:lorite;et:anol;gl5taralde
:yde
Discarding jars
Dis4osa7le a7sor7ent 4a4er
s:eet @6or s4reading on t:e
<or=ing areaA
Dis4osa7le 7ags @6or carrying
in6ectio5s <astesA
A5tocla9e1 4re6erra7ly
incinerator
+ot air o9en.
Needle destroyer;s:redder
Collection of !am5le! ;
Scre<ca44ed lea= 4roo61
non7rea=a7le t57es
Centri65ge 6or se4aration o6
ser58
La=orator+ te!tin# ;
.ater 7at: @3$3""
0
CA
Inc57ator 2OD @3$
0
3""
0
CA
2alance @4:ysicalA
2alance @electricalA
(oc=er @"0360 roc=ings;8in5teA
',ISA reader
',ISA <as:ing syste8 i.e.
a5to8atic ',ISA <as:er
ilters 6or ',ISA reader @"0$ n81
or acti9ities ot:er
t:an +I& testing in
7lood 7an=s
additional
in6rastr5ct5re <ill
7e necessary.
144
"$0 n81 "D# n81 $"0 n8A
',ISA 4a4er role
%lass 4i4ettes 3 #8l1 $8l1 10 8l
)icroti4s @ca4acity #$ l1 $0 l1
100 lA
)icroti4 container
)5ltic:annel 4i4ettes @ $0 3 #00
l A
)icro 4i4ettes @$ 3 $0 l1 $0 3
#00 l1 #00 3 1000 l A
Pi4ette stands
Paste5r 4i4ette1 r577er teats
Plastic 6orce4s 1 4lastic tro5g:s
Tiss5e 4a4er
+I& screening test =its
@',ISA;ra4idA
For !tora#e of 3it! and rea#ent! ;
(e6rigerator @do8estic <it:
6reeJer co84art8entA
Dee4 6reeJe @ 3 #0
0
CA
Plastic storage 9ials
(ac=s 6or :olding storage 9ials
#. &ol5ntary
testing
la7oratories;sen
tinel
s5r9eillance
la7oratories
T.o room! a! in % a=ove
5l6!
an additional roo8 6or
3 sa84le recei9ing and
4rocessing
@Total n587er o6
roo8s U 3A
A! % a=ove 5l6!
Co5nsellor 3 1
ield <or=er 31
@6or collection o6
sa84les etc.A
A! for % a=ove 5l6!
For collection of !am5le!
3 Dis4osa7le syringes @#8l1 $8l1
108l1A or 9ac5tainers <it: gel
3 Dis4osa7le needles @#1 ga5geA
For *eferral
3 Sa84le carrier @e.g. t:er8ocol
7o!A
3. +I& re6erence
la7oratories;
centres
Tree room! a! in "
a=ove 5l6!
an additional roo8
6or3Co5nselling
@Total n587er o6
roo8s U "A
A! a! in " a=ove 5l6!
S54er9ising )edical
O66icer 31
Tec:nician 3 1
,a7oratory assistant 3
#
A! for " a=ove 5l6!
A5to8ated ',ISA syste8 in
4lace o6 se8i a5to8ated ',ISA
syste8
.estern 7lot @4re6era7ly
a5to8atedA
Dee4 reeJe 370
0
C
'B5i48ent and reagents 6or PC(
Note: so8e o6 t:e
s4ecialised
la7oratories;
centres 8ay need to
5nderta=e c5lt5re
6acilities 6or +I&.
T:e necessary
in6rastr5ct5res are
145
@as 8entioned in C:a4ter EA. ela7orate 6or +I&
c5lt5re and are
7eyond t:e sco4e o6
disc5ssion in t:is
8an5al.
KNote ; At 4resent t:ere is :ardly any doc58ented g5ideline regarding t:e reB5ire8ent o6 s4ace 6or +I& testing la7oratories in India.
T:5s1 7ased on t:e cons5ltations a8ong t:e e!4erts an atte84t :as 7een 8ade to 4ro9ide so8e idea in t:is regard. +o<e9er1 so8e
7road co84onents regarding s4eci6ication o6 roo8s1 6or o4ti8al 4er6or8ance o6 t:e la7oratory are 8entioned 7elo<:
Broad !5ecification! of te room! ;$
(oo8s s:o5ld 7e adeB5ately 9entilated and <ell lig:ted.
loors s:o5ld 7e i84er9io5s and easy to clean.
T:ere s:o5ld 7e no o4en drainage syste8.
(oo8s s:o5ld 7e 8ade rodent 6ree and insect 6ree.
All roo8s s:o5ld 7e eit:er centrally air3conditioned or 6itted <it: <indo< air3conditioners.
(oo8s 6or g5est1 6or eating etc s:o5ld 7e a<ay 6ro8 t:e la7oratory area.
AdeB5ate n587er o6 electricity 4oints s:o5ld 7e 4ro9ided so t:at any eB5i48ents 45rc:ased later on can 7e 5sed <it:o5t
additional electrical <or=s.
