ABO Blood Group

System
History: Karl Landsteiner
Discovered the ABO Blood Group
System in 1901
He and his five co-workers began
mixing each others red cells and serum
together and inadvertently performed
the first forward and reverse ABO
groupings
Why is it important?
ABO compatibility between donor cell and
patient serum is the essential foundation of
pretransfusion testing
It is the only system with expected antibodies
Whether they are IgG or IgM, ABO antibodies
can activate complement readily
This means that incompatibilities can cause life
threatening situations (transfusion reactions)
ABO antigens:
Biochemical & Genetic Considerations
ABO and H Antigen Genetics
Genes at three separate loci control the
occurrence and location of ABO antigens

The presence or absence of the A, B, and H
antigens is controlled by the H and ABO
genes
The presence or absence of the ABH
antigens on the red blood cell membrane is
controlled by the H gene
The presence or absence of the ABH
antigens in secretions is indirectly controlled
by the Se gene
ABO Antigen Genetics
H gene H and h alleles (h is an amorph)

Se gene Se and se alleles (se is an
amorph)

ABO genes A, B and O alleles

H Antigen
The H gene codes for an enzyme that adds
the sugar fucose to the terminal sugar of a
precursor substance (PS)
The precursor substance (proteins and lipids)
is formed on an oligosaccharide chain (the
basic structure)

RBC Precursor Structure
Glucose
Galactose
N-acetylglucosamine
Galactose
Precursor
Substance
(stays the
same)
RBC
Formation of the H antigen
Glucose
Galactose
N-acetylglucosamine
Galactose
H antigen
RBC
Fucose
H antigen
The H antigen is the foundation upon which A
and B antigens are built
A and B genes code for enzymes that add a
sugar to the H antigen
Immunodominant sugars are present at the
terminal ends of the chains and confer the ABO
antigen specificity
A and B Antigen
The A gene codes for an enzyme (transferase)
that adds N-acetylgalactosamine to the
terminal sugar of the H antigen
N-acetylgalactosaminyltransferase

The B gene codes for an enzyme that adds
D-galactose to the terminal sugar of the H
antigen
D-galactosyltransferase
Formation of the A antigen
Glucose
Galactose
N-acetylglucosamine
Galactose
RBC
Fucose
N-acetylgalactosamine
Formation of the B antigen
Glucose
Galactose
N-acetylglucosamine
Galactose
RBC
Fucose
Galactose
Genetics
The H antigen is found on the RBC when
you have the Hh or HH genotype, but NOT
from the hh genotype
The A antigen is found on the RBC when
you have the Hh, HH, and A/A, A/O, or A/B
genotypes
The B antigen is found on the RBC when
you have the Hh, HH, and B/B, B/O, or A/B
genotypes
H antigen
Certain blood types possess more H antigen
than others:


O>A
2
>B>A
2
B>A
1
>A
1
B
Greatest
amount of H
Least
amount of H
The O allele
Why do Group O individuals have more
H antigen than the other groups?
The O gene is a silent allele. It does not
alter the structure of the H
substance.that means more H antigen
sites


Group O Group A
Many H
antigen sites
Fewer
H antigen
sites
A
A
A
A
A
Most of the H antigen sites in a
Group A individual have been
converted to the A antigen
ABO Antigens in Secretions
Secretions include body fluids like plasma,
saliva, synovial fluid, etc

Blood Group Substances are soluble
antigens (A, B, and H) that can be found in
the secretions.
This is controlled by the H and Se genes
Secretor Status
The secretor gene consists of 2 alleles (Se
and se)
The Se gene is responsible for the
expression of the H antigen on glycoprotein
structures located in body secretions
If the Se allele is inherited as SeSe or Sese,
the person is called a secretor
80% of the population are secretors
Secretors
Secretors express soluble forms of the H
antigen in secretions that can then be
converted to A or B antigens (by the
transferases)
Individuals who inherit the sese gene are
called nonsecretors
The se allele is an amorph (nothing expressed)
sese individuals do not convert antigen precursors
to H antigen and has neither soluble H antigen nor
soluble A or B antigens in body fluids

