BIOMEDICAL APPLICATIONS OF LASERS

P.K. Gupta
Biomedical Applications Section, Indore, Centre for Advanced Technology 452 013, India

Ab,"ract

The

ose of

la,ser,s

jar or

medical exogenous

diagnosLe,

and jar

therapy i,s

initialed

by photo-excitalian con,sidaable current

<if nalUral altenlioo care, been

chromophures Development' Therejare, to explore the

photosensilisen,

ceceiring

in these an aclivity

arca,e are expeclcd in Ihe", areas

to hare profoand al CAT

ilifluence in 1992-93, jar

on Ihe quality The initial

of health have

wo,s' ,"arled

efforts ond

the use nfla",r effect, can'ied of

indocedj/oarescence of laser imldiatioll

,spectroscopy un lis'ing

cancer

diagnosis, The new article

ineesligate an up at of

photo-hiological of the work devclopme'" obfects

orgollisms, Some and ~flhe loser

prorides taken

oeereiew CAT on

oot at CAT Icchniqoes

in both the,ve areas, for oplical imaging

aclivities

micromanipulation

micrn,snlpic

are aleu briefly

dLecns,sed

Introduction DUE TO THEIR arc SEVERAL finding The

in-situ,

near real time diagnosis radiation suited fm which mass

and the use of makes screening effcm, is the use of applications's of naturally A where developing them and

non-ionizing parTicularly

peoperties applicatiom ym of the invention ulrn-peecise

r'epeated use withour any adv",e Another area of growing interest

in medicine' is now with

use of lams in surgery, which started within a of law, surgery well e"ablished, invasive Lams have made pos,siblc minimally reduced time. The use is also arrracting signifIcant This wurk is by can and rhe

Ia>ers for non-surgical occurring good

rherapeuric activation the of fast

It exploits light induced or administered example is therapy

photosemitiser" cancer'"

patient rnuma and hospirali,",ion of lasers for medical diagnosis considerable advancements motivated imcre>[ ace being made'" and

photodynamic

photo"citation of a rumor' localized drug leads to selective demuction of tumor with mioimal effect on healthy that tissue, There are also of

by the fact that the onset changes,

peogression of a disease are uften accompanied biochemicallmmphological be sensitively techniques, moniroted which

indications therapeuric litetature"

selective

photuexcirarion

native chromophorc

in rhe tissue may lead to

by laser spectroscopic can therefore lead

These rcchniqnes difficult

effem, Clinical repons available in as well as mults from Choithram (CHRCt"', with lasers or

to disease diagnosis disease becomes important spectroscopic

at an early stage befme the to manage, The other offered by laser fur are their poremial

Hospital and Reswch Centct Indore have shown that inadiariun other narrow bandwidrh several thcrapeuric

advantages techniques

light sources can lead to

effects like accelerated wound

I 13 I--

healing, "eatmem tcearmem mechanisms of paio

of pulmona'}' of various

nrheulmis etiologies, pmemial

and The of

induced

fluorcscence

spwroscopy

for

canca

diagnosis. The r<-sults of the Sltldics carried out to investig"te i,mdiation n<xl. This on photo-hiologi,,1 living ""ganisms effects of laser '" pr"sented of

for these thetapeuric The dinid

effeCtS arc nOt thmpeutic work in this is being

vc'}' well undmtood. this cather simple diteetion made", Realizing diagnosis and modality is motivating

and inexpeosive considerable pmgcess

is followed

by a brief discussion

some of tbe new acriviries taken up at CAT.

sobstamial

Laser
the profoond influence can opri,,1 the and photntherapy have on

Induced

Fluorescence

(LIF)

Spectroscopy

for Diagnosis of Cancer

cancer in tWO sYStemic by

qoali'}' of health eare, work in these arca> w" initiared ar CAT in 1992-93. Several Stodies have been made effem and to to investigate the me the photO-biologic-,1 on liviog ocgaoisn1' of laver induced diagnosis, at surgery fot cancer reswed

!.IF has heen us"d for diagnosing ways, One approach administration rhe rumoL

involves

of" dntg like haemaotOporpby,in which is selectively mained photoexcited with When light of

of Iasec icradiation explore speCtroscopy

deriv,nive (HpD)

tlooreseence

Following stodies on sampb or binpsy from patietm arga",.