146
%?)")( La=oratorie! monitorin# te 5ro#re!!ion of HI0 infection ; T:ese la7oratories
carry o5t tests <:ic: can acc5rately assess t:e stage o6 +I& in6ection
in an indi9id5al as <ell as 4redict its co5rse and 8onitor its
4rogression. T:ese tests are 9al5a7le d5ring t:e 4eriod o6 clinical
latency in 8onitoring disease 4rogression and e9al5ating res4onse to
antiretro9iral t:era4y.
T:e details o6 t:e in9estigational set54s necessary 6or 8onitoring t:e
stage o6 disease in +I& in6ected indi9id5als :a9e 7een disc5ssed in
details in c:a4ter 1#. T:e 8ain eB5i48ents;6acilities reB5ired are
1
.
lo< cyto8etry set 54
6or
3 'n58eration o6 4eri4:eral ly84:ocyte
s574o45lations.
@ s4ecially CD" ly84:ocyte co5nts A
#
.
PC( set 54 6or 3 K5antitating 9iral load
3
.
Diagnostic =its 6or 3 4#" antigen assay
3 Ser58 3# 8icroglo75lin assay
3 Ser58 3neo4terin assay
"
.
S=in testing 4anel
antigens
and a44licators 6or
3 Delayed c5taneo5s :y4ersensiti9ity testing.
%?)")' Dia#no!i! of o55ort6ni!tic infection! ; )ost o6 t:e in6rastr5ct5re necessary
6or diagnosis o6 s4ectr58 o6 o44ort5nistic in6ections associated <it:
+I&;AIDS are a9aila7le in a 85ltidisci4linary :os4ital. So8e o6 t:e8
t:at are rele9ant 6or +I&;AIDS 4atients are 8entioned 7elo< :
1. (o5tine 9irology la7oratory.
#. I885no6l5orescent 8icrosco4y 6acilities and necessary
reagents.
3. 2acterial c5lt5re @anaero7ic and aero7icA.
". T57erc5losis la7oratory.
$. (o5tine :isto4at:ology set 54.
6. S3ray eB5i48ent1 6il8 and reagents 6or 6il8 4rocessing.
7. 'B5i48ent 6or 5ltrasonogra4:y1 7ronc:osco4y 1 endosco4y.
E. )(I eB5i48ent1 CT eB5i48ent.
B$ (%hthal#oso%e$
'ine, this dou#ent is on HIV testing the details of infrastrutures for abo"e
#entioned laboratories ha"e not been inluded in this dou#ent$
147
%7) 1OST E91OSU*E 1*O1H:LA9IS GUIDELINES
%7)% Introd6ction ; +ealt: care <or=ers @+C.A are nor8ally at a 9ery lo<
ris= o6 acB5iring +I& in6ection d5ring 8anage8ent o6 t:e in6ected
4atient. +o<e9er1 ins4ite o6 a lo< statistical ris= o6 acB5isition o6 +I&1
t:e a7sence o6 a 9accine or e66ecti9e3c5rati9e treat8ent1 8a=es t:e
:ealt: care <or=er a44re:ensi9e. So1 it is 9ery necessary to :a9e a
co84re:ensi9e 4rogra88e in 4lace to deal <it: antici4ated accidental
e!4os5re.
)ost e!4os5res do not res5lt in in6ection. T:e ris= o6 in6ection 9aries
<it: ty4e
o6 e!4os5re and ot:er 6actors s5c: as:
3 T:e a8o5nt o6 7lood in9ol9ed in t:e e!4os5re
3 T:e a8o5nt o6 9ir5s in 4atient0s 7lood at t:e ti8e o6 e!4os5re
3 .:et:er 4ost e!4os5re 4ro4:yla!is @P'PA <as ta=en <it:in t:e
reco88ended ti8e.
1revention is t:e 8ainstay o6 strategy to a9oid occ54ational e!4os5re to
7lood;7ody 6l5ids. All t:e 7iosa6ety 4reca5tions e84:asiJed in
c:a4ter " 85st 7e 4ractised at all ti8es 6or all 4atients1 7lood and 7ody
6l5ids <:ile 4ro9iding 8edical ser9ices.
%7)" Definition of an occ65ational eH5o!6re ; An occ54ational e!4os5re t:at 8ay
4lace a <or=er at ris= o6 +I& in6ection is a 4erc5taneo5s inj5ry1 contact
o6 85co5s 8e87rane or contact o6 s=in @es4ecially <:en t:e s=in is
c:a44ed1 a7raded or a66licted <it: der8atitis or t:e contact is
4rolonged or in9ol9ing an e!tensi9e areaA <it: 7lood1 tiss5e or ot:er
7ody 6l5ids to <:ic: 5ni9ersal 4reca5tions a44ly.