Secretor Status Summary
The Se gene codes for the presence of the H
antigen in secretions, therefore the presence
of A and/or B antigens in the secretions is
contingent on the inheritance of the Se gene
and the H gene

Se gene
(SeSe or Sese)
H antigen in
secretions
A antigen
B antigen
se gene
(sese)
No antigens secreted
in saliva or other
body fluids
and/or
ABO Group
ABH
Substances
Secretors (SeSe or Sese): A B H
A +++ 0 +
B 0 +++ +
O 0 0 +++
AB +++ +++ +
Non-secretors (sese):
A, B, O, and AB 0 0 0
Sese + h/h (no H antigen) no antigens in secretions
Type I and Type II Precursors
There are two potential precursors substances for
ABH antigens Type I and Type II
Both are comprised of identical sugars but the
linkage of the terminal sugars differs in the two types
Type I precursor has a terminal galactose linked to a
subterminal N-acetylgluosamine in a 1-3 linkage
These same sugars combine in a 1-4 linkage in type
II precursor
ABH Ags on red cells are derived from Type II
chains whereas the ABH Ags in plasma are made
from both types I & II precursors
Type II H
After fucose is added to Type II chains, the
structure is termed Type II H
Four kinds of Type II H have been identified
H1, H2 are simple straight chain glycolipids
Whereas H3 & H4 have branched chains
ABO Subgroups
ABO subgroups differ in the amount of antigen
present on the red blood cell membrane
Subgroups have less antigen
Subgroups are the result of less effective
enzymes.
They are not as efficient in converting H
antigens to A or B antigens (fewer antigens are
present on the RBC)
Subgroups of A are more common than
subgroups of B
Subgroups of A
The 2 principle subgroups of A are: A
1
and A
2

Both react strongly with reagent anti-A
To distinguish A
1
from A
2
red cells, the lectin
Dolichos biflorus is used (anti-A
1
)
80% of group A or AB individuals are subgroup A
1
20% are A
2
and A
2
B
A
2
Phenotype
Why is the A
2
phenotype important?
A
2
and A
2
B individuals may produce an anti-A
1
This may cause discrepancies when a crossmatch is done
(incompatibility)
Whats the difference between the A
1
and A
2

antigen?
Its quantitative
The A
2
gene doesnt convert the H
3
& H
4
to A very well
The result is fewer A
2
antigen sites compared to the many
A
1
antigen sites
A
1
and A
2
Subgroups
Anti-A
antisera
Anti-A
1

antisera
Anti-H
lectin
ABO
antibodies
in serum
# of
antigen
sites per
RBC
A
1
4+ 4+ 0 Anti-B 900 x10
3

A
2
4+ 0 3+ Anti-B &
anti-A
1

250 x10
3


Other A subgroups
There are other additional subgroups of A
A
int
(intermediate), A
3
, A
x
, A
m
, A
end
, A
el
, A
bantu

A
3
red cells cause mixed field agglutination
when polyclonal anti-A or anti-A,B is used
Mixed field agglutination appears as small
agglutinates with a background of
unagglutinated RBCs
They may contain anti-A
1
B Subgroups
B subgroups occur less than A subgroups
B subgroups are differentiated by the type of
reaction with anti-B, anti-A,B, and anti-H
B
3
, B
x
, B
m
, and B
el

Other ABO conditions
Bombay Phenotype (O
h
)
Inheritance of hh
The h gene is an amorph and results in
little or no production of L-
fucosyltransferase
Originally found in Bombay (now Mumbai)
Very rare

Bombay
The hh causes NO H antigen to be produced
Results in RBCs with no H, A, or B antigen
(patient types as O)
Bombay RBCs are NOT agglutinated with
anti-A, anti-B, or anti-H (no antigens present)
Bombay serum has strong anti-A, anti-B and
anti-H, agglutinating ALL ABO blood groups
What blood ABO blood group would you use
to transfuse this patient??