appropliare wavelength, tumor tloores<:es. This detection triplet swe and via

the dntg localized in the fluores<:cn<:e is used fot of the cr",sing. tumoL thc The

wirh cancer of difTeretlt

imaging a"o leads intersystem

systems suirable foc clinical use have been

Pboroex"irotion

ro populating

develnped at CAT. One unit has been i",ralled at a cancer screeuing center for screening patients with neopl"m h" patients wirh of urerine eervix. The other unit histopatbologically confirmed mults been used to Cat'}' OUI a pilot study on 25

molecule in exdted rripla with biomolecules oxygen, which resulting exploited hom approach

Stare can dire"tly teact The is

01 lead ro generation of singlet of the host tisslle

is roxie tu the hoSt ti"ue. thmpy

de","etioo

canw of oral eaviry and eocouraging obrained. Work has also been initiated oth" through withom promising areas, One

fa, phorody,,"mic has cwo drawback"

of tumoL dack

the poiot of view of nse in diagnosis this a po"ible

,It CAT in some of these is the in"ging imaging and

tOxicity of the drug and th" possibility of drug induced photOsensitization. ,hawback induced necessitating week>. Thae tumo,' mackm quenched avoided. and photosmsitizarion the patient is therefore whm thcceby For example a major of the skin developmem ro pmvide of teehni,]ues sob-millimeter for optical resolotion with the lIse of HpD has been dntgto avoid light for a few intereSt in devdoping phorosensitizarion is

turbid media. Thesc have rhe porential rhe need ofbser for ionizing radiation

associated risks". The orher aetivi'}' taken op is devdopment trap and mierobeam vcr up, These set ops "'c finding widesp<ead applie",ions in biomedicine for non-comaet mieromanipolation just being 'esearch'". of microscopic tool objem and

the tr'iplet State is mpldly the one that h" InStead it

The Other approach,

received mote attention exploitS fOt diagnosis

in recent years, does not markers. the subtle ehangcs in thc

use any <'Xogenou.s tumor

are expecred ro have an impact far beyond than a new physic!1 for biological

'peen-urn as well as the decay time of fluoI",eenee from native tissuc> as it rransfolms from normal to the malignant cartied state. The studies

In this article we first provide an owvi"w of the work cartied out at CAT on the use of laser

Out over the laSt few years have shown

14

I--

considenble diagnosis m"ioc covity" cmix,

pwmi;e of .hc canee. csoplugus,

af

this

appmach mgans

fa. like

beam

splinre-phorodiode

combioation, by thc "me slit uf

Thc

of va.ious

fluoecscence fcom .he tissue, kept io contact with the fibre, is collected imaged 00 the fibre and a scanning emeance

lung, b,'east, and onl

In-virm <t"die; at CA T A schen",ic of the expccimental used at CAT for in-vino aotofluorescence I"" tI", spemoscupy mangcmcm m,dies on

monochmmatoL The wavelengtb-dispmcd light " tbc exit slit of the monochmmatm is detected by a phatomultiplire spem tube detector. A

micmpmceo.-bascd powe. and

developed at CAT of Nt lase. data, Pm specteal

is being ",cd fm on-line fluoeesccncc excitation/emission wavelengths (SPEX, Flumolog

acquisition

of human tiucs is pulsed Nt minm and by a

shown in fig, 1. It mes a home-buih fibet (core diamcte. ,'cflew Nt 400~m)

spectwscopic

studics at othee

the output of which is coupled to an opti,,1 via dichmic (3370m) lase. rodi"ion

a commercial

spcwofluarometet

II) w" used, In the following, the eesuhs of the "udies on autofluoeescence fmm

transmits Inogcc wavckogth

fluoecsccncc ompur.

we bticfly summ"i" cmied out at CAT

The powet of the lasre pube is manitated

tissues fmm diffeccnt otgans,

Fig 1

Sth""","

diag",m

"fthe expaimem,,!

rd-up ftr au"1Iu"""""

,p"""""py

"ftisru",

IS I--

PATIENT
Fig.2. &a""pl"'fl"h, in"g.,."djlua",nin"miryfl'N, /&"""i"d 'po'" afpai"dcan"'maud",",,1 a,,1Un""i", ftam 12.parim""kmd","udam.P"im' numb"1" 4badcau",afal",lm (A); " 8hadca"",afbuoalm"a" (BM)aud9 5 " 12badcan",af"ng.u,(T).
Cancer of oral cavil] Oral cane is one of the most common cancers and 4 tongue) selected The I,(A) mean at random &om the higher

patienrs in India and several other Sourh Asian counrries. Laser induced fluorescence technique is

investigated.

considerably

values of L. apparent. normal compared The

for normal value

tissue sites is I,(A) from

of L.