%7)< Ste5! to =e ta3en on eH5o!6re to HI0 infected =lood8=od+ fl6id! and
contaminated !ar5! etc) ;
I88ediately 6ollo<ing an e!4os5re :
3 Needlestic=s and c5ts s:o5ld 7e <as:ed <it: soa4 and <aterF
3 S4las:es to t:e nose1 8o5t: or s=in1 s:o5ld 7e 6l5s:ed <it:
<aterF
3 'yes s:o5ld 7e irrigated <it: clean <aterF saline1 or sterile
irrigantsF
3 Pric=ed 6inger s:o5ld not 7e 45t into 8o5t:1 re6le!ly.
No scienti6ic e9idence e!ists as to t:e 6act t:at t:e 5se o6 antise4tics
6or <o5nd care or sB5eeJing t:e <o5nd <ill red5ce t:e ris= o6
trans8ission o6 +I&. +o<e9er1 t:is 85st al<ays 7e done. T:e 5se o6
a ca5stic agent s5c: as 7leac: is not reco88ended @Anne!5re 17A.
3 (e4ort t:e e!4os5re to t:e a44ro4riate a5t:ority and condition
85st 7e treated as an e8ergency. Pro84t re4orting is essential
7eca5se in so8e cases1 +I& 4oste!4os5re 4ro4:yla!is @P'PA 8ay 7e
reco88ended and it s:o5ld 7e started as soon as 4ossi7le1 4re6era7ly
<it:in a 6e< :o5rs.
148
2ased on ani8al 8odels1 t:e s5ccess o6 P'P t:era4y is re4orted to 7e
8a!i8al <:en started <it:in a 8atter o6 :o5rs a6ter t:e e!4os5re.
Alt:o5g:1 any c5to66 ti8e is ar7itrary initiating treat8ent 8ore t:an 7#
:o5rs a6ter t:e e!4os5re is not reco88ended. Alt:o5g: 4er:a4s not
as e66ecti9e as 4ro4:yla!is1 late P'P @a6ter 7# :o5rsA 8ay still 7e 5se65l
as early treat8ent o6 +I& in6ection1 in case in6ection :as occ5red.
%7)( T+5e! of occ65ational eH5o!6re! to HI0 for .ic 1E1 i! recommended ; )ost
occ54ational e!4os5res do not lead to +I& in6ection. T:e c:ance o6
4ossi7le serio5s side e66ects @to!icityA o6 t:e dr5gs 5sed to 4re9ent
in6ection 8ay 7e 85c: greater t:an t:e c:ance o6 +I& in6ection 6ro8
so8e =inds o6 e!4os5res. 2ot: ris= o6 in6ection and 4ossi7le side
e66ects o6 dr5gs s:o5ld 7e care65lly considered <:en deciding <:et:er
to ta=e 4oste!4os5re 4ro4:yla!is. '!4os5res <it: a lo<er in6ection ris=
8ay not 7e <ort: t:e ris= o6 t:e side e66ects associated <it: t:ese
dr5gs. T:e decision to start P'P is 8ade on t:e 7asis o6 degree o6
e!4os5re to +I& and +I& stat5s o6 t:e so5rce 6ro8 <:o8
e!4os5re;in6ection :as occ5rred: @ig. 17.11 17.#1 17.3A.
149
Fi#) %7)% Determination of te EH5o!6re Code 2EC4
Is t:e so5rce 8aterial 7lood1 7loody 6l5id1 ot:er 4otentially
in6ectio5s 8aterial @OPI)A1 or an instr58ent
conta8inated <it: one o6 t:ese s57stances L
*es No No 1E1 needed
OPI) 2lood or 7loody 6l5id
.:at ty4e o6 e!4os5re :as occ5rred L

)5co5s 8e87rane or Intact s=in
Perc5taneo5s
s=in1 integrity co84ro8ised only
e!4os5re
&ol58e No 1E1 Needed Se9erity
Small Lar#e Le!! Severe More
Severe
@e.g.1 6e< @e.g.1 se9eral @e.g.1 solid
@e.g.1 large37ore
dro4s1 dro4s1 8ajor needle1 :ollo<
needle1 s:ort 7lood s4las: s54er6icial
dee4 45nct5re1 d5rationA and ;or longer
scratc:A 9isi7le 7lood
d5ration @i.e.1 on
de9ice1 or
se9eral 8in5tes needle
5sed in so5rce
or 8oreA 4atient0s artery
or 9ein
'C1 'C# 'C# 'C3
150
Fi#) %7)" Determination of te HI0 Stat6! Code 2HI0 SC4
T:e +I& stat5s o6 t:e e!4os5re so5rce

+I& negati9e +I& 4ositi9e Stat5s
So5rce
5n=no<n 5n=no<n

No 1E1 needed
,o<er titer e!4os5re +ig:er titer e!4os5re
@e.g.1 asy84to8atic @e.g.1 ad9anced AIDS1
and :ig: CD" co5ntA 4ri8ary +I& in6ection1 :ig:
or increasing 9iral load or
lo< CD" co5ntA
+I& SC 1 +I& SC# +I& SC
Cn=no<n
Fi#) %7)< Determine te 1E1 recommendation
EC HI0 SC 1E1 recommendation
1 1 P'P 8ay not 7e <arranted
1 # Consider 7asic regi8en.
'!4os5re ty4e 4oses a negligi7le ris= 6or +I&
trans8ission.