ANSWER:
Another Bombay
Group O RBCs cannot be given because they still
have the H antigen
You have to transfuse the patient with blood that
contains NO H antigen

ABO Blood Group
ABO Antibodies
Landsteiners Rule:
Normal, Healthy
individuals possess
ABO antibodies to
the ABO antigen
absent from their
RBCs


ABO Blood Group System
The ABO Blood Group System was the first
to be identified and is the most significant for
transfusion practice
It is the ONLY system that the reciprocal
antibodies are consistently and predictably
present in the sera of people who have had
no exposure to human red cells
Blood Group Systems
Most blood group systems (ABO and others)
are made up of:
An antigen on a red cell and the absence of its
corresponding antibody in the serum (if youre A,
you dont have anti-A)
If you do NOT have a particular antigen on
your red cells then it is possible (when
exposed to foreign RBCs) to illicit an immune
response that results in the production of the
antibody specific for the missing antigen
ABO
Remember:
The ABO Blood Group System does NOT require
the presence of a foreign red blood cell for the
production of ABO antibodies
ABO antibodies are non-red blood cell
stimulated probably from environmental exposure
and are referred to as expected antibodies
Titer of ABO Abs is often reduced in elderly and in
patients with hypogammaglobulinemia
Infants do not produce Abs until 3-6 months of
age
ABO antibodies
RBC
Phenotype
Frequency
(%)
Serum Ab
A 43 Anti-B
B 9 Anti-A
AB 4 --------
O 44 Anti-A,B
Anti-A
1
Group O and B individuals
contain anti-A in their serum
However, the anti-A can be
separated into different
components: anti-A and
anti-A
1

Anti-A
1
only agglutinates the
A
1
antigen, not the A
2

antigen
There is no anti-A
2
.
Anti-A
1
Clinically
Significant
Sometimes
Abs class
IgM
Thermal
range
4 - 22
HDNB
No
Transfusion Reactions
Extravascular Intravascular

No Rare
Anti-A,B
Found in the serum of group O individuals
Reacts with A, B, and AB cells
Predominately IgG, with small portions being
IgM
Anti-A,B is one antibody, it is not a mixture of
anti-A and anti-B antibodies
ABO antibodies
IgM is the predominant antibody in Group A
and Group B individuals
Anti-A
Anti-B
IgG (with some IgM) is the predominant
antibody in Group O individuals
Anti-A,B (with some anti-A and anti-B)
ABO antibody facts
Complement can be activated with ABO antibodies
(mostly IgM, some IgG)
High titer: react strongly (4+)

Anti-A, Anti-B, Anti-A,B
Clinically Significant
Yes
Abs class
IgM, less IgG
Thermal range
4 - 37
HDNB
Yes
Transfusion Reactions
Extravascular Intravascular
Yes Yes
ABO Antibodies
Usually present within the first 3-6 months of
life
Stable by ages 5-6 years
Decline in older age & in
hypogammaglobulinemia
Newborns may passively acquire maternal
antibodies (IgG crosses placenta)
Nature of antibodies
Non-red blood cell stimulated
ABO antibodies
Red blood cell stimulated
Antibodies formed as a result of transfusion, etc
Usually IgG
Active at 37C
Can occur in group O (may occur in group A or B)
These antibodies also occur in the other Blood Group
Systems
Anti-H
Auto-Anti-H
Clinically
Significant
No
Abs class
IgM
Thermal range
4 - 15
HDNB
No
Transfusion Reactions
Extravascular Intravascular
No No
Allo-Anti-H
Clinically
Significant
Yes
Abs class
IgM, IgG
Thermal range
4 - 37
HDNB
Yes
Transfusion Reactions
Extravascular Intravascular
Yes Yes
RH System
M. Zaharna Blood Bank 2009
The Rh(D) Antigen
Rh is the most complex system, with over
45 antigens
The complexity of the Rh blood group
Ags is due to the highly polymorphic
genes that encode them.
Discovered in 1940 after work on Rhesus
monkeys
The 2
nd
most important after ABO in the
crossmatch test
Only the most clinically significant Ags
will be discussed
Rh Genetics
The genes that control the system are
autosomal codominant located on the short
arm of chromosome 1.
M. Zaharna Blood Bank 2009
Rh blood group antigens are proteins
The antigens of the Rh blood group are proteins.
The RhD gene encodes the D antigen, which is
a large protein on the red blood cell membrane,
& the most important.
RHD gene RHCE gene
Chromosome 1
Proteins
M. Zaharna Blood Bank 2009
Rh Antigen Frequency
D antigen 85%
d antigen 15%
C antigen 70%
c antigen 80%
E antigen 30%
e antigen 98%