particularly

wdl suited for early detection of oral

tissue sites was larger by a factor of 2 to that &om cancerous tissue sites. At wavelengths significant (A. : 300 nm difference in

cancers due to the easy accessibility of this organ. Studies on 337 nm excited autofluorescence spectra from oral tissues (alveolus. buccal mucosa and the tongue significant integrated tissue samples in the obtained total. from patients with the cancer of oral cavity) revealed differences specrrally

the other excitation and 460 nm). no

fluorescence yidd betWeen cancerous and normal oral tissue sites was observed. With 337 nm excitation discrimination discrimination use ofLI,(A) alone as a

fluorescence intensity <L. I,(A) ;360 ,; and normal intensity normal oral for tissue

As 600 nm) &om cancerous spectrally paired integrated and

parameter provided exedlent betWeen cancerous and normal

tissue sites. In Fig. 2 we show a plot for the fluorescence adjoining cancerous

oral tissue sites. The scatter plots for the values L.I.(A) tissue from all cancerous and the normal in

sites of the oral

cavity

arc shown

sites of 12 patients

(4 alveolus. 4 buccal mucosa

Fig. 3. The sensitivity

and specificity

values for

16

t---

'"

Ftc

ORAL:'c'
Fig.3. S",," plo,ft' ,bt 'p"",/ly imwa"d fI~mtm" in"mi,i" Jivm tan"rom IORAL-q and ~,,"a' IORAL-N)';", nf,}" nml ,auiry I "andad ,",iaNnn. "" ,ht ",,' >amp/' iu in""iga,,d. Th, b", ,how mtan "',, ,

1600 F I (a. u.) 1400

1
0

I. I. .

00

Iii I::i:
~

i +~. i ~
2 4

t
8

t t
-9
tinut>ampktJivm

PATIENT
Fig. 4. &m" plotft, tht 'P"aI/y in"f7a"dfl."""'mtt imtmiry nfpaim!

NUMBER
,"'urow and adjoining nanna/Im..t

'"parim..

117 r-

dimiminating rissues were

cancerous -90% 0=

from the

notmal sample

oral size

specificity,

the discrimination

results were poor

invesrigated". pmmetetS nore rhat

Use of a stepwise

mulri-varia improvement to of a since parameter

for discriminating cancerous from benign tumor tissue. With 337 nm excitation, the use of fluorescence tissue from, intensity both as a and discrimination cancerous tumor benign

linear regression (MVLR) analysis with ten input led to only a muginal suits". having a discrimination in the discriminarion based only on L,I,().) much simpler It is pertinent

parameter could howevcr, discriminate normal

h", the advantage =gemcnt,

rissue, with sensitivity and specificity values of> 99% ". Excitation-emission spectroscopy" and timeresolved m=utements. carried. out on breast tissue autofluorescence variation malignant, types. benign of have revealed a significant of fluorophores and normal of in the tumor rhe tissue

experimental

no spectral resolution is required. It can however be used as a good discrimination whcn rhe the diflerence and normal possible in malignant parameter only values the to for intensity due to

sites is much more than various in-vivo The

in the concentration Some

variations

cxperimental UF

factOrs. Applicability is expected

predictions

studics therefore

needs to be investigated.

based approach

to lead to an of preno

spectroscopic analysis have been confirmed by biochemical estimation". This variation in the concentration of the fluorophores explained why the discrimination resultS obtained with 337 nm excitation obtained we" with much better compared to that the use of orber excitation

early diagnosis and may allow detection

malignant alterations for which ptesently dlective non-in=ive method exists. Breast cancer Autofluorescence with N, laser tissue comp=d tissue""'. fluorescence sires and cancerous fluorescent benign integrated cancerous spectroscopy excitation was of bre"'t sbowed considerably A scatret intensity adjoining thar

wavelengrhs by other researchers". tissue rhe more and the paired breast Although b,,"'t cancer is not a superficial canCer

disease, the UF based diagnosis of brmt can be conveniendy the best available

done during needlc biopsy. means of detecting br=t of false (60-90%) turn biopsy,

to the normal from normal

This is of interest because X-ray mammography, cancer at present, has two important positives, i.e., a vety large ptOportion of mammograpbically out to be benign leading Stress to patients has potential ",sociated Furtbet, to avoidable abnormal trauma upon invasive br=t drawbacks.

tumor

plot for the

First it leads to a vety large number

tissue sires from 10 patients

is shown in Fig. 4. intensity for and bcnign

The ratio of the mean fluorescence cancerous sites to that from normal tumor sites were .2.82

detection

and 3.23 respectively in

and psychological frequent exposure remote. Laser

the study involving 63 patients, 28 with ductal carcinoma and 35 with fibroadenoma". With 300 nm and 488 nm excitation sites were more However, fluorescent rhan the canccrous the normal. diffetence intensity of

n. Secondly,

to ionizing x-ray tadiation hazards,

during mammography the adverse effects radiation.

howsoever

screeuing can be used without with

no statistically

significant

the use of ionizing

was observed

in the fluorescence

the tesultS of the in-vitro studies suggest may offer much imptoved

cancerous and benign tumor sites. Therefore, while cancerous tissue could be discriminatcd from normal with good sensitivity and

that the UF technique specificity.