# 1 (eco88end 7asic regi8en
)ost +I& e!4os5res are in t:is category F no increased
ris= 6or +I& trans8ission :as 7een o7ser9ed 75t 5se o6
P'P is a44ro4riate.
# # (eco88end e!4anded regi8en.
'!4os5re ty4e re4resents an increased
+I& trans8ission ris=.
3 1 or # (eco88end e!4anded regi8en.
'!4os5re ty4e re4resents an increased
+I& trans8ission ris=.
CNHNO.N I6 t:e so5rce1 @in t:e case o6 an 5n=no<n so5rceA1 t:e
setting <:ere t:e e!4os5re occ5rred s5ggests a
4ossi7le ris= 6or +I& e!4os5re and t:e 'C is # or 31
consider P'P 7asic regi8en.
151
%7)' 1re and 1o!t te!t co6n!ellin# and te!tin# ; T:e 4erson s:o5ld 7e
4ro9ided <it: 4re test co5nselling and AGT 7e started as disc5ssed
a7o9e. 2e6ore starting AGT1 33$ 8l o6 4erson0s re6erence 7lood
sa84le is ta=en and tested 6or anti3+I& anti7odies i88ediately a6ter
t:e e!4os5re. In case t:e sa84le tests 4ositi9ed as 4er t:e strategy
o6 +I& testing t:e indi9id5al is re6erred to t:e clinician 6or
8anage8ent1 as a case o6 +I& in6ection. In case t:e sa84le tests
non reacti9e1 a #
nd
sa84le is collected at 6 <ee=s and 3
rd
at 1#
<ee=s a6ter t:e e!4os5re and tested 6or +I&3anti7odies. T:e
6acilities 6or (T3PC( are a9aila7le 4resently at )C )edical College1
)587ai and NA(I1 P5ne and AII)S1 Ne< Del:i and t:is can gi9e 5s
res5lts e9en at #
nd
or "
t:
<ee=s a6ter e!4os5re. Post test co5nselling
is done in all cases.
D5ring t:e 6ollo<354 4eriod1 es4ecially t:e 6irst 631# <ee=s
<:en 8ost in6ected 4ersons are e!4ected to s:o< signs o6 in6ection1
t:e reco88endations 6or 4re9enting trans8ission o6 +I& are to 7e
6ollo<ed 7y t:e +C.. T:ese incl5de re6raining 6ro8 7lood1 se8en1
organ donation and a7staining 6ro8 se!5al interco5rse. In case
se!5al interco5rse is 5nderta=en a late! condo8 85st 7e 5sed
correctly and consistently. T:is red5ces t:e ris= o6 +I& trans8ission.
In addition1 <o8en s:o5ld not 7reast36eed t:eir in6ants d5ring t:e
6ollo<354 4eriod a6ter e!4os5re to 4re9ent e!4osing t:eir in6ants to
+I& in 7reast 8il=.
%7)? Dr6#! recommended for 5o!teH5o!6re 5ro5+laHi! 8 treatment ; It is
reco88ended t:at in India Jido95dine @GD&A and la8i95dine @3TCA
7e 5sed as 6ollo<s. 2ot: t:ese dr5gs s:o5ld 7e considered 6or
treat8ent o6 all e!4os5res in9ol9ing +I&3in6ected 7lood1 6l5id
containing 9isi7le 7lood1 or ot:er 4otentially in6ectio5s 6l5ids or
tiss5es @GD&3#00 8g S E :lyA. Csed in co87ination1 GD& and 3TC
are 9ery e66ecti9e in treating +I& in6ection a6ter e!4os5re and
considera7le in6or8ation s:o<s1 t:at t:ey are sa6e <:en 5sed 6or a
s:ort ti8e @,a8i95dine 1$0 8g S 1# :rlyA.
In selected cases @+I& stat5s code to and 'C code # and 3A1
Indina9ir @or one o6 t:e ot:er 4rotease in:i7itorsA is also 7eing 5sed as
4er P'P g5ideline.
%7) 7 D6ration for .ic dr6#! need to =e ta3en ; T:e o4ti8al co5rse o6
treat8ent is 5n=no<nF since " <ee=s o6 GD& a44ears to 4ro9ide
4rotection against +I& in6ection1 i6 tolerated1 treat8ent s:o5ld
4ro7a7ly 7e ta=en 6or " <ee=s.
%7)@ 1re#nanc+ and 1E1 ; 2ased on li8ited in6or8ation1 GD& ta=en d5ring #
nd
or 3
rd
tri8esters o6 4regnancy :as not ca5sed serio5s side e66ects in
8ot:ers or in6ants. T:ere is 9ery little in6or8ation on t:e sa6ety o6 GD&
<:en ta=en d5ring t:e 1
st
tri8ester or on t:e sa6ety o6 ot:er anti9iral
dr5gs ta=en d5ring 4regnancy. I6 t:e +C. is 4regnant at t:e ti8e o6
occ54ational e!4os5re to +I&1 t:e designated a5t:ority;4:ysician 85st
7e cons5lted a7o5t t:e 5se o6 anti39iral dr5gs 6or 4ost e!4os5re
treat8ent.