The presence or absence of D Ag determines if
the person is Rh+ or Rh-
Rh Positive
Rh Negative
M. Zaharna Blood Bank 2009
Weak D Phenotype
Most D positive rbcs react macroscopically with
Reagent anti-D at immediate spin
These patients are referred to as Rh positive
Reacting from 1+ to 3+ or greater

HOWEVER, some D-positive rbcs DO NOT
react (do NOT agglutinate) at Immediate Spin
using Reagent Anti-D.
These require further testing (37
o
C and/or AHG)
to determine the D status of the patient.
M. Zaharna Blood Bank 2009
Rh Deleted
Red cells that express no Ags at the C & E
loci ( D )
Number of D Ags greatly increase
Anti-D IgG Abs can agglutinate these cells



M. Zaharna Blood Bank 2009
RH null: individual that appears to have no Rh antigens
( , , )
RBC has fragile membrane- short lived
Must use autologous blood products
No D, C, c, E, e antigens present on the RBC membrane
Demonstrate mild hemolytic anemia (Rh antigens are
integral part of RBC membrane and absence results in
loss of membrane integrity)
Stomatocytosis.
When transfusion is necessary ONLY Rh Null blood can
be used to transfuse.
Rh null
M. Zaharna Blood Bank 2009
Rh antibodies
Result from the
exposure to Rh
antigens
IgG form
Bind at 37C
Form agglutination in
IAT phase
Rh Abs

Clinically
Significant
Yes
Abs class
IgG
Thermal
range
4 - 37
HDNB
Yes
Transfusion Reactions
Extravascular Intravascular

Yes No
M. Zaharna Blood Bank 2009
Related to Hemolytic transfusion reactions
Re-exposure to antigen cause rapid
secondary response
Always check patients history for previous
transfusion or pregnancy to avoid re-
exposure.
Clinical Significance of Rh antibodies
M. Zaharna Blood Bank 2009
Usually related to D antigen exposure and the
formation of anti-D
Usually results from D negative female and D
positive male producing and offspring.
The baby will probably be D positive.
1
st
pregnancy not effected, the 2
nd
pregnancy
and on will be effected-results in still birth,
severe jaundice, anemia related to HDN.
To prevent this occurrence the female is
administered RHIG.
Hemolytic disease of the Newborn
(HDN)
M. Zaharna Blood Bank 2009
Rh factor can
cause
complications in
some
pregnancies.
Mother is exposed
to Rh antigens at
the birth of her Rh
+

baby.
First pregnancy
Placenta
Rh
+
antigens
Rh factor
M. Zaharna Blood Bank 2009
Anti-Rh
+

antibodies
Possible subsequent
pregnancies
Mother makes anti-
Rh
+
antibodies.
During the mothers
next pregnancy, Rh
antibodies can cross
the placenta and
endanger the fetus.
M. Zaharna Blood Bank 2009
Significance
After ABO, the Rh system is the second most important
system. This is because:
The D antigen is extremely immunogenic.
It causes the production of anti-D in 50 - 70% of Rh(D)
negative people who are exposed to the D antigen.
Moreover, anti-D is the most common cause of severe
HDN and can cause in Utero death.
Because of this, in blood transfusion, the patient and
donor are matched for Rh(D) type as well as ABO
groups.
The C and E Ags are not as immunogenic as D, routine
typing for these Ags is not performed