J 18 I--

Uterine

cancer

differences requites that the fluowphores spectta

present and

The N, laset excited, autofluo<cscence

in the different tissue types ace identified their telative concentrations estimated. The tesulrs of exciwion/

fmm uwine tissue also showed significant differences between notmal and cancemus tissue. The diffecences in the spectra using a stepwise disccimination scote MVLR analysis could we" quantified

emission""',

MVLR analysis. The based on a 4 vatiable disctiminate cancewus

synchwnous luminescences"'" and time-tesolved studies'" cattied out at CAT on breast and oral cavity tissues suggest a significant vaciation in the concentwion while of the fluowphmes of NADH in the different is higher in tissue types. In particular, concentration the studies reveal that

sites fwm notmal with sensitivity and specificity values of gteater than 85% in genetal and up to 100% when the cancewus site showed red fluorescence porphyrins". characteristic of endogenous

malignant breast tissues compaced to benign tumm and normal breast tissues", the "verse is ttue for tissues fwm oral cavity where NADH

Reasom

for

the spectra!

differences

concentwion is higher in nmmal oral rissues". These results have also been confirmed by for rhe benign vety in on help emymatic concentration fluowphore spectroscopic observed autofluorescence tissues. measurements in malignant concentration studies spectral spectra of and normal inferrcd explain in NADH tissues fwm the the

An understanding the significant autofluorestence important. oprimization the tissue. This

of the factors tesponsible differences spectra may between of cancewus, nor only

fwm breast and oral cavity". The differences in qualitatively differences

tumoc and normal

tissue sites is deady

of the diagnostic Comprehension

system but may of the spectral

also pmvide valuable biochemical infmmarion

of the oral and breast

F;g.5.

A "b,m";, vf,h, p""'yp'

LIF ba;d 'pOemdmi,pdat

C4Tjb, ;n-v;vvtNdi" vn can",d;agn"".

19

I--

In-vivo studies A schematic of the UF based system developed at CAT for in-vivo elinical studies is shown in Fig. 5. It consists of a sealed-off N, laser (7 ns, 100 I'}, 10 Hz), an optical gareable diagnostic quartz and iotensified probe, CCO developed fiber probe, and a detector. in-house The is a fIber

polychromator coupled to the intensifIed CCD. The fiber bundle is enclosed in an 55 tube (9 mm outer diameter). The tip of the probe is betWeen the fluor=ence of the of the

shielded by a quartz optical flat (2 mm thick). This is to provide a fixed distance tissue and the fibers for improved collection a.=mbled and to protect

contamination

fiber tips wirh body fluids. A photograph unit is shown in Fig. 6.

bundle, whieh has tWOlegs; one contains a single tibet (NA 0.22, contains core diame<er 400l'm) six quartz fIbers (NA fiber The central the other

One unit has been installed at a cancer screening center at Indore and is being used to record aurofluorescence spectra of different regions from

0.22, core diamete< 400l'm). the six surrounding fluorescence from the illuminated fIbers

delivers excitation light to the tissue surface and fibers colleCt tissne surface area directly light. The slit light of a

uterine cervix of patienrs screened for neoplasm of uterine cervix. The orher unit at CAT has been used to car')' with out a pilot stUdy on 25 confirmed

by the excitation at the

coming from the distal ends of the six collection is imaged entrance

patients squamous

histopathologically

cell carcinoma

of oral caviryD. The

Fig. 6.

Aph"r'Wnph

"frhr LIFb""dry",m

4wkp,da<CATftrcnnmdingna<;'.

I 20 I----

discrimination differentiare semiriviry 86% and

algorithm rhe squamous

developed cell carciuoma ti"ue towards The

could of rhe wirh a of

region of

surrounding

rhe cancero", Indeed,

rurnm when

may rhe

have biochemical rhe discriminarion

changes due to rhe field.effect analysis was carried our on rhe averaged over all normal

oral caviry from normal and 63%,

"Iuamo",

malignancy.

specificiry, The

cancer,

respectively, reason

resultS are

basis of the spectra averaged over all cancerous sites and the spectra sites from a patient, a sensiriviry and specificiry towards cancer of 100% were obrained. The remarkably averaged normal good spectra results suggests obrained rhar on sire

shown in Fig. 7.

for rhe relatively in rhis

lower specificiry values appears ro be rhe fact rhat most of rhe parienes who panicipated Some of rhe visually uninvolved be nmmal This may therefo because follows the study were at an advanced stage of malignancies. sires assumed to appearing nOt be truly normal. normal

a few of rhe

sires had signarures

very different from

the orher sites of the group.