%7)A Fact! 3no.n a=o6t te !afet+ and !ide effect! of te!e dr6#! ; )ost o6 t:e
in6or8ation =no<n a7o5t t:e sa6ety and side e66ects o6 t:ese dr5gs is
7ased on st5dies o6 t:eir 5se in +I&3in6ected indi9id5als. or t:ese
152
indi9id5als1 GD& and 3TC :a9e 5s5ally 7een tolerated <ell e!ce4t 6or
na5sea1 9o8iting1 diarr:oea1 tiredness1 or :eadac:e 6or 4eo4le ta=ing
GD&.
%7)%& Ste5! to =e 6nderta3en =+ te infection control officer on receivin# information
a=o6t occ65ational eH5o!6re ;
3 All t:e needle stic= inj5ries s:o5ld 7e re4orted to t:e AIDS State
Society gi9ing t:e e!4os5re code and t:e +I& stat5s code in a
4ro6or8a enclosed @Anne!5re 17.1A.
3 T:e State AIDS Societies s:o5ld in t5rn in6or8 NACO a7o5t t:e cases
4eriodically.
3 A registry is 4lannned to 7e o4ened in NACO soon1 6or 6ollo<354 o6 all
s5c: cases.
3 NACO :as decided in 4rinci4le to s544ly antiretro9iral dr5gs to all cases
6or 4ost e!4os5re 4ro4:yla!is in %o9t. :os4ital settings 6or +C..
3 In6ection control o66icers in all :os4ital :a9e 7een directed to ens5re
t:at anti retro9iral dr5gs 6or P'P are a9aila7le in cas5alty at all t:e
ti8es.
153
Ca5ter %7 $ AnneH6re $ "
Pro6or8a 6or re4orting e!4os5re to 7lood;7ody 6l5ids.
Needle Stic3 Sar5 In>6r+ 1rotocol
Na8e o6 +.C... :
Section o6 +C. :
'84loy8ent No. :
Date o6 Needle Stic=;S:ar4 Inj5ry :
Date o6 (e4orting to Cas5alty :
Site - De4t: O6 Inj5ry
Nat5re o6 Inj5ry : Needle Pric= ; S:ar4 C5t ; ,aceration ;
S4las: O6 l5ids ;
S4lattered %lass
Action ta=en in cas5alty
+e4. 2. 9accination gi9en *es ; No
+2I% *es ; No
Hi!tor+ of in>6r+
+istory O6 +e4atitis 2 I885niJation : Date: Intrader8al ;
Intra85sc5lar
Anti +2s Titre @Date o6 recent testingA
01. +2sAg *es ; No
0#. +I& *es ; No
So6rce Of In>6r+ 2If Availa=le4
Ser58 sent 6or: @re4orts to 7e entered in 65ll: 54on 9isitA
01. +I&
>=$ HBs2g
154
%@) TESTS -HICH HA0E BEEN E0ALUATED B: -HO COLLABO*ATING CENT*E ON
AIDS IN THE
DE1A*TMENT OF MIC*OBIOLOG:, INSTITUTE OF T*O1ICAL MEDICINE, ANT-E*1,
BELGIUM
T:e list o6 assays gi9en 7elo< @',ISA;(a4id;S544le8entalA is rele9ant 6or
5se in India as t:ese assays detect anti7odies to 7ot: +I&31 as <ell +I&3#.
T:e list :as 7een 4re4ared 6ro8 .+O doc58ent n587er .+O;2,S;DE.1 <it:
d5e 4er8ission 6ro8 .+O.
%@)% ELISA8EIA For te detection of anti=od+ to HI0$% and HI0$"
Man6fact6rer Sen!itivit+ S5ecificit+ val6e! *eader
8
vi!6al
2*804
S6ita=ilit
+
-BL
!era
-B$!era
Detect3+I&
T)
@2ioc:e8
I885nosyste8sA
100.0
@DE.63
100.0A
D7."
@D".03
DD.#A
1#.6$ 3#.#1 ( ,S
D5Pont +I&31;+I&3#
',ISA
@D5Pont de Ne8o5rsA
100.0
@DE.73
100.0A
E$.6
@7D.E3
D0.#A
D.3" 30.D6 ( ,S
Anti3+I&31;+I&3# 'IA
P(oc:eO
@. +o668an3,a(oc:eA
100.0
@DE.73
100.0A
D6.D
@D3."3
DE.DA
11.30 3#.37 ( ,S
.ellcoJy8e +I&31R+I&3
#
@.ellco8e DiagnosticsA
100.0
@DE.73
100.0A
D6.D
@D3.33
DE.DA
3E.$1 31.DD ( ,S
Pe4tide +I& ',ISA
@Cal3Tec: DiagnosticsA
7#.6
@6D."3
77.6A
D$."
@D1.33
D7.DA
& S
%enela9ia )i!t
@Sano6i Diagnostics
Paste5rA
100.0
@DE.63
100.0A
DE.$
@D$.63
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158
$4. LIST OF REFERENCES FOR FURTHER STUDY
0$ 2ID' : 2etiology, Diagnosis and Treat#ents Eth ;d$ ;ditors Vinet T$ De"itta 9r$,
'a#uel Hell#an and 'te"en 2 6osenberg$ 2ssoiate ;ditors 9a#es Curran,
Ma/ ;sse/ and 2nthony '$ -aui$ Li%%inot A 6a"en, 8$L$ 0BBH$
=$ HIVAVolu#e 0 : 2 )ratial 2%%roahJ Virology and I##unology$
;dited by 9$ :arn$ (/ford <ni"ersity )ress$ (/ford 0BBG$
@$ HIVAVolu#e = : 2 )ratial 2%%roahJ Biology, Moleular Biology and Drug
Diso"ery$ ;dited by 9$ :arn (/ford <ni"ersity )ress, 0BBG$
E$ I##unology of HIV Infetion$ ;dited by 'udhir 3u%ta$ )lenu# )ublishing
Cor%oration, 8e, Lor. 0BB?$
G$ 2ID'!HIV : 6eferene 3uide for Medial )rofessionals$ ;ditors 9ohn L$ -ahey
and Diana 'hin -le##ing$ 5illia#s and 5il.ins$ 2 ,a"erly o#%any,
Balti#ore, 0BB?$
?$ 3uidelines for )re"enting HIV,HBV and other Infetions in the Health Care
'etting$ Dou#ent 8o$ I'B8 BHEAGG1AHMAG$ 0BB?$
H$ (%erational Charateristis of Co##erially a"ailable 2ssays to Deter#ine
2ntibodies to HIVA0 and ! or HIVA= in Hu#an 'era$
Dou#ent 8o$ 5H(!BL'!B1$0 <82ID'!B1$0E$
1$ 6e"ised 6eo##endations for the 'eletion and use of HIV antibody tests$
5H(!<82ID', 0BB1$
B$ 8ational HIV Testing )oliy$ 82C( Dou#ent, 0BBG$
0>$ 6etro"iral Testing : ;ssentials -or Quality Control and Laboratory Diagnosis$
;ditors 8eil T$ Constantine$ 9ohnny D$ Callahan and Donglas M 5att$ C6C
)ress$ 2nn 2rbor, 0BB=$
00$ Hu#an I##unodefiieny Virus : Laboratory Training Manual$
2'T)HLD, Connetiut 2"e$ 5ashington D$C$ 0BBE$
0=$ )ubli health ser"ie guidelines for #anage#ent of health are ,or.er e/%osure
to HIV and reo##endations for %ost e/%osure %ro%hyla/is MM56, Vol$ EH,
8(66AH, May 0G, 0BB1$
0@$ :artD MH and 3erberding 9L$ )ost e/%osure treat#ent of %eo%le e/%osed to the
HIV through se/ual ontat or infetion during se/$ 8;9M$ @@? &0G*: 0>BHA
BB,0BBH$
0E$ 8ational HIV testing net,or. o%erational guidelines 82C( Dou#ent$
159
CHA%TER .I
%ROFORMAS TO BE USED
FOR SENDING RE%ORTS TO
DI('CTO(1 STAT' AIDS SOCI'T* AND
ADDITIONA, DI('CTO( T'C+NICA,1
NATIONA, AIDS CONT(O, O(%ANISATION
160
Montl+ la=orator+ re5ort on !erolo#ical te!tin#
0ol6ntar+ =lood te!tin#8!6rveillance centre!
Na8e o6 t:e Testing Centre;,a7oratory
ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
City ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ State
ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
Date and year o6 re4ortZZZZZZZZZZZZZZZZZ (e4ort 6or t:e 8ont:
o6;yr.ZZZZZZZZZZZZZZ
N587er o6 4ersons tested
Po45lation;
Area
2y one test 2y s544le8ental tests
Tested Positi9e Percent Tested Positi9e Percent
STD clinic attendants
)ales
e8ales
Co88ercial se! <or=ers
)ales
e8ales
+o8ose!5als
Injecting dr5g 5sers
)ales
e8ales
)o7ile 8en1 s4eci6y
Antenatal1 clinic
attendants
2lood donors1 all
&ol5ntary
(e4lace8ent
S5s4ect aids;arc cases
Ot:er gro54s1 s4eci6y :
Not s4eci6ied1
Total1 d5ring t:is 8ont:
Total1 since ince4tion
Total1 d5ring
or re6erence la7. only
No. o6 con6ir8atory;
s544le8entary tests
No. o6 tests 6or B5ality
control
To 7e sent to Addl. Director Tec:nical1 NACO1 %.O.I. d5ring t:e #nd <=. o6 e9ery
8ont:.
161
Montl+ la=orator+ re5ort on !erolo#ical te!tin#
*eference Centre
Na8e o6 t:e testing centre
ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
City ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ State
ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
Date o6 re4ort <it: yr. ZZZZZZZZZZZZZZZZZZZZZZ(e4ort 6or t:e 8ont:
o6;yr.ZZZZZZZZZ
A#e8!eH di!tri=6tion of HI0 te!ted 5er!on!