1.0-1"

i'~- . . . .;. ~

:
0

0 0
0

g8
00 0'J} ,t
0

ni/iII!!:!,

:s =
,Q
Q., ... = 'C

. .~1?~ .It .1Ii'?"

.
Ii"

2 '"

J:. , .... ..... ... .. . '. . ." . . I .. .. .III

I

:.~. .'"

(J0 '0
o~ '1: 0 0

80~
0

i'~i
~"8

0 ~o~:
f!.{{p

Site number

F;g.7.

Th, p"'"i,,

p"babiliri" 'Ib,ing ""'ifi,d",

'q..m,m ,,1/ wdnoma (SCC)I" ,h, ,iou, ,i", in""iga,,d.

I 21 I--

Studies on Photobioactivation
The u,e of light by for non-,urgical photochemical of rherapeutic reactions natural

Furmer,

me

fact mat was

an

inhibition at higher

of cell dose

proliferation highlights appli"",io", initiated chromophores tberapeuuc tuberculo,is, inadiation exploits me parametric an important been studied

observed

the importance dependence role

of a carefrd study of in order ro elicit me of wound

pboto-excitation or exogenous

dC1iired clinical response. Since macrophages play in rhe process healing, effi:cr of light on macrophagC1i bas also and stimularion of macrophagC1i observed following He-Ne laser inadiation". The studi", on narrow bandwidm E-coli baCterial ')'stems canied lighr <fTem on

drugs localized in example.. of the are therapy its use in

the tissue. Some well-known use of light"

the teeatment of psoriasis, neonatal jaundice, skin photodynamic wirh narrow of cancer, mat light Oaser) etc. Several ",;carchers can have profound have also reported bandwidth

Out at CAT have

effect on cellular cultures and

animal models' and can alw lead to rherapeutic effects in humans'."', like, accelerated wound hcaling, treatment of pain nf different applicatinns is gtowing origins, it has

provided two inreresting remlrs. First, ir has been observed" that He-Ne laser irradiation can stimulate through components. respiratory change electron transfer process

in redox

stace of mpirarory of ATP, the

etc.. Although

the usage of lasm for non-,urgical clinical modality"

Since me production

laser therapeutic

not yet become an established because the mechanisms thetapeutic inexpensive considerable substantial applications generating effem The clinical porential

source of cellular energy, i, linked to eleCtron transfer such stimulation can lead to enhanced metabolism. different components phororecepror cytochrome thar several protection imensity, incubation subsequem He-Ne Ecoli Experiments for me that las ,uggesr ar He-Ne laser canied the our wing chain primacy is

for many of rhe photo-

ace not vety well undersrood. of mi, tatber simple and

inhibirors

respiratory

therapeutic modality is motivating work in this direction and progtess any is being conventional made". light It is

wavelength pee-irradiation irradiation magnitude the The period

d. Secondly, it has been observed"'" (632.8nm) rowards uve The and in of of aud

perrinent to note rhat for these phoro-therapeuric source and with the appropriate wavelength

induces protection

strains. rime

was found to depend on He-Ne laser exposute between uve He-Ne inadiation. laser exposure

the desired paramerers (engy, pulse duration etc.) can be used. However, rhe berrer control on laser light characreristics more convenient ofren makC1i

results also

phototherapy lasers.

with rhe use of

suggest involvement

of singler oxygen in the HeRecent experiments 'uggesr mat rhe to He-Ne laser of ph, gene and

Ne laser induced protecrion. out to induced irradiacion protection induced is due induction

At CAT ,everal studies have been canied

with E-Co/i strain KY706/pPL-I

investigate the effect of laser radiarion on cellular culrures and animal models. Our studies" on rhe effecr of N, laser irradiation on the skin of animal models (albino rabbirs and mice) showed that N, laser inadiation to a proliferation layer of me rabbit/mice of cells can comribute wounds reponed in at cerrain doses can lead skin. Such proliferation to fuster healing of me several clinical studies. of cells in rhe active epidermal

acrivarion of 50S repair. SrudiC1i carried showed oxidarion that our on red blood N, laser inadiation Mechanistic cell lysate" induces studies ted blood is

of hemoproreins.

and experiments

wim deoxygenaced

cell lysate rule out involvecnem

of any reacrive

oxygen species and suggests mat rhe proms

122

I-

photOchemical electwphmesis

and not intrinsic action.. '"

photosensitiw A singl~ cell gel and DNA damage

bluccing of ehe image. Sevecal appwaches fwm the backgwund-scatteeed light"

can be One and w

induced photodynamic med to investig"e in mice pedton,,1 Studies on

used to pick out the useful image beating light appwach is w exploit the fact that the scattered

up was also developed UVA induced macwphages"'. photodynamic effect

light emerges fwm ehe dssue in all dictions of also takes longer time to emerge as compad the component any scattering scattered. oe is peedominantly using sp"ial duration) can be filted achievable imaging mpmal

photnsensidsm on cellulae cultutes and animal have also been initiated" CAT. Studies wece c",ied psence both out on the photodynamic of methylene and TB inactindon of in Bacillus ,"brill, cells by He-Ne la"e itradi"ion blne (TB). Results show th" MB lead to genemion

of light which does not undergo foewatd filtecs and by beating ehe

Therefo,

use of ultta-shott sub-pico"cond components 'patial be solution fm

gates (of picosecond I image out. Though

blue (MB) and toludine outside: the cell photodynamic

in these techniques these can not laege dssue beaeing in One

of singlet oxygen causing a decase in

is vel)' high (few tens of micwns), used thwugh thickness since the amount

cell memb"ne fludity. Howevet, whereas TB eenuins outside ehe cell and does nm lead to genecation photodynamk of feee "dicals, action, in MB supewxide mediated cadica!

of image

lighe hils off exponentially the thickness appwach optical to addss eechniques.

with the incase medium.

of the scatteeing this pwblem Experiments

generated int"cellulacly to the cell damage". c",ied AlA mice. out photodynamic induced Effect

is the majoe contributm have also been and and on beating and effect of MC540

is to ',electively on optical

Investig"ions

amplifY the image beacing light using non-lineae imaging through wlution stimulated of eutbid media have been c",ied

AlA on cell lines of epithelial neoplasm accumulation of glucose (Pp) intumm and skin of fibwsaccoma p-treatment

of pwtopotphydn

our ar CAT using a Streak Cameca with tempoca! Raman scattering (SRS) appwach of

- 10 ps and also by urilizing

feactionated delivery of AlA on Pp accumulaeion in tumoe and tumoe investigated. A pilot to skin mio study on of Pp was photodynamic

rime garing. A sparial solution

foe

300 ~m has

been obsecved in rhe present set-up.

thecapy of animal tumms " RMC to chatacteei"

has also been weied a photosensitisee Medicine dcug Centre

Development

of Laser Optical

Trap-

our'". This was a collaboeative wack with a gwup p"ented jointly by Radiation

Microbeam Ser Up
Development finding of laser trap and micwbeam up ar CAT becau" applications Laser lasee beam micwmanipularion micwbeam coupled set up it is of is to a at Cut, arise

(RMC), Mnmbai, and liT, Mnmbai.

has also been taken widespread non-contact objects". a pulsed for

in biomedicine

Development Imaging

of Techniques for Optical

micwscopic essentially micwscope. W/cm'

Optical imaging has the pmential w pwvide submillimme foe ionizing fundamental th" eesolution "di"ion pwblem imaging without with optical optical which the need imaging photons is ace and associated eisks. The

The lacge intensities

(of up to 10" generated used to apetture

with UV lasec micwbeam) objecrive can be

rhe focal point micwscope perforate micwmetee

of the large numerical

in contrast

w Hays in tissue,

m fuse micwscopic

objects with sub-

StCongly scattered

leads to a

accuracy. As these intensities

123

I-

only ac the focal point. opening it. Optical

it is possible to work object without

bandwidth models

light on celiular cultUres and animal tesponsible for

within the depth of a transpacent

so that the mechanisms

tWeeze, or laser optical trap

uses the light of a CW infrared laser for transpOt( of microscopic objecrs. Here. the gradient forces laser beam and the resulting objecrs at the is gentle and arising due to the lacge gradient of light intensity in rhe focussed gradient

the various photo-therapeutic effecrs are better understood. Several studies in this direction have also been carried out ac CAT.