Tested 7y one
screening test
Tested 7y s544le8ental tests
Age gro54 2y one test e8ales 2ot: se!es
Tested Positi9e Tested Positi9
e
Tested Positi9e
0310
113#0
#1330
313"0
"1 and a7o9e
All ages
Te!t 3it! !655l+
Test =it ty4e and
n587er o6 tests 4er
=it
(ecei9ed t:is
year
Csed 7y re4ort
date
In
stoc=
(e4lace8ent
reB5ired
Co88ents1 4ro7le8s
ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
ZZZZZ
(e4orting O66icer ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
@Na8e - DesignationA
Date: ZZZZZZZZZZZZZZZZZZZZZZZ Signat5re :
ZZZZZZZZZZZZZZZZZZZZ
To 7e sent to Addl. Director1 Tec:nical1 NACO1 )+ - .1 Nir8an 2:a9an1 Ne< Del:i3
110011 d5ring t:e 6irst <eee= o6 e9ery 8ont:.
162
Montl+ !tate re5ort on HI0 !erolo#ical te!tin#
0ol6ntar+ =lood te!tin#8!6rveillance
State ZZZZZZZZZZZZZZZZZZZZZZ Na8e o6 t:e Testing Centre;,a7oratories
ZZZZZZZZZZZ
Date and re4ort o6 year ZZZZZZZZZZZZZZZZZZZZZ (e4ort 6or t:e 8ont: o6;yr.
ZZZZZZZZZZZZZ
N587er o6 4ersons tested
Po45lation;
Area
2y one test 2y s544le8ental tests
Tested Positi9e Percent
Positi9e
Tested Positi9e Percent
Positi9e
STD clinic attendants
)ales
e8ales
Co88ercial se!
<or=ers
)ales
e8ales
+o8ose!5als
Injecting dr5g 5sers
)ales
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)o7ile 8en1 s4eci6y
Antenatal1 clinic
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2lood donors1 all
&ol5ntary
(e4lace8ent
S5s4ect aids;arc
cases
Ot:er gro54s1 s4eci6y :
Not s4eci6ied1
Total1 d5ring t:is
8ont:
Total1 since ince4tion
Total1 d5ring t:e year
or re6erence centres
No. o6 con6ir8atory;
s544le8entary tests
No. o6 tests 6or B5ality
control
To 7e sent to State AIDS Progra88e O66icer 7y eac: testing 6acility d5ring t:e 6irst <=. o6
e9ery 8ont:.
163
Montl+ donor! !creenin# re5ort of =lood =an38tran!f6!ion centre
2lood 7an=;trans65sion centre
ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
City ZZZZZZZZZZZZZZZZZZZ State ZZZZZZZZZZZZZZZZZZZZ Date o6 re4ort;yr.
ZZZZZZZZ
(e4ort 6or t:e 8ont: o6 ZZZZZZZZZZZZZZZZ year ZZZZZZZ
Na8e o6 t:e centre;
No. o6 sa84les 6ro8 donors
No. o6 tests 4er6or8ed; &ol5ntary (e4lace8ent Total
No. o6 4ositi9e
) T ) T ) T
G2TC
No. Tested
No. Positi9e
2lood 7an=s attac:ed to G2TC
1.
No. Tested
No. Positi9e
#.
No. Tested
No. Positi9e
3.
No. Tested
No. Positi9e
".
Tested
Positi9e
$.
Tested
Positi9e
6.
Tested
Positi9e
Total 6or t:e re4orting
8ont:
Total since ince4tion
Total d5ring t:e c5rrent
year
TOTAL DU*ING THE 1*E0IOUS :EA* 2 )))))))))))))))))))))))) 4
164
Montl+ donor! !creenin# re5ort of =lood =an38tran!f6!ion centre
2lood 7an=;trans65sion centre
ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
City ZZZZZZZZZZZZZZZZZZZ State ZZZZZZZZZZZZZZZZZZZ Date and year o6 re4ort
ZZZZZ
(e4ort 6or t:e 8ont: o6 ZZZZZZZZZZZ *ear ZZZZZZZ
+I& testing 4rotocol: One screening test @';(;SA ZZZZZZZZZ t<o or 8ore
@';(;SAZZZZZZ
T:ree @';(;SA ZZZZZZZZZZZZZZZZZZZZZZZZZZ @';(;SA and .estern 2lot tests
No. o6 sa84les 6ro8 donors
Tested and res5lts &ol5ntary (e4lace8ent Total
) T ) T ) T
+2s Ag: Tested
Positi9
e
&D(, : Tested
Positi9
e
)A,A(IA : Tested
Positi9
e
Te!t! 5erformed d6rin# te calendar +ear
Tests
&ol5ntary ; (e4lace8ent
) T
+2s Ag:
&D(,
)alaria
HI0 te!t 3it! !655l+
Test =it ty4e
And no. o6 (ecei9ed Csed 7y In stoc= (e4lace8nt
Tests 4er =it t:is year re4orting
ti8e
reB5ired
Pro7le8s1 co88ents ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
(e4orting O66icer ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
@Na8e - DesignationA
Date : ZZZZZZZZZZZZZZZZ Signat5re ZZZZZZZZZZZZZ
165
Montl+ donor! !creenin# re5ort of =lood =an38tran!f6!ion centre
State ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ
(e4ort 6or t:e 8ont: o6 ZZZZZZZZZZZZZ year ZZZZZZZZZZZZ
Na8e o6 t:e centre; No. o6 sa84les 6ro8 donors
No. o6 tests 4er6or8ed; &ol5ntary (e4lace8e
nt
Pro6essional Total
No. o6 4ositi9e
) T ) T ) T ) T
G2TC
Tested
Positi9e
2lood 7an=s attac:ed to J7tc
1.