Acknowledgments
The author will like to thank Biomedical Applications his colleagues at and Laser

in the light pressure and the electric Section

field ace used to trap microscopic microtools. absolurely move and panicles made the optical trap

focal point of the laset beam. Unlike mechanical stetile and can be used to capture, position single cells or subcellular contact and and or significant been used to trap motile bacteria without direct

Applications and Electronics Division, CAT who have contributed to the wack described in the paper. It is also a pleasure to thank Bhawalka<. Ditectot. encouragement. Dr. D. D. and

CAT for his suppott

damage. A laset trap set up has recently operarional ar CAT poiystyrene "'""damo"", microsphetes and E-Coh).

References 1. See for example (i) Wolbacsht, M. L, ed.. "Laserapplicationsin medicineand biology", Vol. ]-5, Plenum Press, New York, 1971, 1974.1977.1989 and 199]; (il) Caro. R C. and Choy. D. S. J.. (Guest eds.) Optics and Photonics News. Specialissueon Optics and Light in Medicine, Oct. 92, pp. 9.44 and (iil) Gupta, P. K.. in "Medical Physics for Human Health Cace". (eds. Bhamagat. P.K.. Pradhan. A S., and Reddy. A. R). Scientifie Publishers,Jodhpur, 1997, pp. 143-161 and referencestherein. 2. Servick-Muraca,E.. and Benaron, D., (eds.). "OSA Trends in Optics and Photonics on Biomedical Optical Spectroscopy and Diagnostics", Optical Society of America. Washington, D.C.. Vol. 3. 1996. 3. Wagnieres, G. A, Srar, W. M.. and Wilson, B. C.. Photochem. PhotobioI. 1998. 68. 603-632. 4. Special Section: Biomedical Applications of Lasers,CurroSci.. 1999,77,885-941.

Conclusions
There exist considerable oflaser-based current interest in the for in-,itu.

development

techniques

near teal time diagnosis. Sevetal studies have been carried our at CAT on laser induced fluorescence biopsy from of tissues resected pacients suffering at surgety from cancer or of

oral cavity. breast or uterus. These studies have provided very encouraging resulrs on discrimination of cancerous tissue sites from benign tUmor tissue Ot normal systems have been tissue sites. N, developed for

laser based

clinical studies and ace being used for in-vivo studies on the diagnosis of cancer of oral cavity and urerine cervix. A pilot study on 25 patienrs wirh histopathologically confirmed squamous cell carcinoma of oral cavity has been completed results obtained. endogenous and

and satisfactory discrimination Photobioactivation exogenous using

chromophores

also holds considerable applications.

promise for a variety of therapeutic However, realizations careful studies on

of this potential requires the effects of narrow

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5. Karu, T I., "Phorobiology rherapy", Harwood London, 1989. 6. Ohshirao,

oflow-power

laser

Aeademic

Publishers,

T, and Calderhead,

R. G., "Low inrroducrion",

level laser therapy a prKrical

John Wiley and Sons, Chichester, 1988; Ohshirao, T, "Low reacrive level laser therapy: pracrical applicarions", and Sam, Chichester, 1991. 7. Hender,"n, (eds.), John Wiley

17. Majumder, S. K, Uppal, A., and Gupra, P. K., in "Trends in Optics and Photonics Volume on Biomedical Speetroscopyand Diagnosis" Eds. Benavon, D., and SevickMuraca, E., Oprical Societyof America,Vol. 3,1996, pp. 142-146. 18. Gupta, P.K., Majumder, S. K., and Uppal, A."LasersSurg. Med., 1997,21, 417-422.
19. Majumder, S. K, Gupta, P. K, Jain, B., and Uppal, A, Lasers Life Sci, 1999, 8, 249-264. 20. Jain, B., Majumder, S. K, Gupta, Lasers Life Sci., 1998,8,163-173. 21. Uppal, A, Majumder, P. K,

B. W., and Dougherty, "Phorodynamic Therapy

J.,

Basic

Principles and Applicarions", NewYork,1992. 8. Baxrer, G. D., "Therapeuric and PraCtice", Chmchill Edinbmgh,1994.

Ma<eel Dekker,

S. K, and Gupta,

P.

lasers, Theoty Livingsrone,

K, Proc. Narional Kanpm,

Laser Sympos., 1998, !IT

pp. 219-220. R. R., in Serviek-Muraca, D., (eds.), "OSA Trends in on Biomedical Oprical Diagnosrics", Washington, Optical D.c., and America,

22. Katz, A, Alfano, 9. Yamamoto, H., Okunaka, T, Furukawa, and Kato, K, H., E., and Benaron, Spectroscopy Society of Hiyoshi, T, Konaka, c., Cun. Sci., 1999,77,894-903. 10. Bhagwanani,

Oprics and Photonics

N. S., Laser News, 1995,6(3),

8-12, and 6(4), 6-8.