Tested
Positi9e
#.
Tested
Positi9e
3.
Tested
Positi9e
".
Tested
Positi9e
$.
Tested
Positi9e
6.
Tested
Positi9e
Total 6or re4orting
8ont:
Total since ince4tion
Total t:e year
TOTAL DURING
L-s( )3 3-2'+r"s '* ''"A+r"s
(62"s ')( '+1/"r"*)
Ch6("r ; $ %2" N).
166
-ig$ 0$0 Cu##ulati"e HIV Infetions in India
&May 0B1? A Marh 0BBB*$
0? A a
-ig$ 0$= Cu##ulati"e 2ID' Cases in India
&May 0B1? A Marh 0BBB*$
0? A b
-ig$ 0$@ HIV Infetion in India A
Cu#ulati"e 'ero%ositi"ity rate &0BB=A'e%te#ber 0BB1*$
0? A
-ig$ 0$E 2ge and se/ distribution of 2ID' Cases in India$ 0? A d
-ig$ 0$G )robable soure of infetion of 2ID' ases in India
&nR??>H*$
0? A e
-ig$ 0$? 8ational 2ID' Control )rogra##e$ 0? A f
Ch6("r ; .
-ig$ =$0 6e%resentation of the struture of the hu#an
i##unodefiieny "irus$
=> A a
-ig$ =$= 'truture and funtions of HIV A 0 geno#e$ => A a
Ch6("r ; =
-ig$ @$= :inetis of CDE ly#%hoytes during the ourse of HIV
infetion$
=B A a
-ig$ @$@ Viral load and ourse of infetion$ =B A b
Ch6("r ; >
-ig$ E$0 2reas o##only #issed in hand ,ashing$ E1 A a
-ig$ E$= 'te%s of effeti"e hand,ashing$ E1 A b
Ch6("r ; ?
-ig$ G$0 )a.ing infetious substanes for the %ost$ G> A a
-ig$ G$= Infetious substane label$ G> A b
Ch6("r ; F
-ig$ ?$0 HIV #ar.ers fro# ti#e of infetion to endstage disease$ G= A a
-ig$ ?$= )rini%le of the indiret ;LI'2$ G= A a
-ig$ ?$@ )rini%le of the o#%etiti"e ;LI'2$ GH A a
-ig$ ?$E 2n antigen sand,ih ;LI'2 for deteting HIV antibody$ GH A a
-ig$ ?$G )rini%le of the indiret antigen a%ture ;LI'2$ G1 A a
-ig$ ?$? The Ig3 antibody a%ture ;LI'2$ G1 A a
-ig$ ?$H 2gglutination assay %ositi"e$ ?> A a
-ig$ ?$0> 'he#ati re%resentation of 5B results$ ?? A a
2nne/ure 0 'he#ati re%resentation of the <82ID' and 5H( HIV
testing strategies$
?1 A a
167
2nne/ure @ )rofor#a &infor#ation* for re+uisitioning HIV testing$ ?B A a
Ch6("r ; 5
-ig$ 1$0 )rini%le of )C6 %roedure$ H1 A a
-ig$ 1$= 3el eletro%horesis of ;8V )C6 %roduts$ 1> A a
Ch6("r ;
$.
-ig$ 0=$0 Quantitati"e HIV 682 )C6 for "iral load #easure#ent$ 00H A a
-ig$ 0=$= Branhed D82 assay$ 00H A b
-ig$ 0=$@ 82'B2$ 001 A a
Ch6("r ;
$=
-ig$ 0@$0 'truture of HBV$ 0=1 A a
-ig$ 0@$= 'erologial %rofile of aute HBV infetion$ 0=1 A a
-ig$ 0@$@ 'erologial %rofile of HBV infetion$ 0@> A a
-ig$ 0@$E )rini%le of ;LI'2 for HBs2g detetion$ 0@> A a
Ch6("r ;
$G
2nne/ure A0 0G? A a
Fi#) <)< 0iral load and co6r!e of infection
Sero conver!ion
10
7
AIDS
10
6
ra4id
4rogression
10
$
ty4ical
4rogression
10
"
long3ter8
4rogression
10
3
Clinical latenc+
168
-ee3! +ear!
Time
169