11. Gupra, P. K., and Bhawalkar, D. D., Cmf. Sci., 1999,77,925-933. 12. Karu, T I., J. Phorochem. Phorobiol. B: BioI., 1999,49,1-17. 13. Rudolph, W., and Kempe, M., J. Mod. Optics, 1997,44,1617-1642. 14. Greulich, K. 0., "Micromanipulation by light in biology and medicine", Birkhauser Verlag, Berlin, 1999. 15. Majumder, S. K., Gupta, P. K and Uppal, A.,LasersLifeSci,1999,8,211-227. 16. Majumder, S. K., Gupra, P. K and Uppal, A, in "Oprical Diagnosrics of Biological Fluids III", (ed. Prienhev, A V.), Proc. Soc. Phoro-Opf. 1nstrum. Eng., 1998, 3252, pp. 158-168.

1996, Vol. 3. pp. 132-133, therein.

and references

23. Maranro, G., Scientific American, 1996, 275,113. 24. Majumder, S. K, Uppal, A, and Gupta, P.K, Cun. Sci., 1996,70,833-836. 25. Majumder, S. K., and Gupta, P. K., in "Optical Diagnosrics of Biological Fluids III", (ed. Priezzhev,A V.), Proc. Soc.PhoroOpt. Instrum. Eng., 1998, 3252, pp. 169178. 26. Majumder, S. K., and Gupta, P. K, Lasers LifeSci., 2000, 9, 143-152 27. Majumder, S. K., Mohanty, S. K, Ghosh, N., Gupta, P. K, Jain, D. K, and Khan, F., Cun. Sci., (in press).

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28. Regan, J. D., and Parcish, J. A., (eds.), "The science of photomedicine", Plenum P<ess, NewYork,1982. 29. Basford, J. R., Lasers Surg. Med., 331-342. 1995, 16,

34. Kohli, R., Gupta, P. K, and Dubey, A , Radiation Research,2000,153,181-185. 35. Murali Krishna, c., Bose,B., and Gupta, P. K, Radiation Research,2000, 153,411-415. 36. Bock, c., Dubey, A., Greulich, K 0., and Gupta, P. K, Mutation Research,1999, 439, 171-181. 37. Dubey, A., Bansal, H., and Gupta, P. K, Indian J Biochem. & Biophysics, 2000, 37, 245-250. 38. Murogesan, S., Gupta, P. K, Sharma, M., Dube, A., Noronha, O. P. D., Samuel, A. M., Sherry, S. J., Srivastava, S., Khare, R., Bharnagor, R., and Parhak, S., Presented at XV A>ia Pacific Cancer Conference, 12-15 December, 1999, Cancer Institute, Chennai.

30. Agnihotri, S., Sachdeo, S., Sharma, A., Kemi, Y., and Gupta, P. K., LasersLifeSci, 1997,7,227-235.
31. Bansal, H., Dubey, Proc. National 1998, pp. 227-228. A., and Gupta, P. K., Laser Sympos., !IT K,npur,

32. Dubey, A., Gupta, P. K, and Bhatti, S., LasersLifeSci, 1997, 7,173-180. 33. Kohli, R., Gupta, P. K, and Dubey, A., in "Biologiceffecrsof light 1995", (eds.Holick, M. F. and Jung, E.G.), Walter de Groyter and Co. Berlin, 1996, pp., 243-245.

Abaut the auth".. (MSr., Lurk",w Uni"mity, 1973) i"iucd the mtwhile La", Sreti", BARC, in ,ating thwugh the BARC haining Seh"l. Hi, ""a"h intem", while at BARC, included n,,-line., aptiealfrequency ,,""minn, CO, I.,m and CO, /a", pumped malew/a, g" Imm. Dr. Gupta war at H"int Watt Uni",ri'y, UK frum N"emb" 1979 to N"emb" 1981 " a Camm"wealth Schola"hip Awa,d and late' nn a pnttdnttocal ftllawrhip dueing 1988-89 He ubtained hir Ph.D. "'pre fram H"int Watt Univmity in 1981 ft' hit w"k nn 4fttient midinfra"d genreati" by optical pumping mnlreul., garer and ",n-line., aptieal mixing. Dr. Gupta m"ed tn Cen"eftcAdvanrrd he initiated w"k nn CO, I.", pumpedfir,-infra"d Biumedical Applieati'" Seai", the Indian PhyrirrA",tiatinn fir, hit "m,ibutia", at CA T, D,. Gupta h., raclie"eaintd (CAT), Indu", in Ma"h 1990. At CAT, the 1988 NS Satyamucthy Mem"ia/ awaed of

larm and an biamedieal applieati"r aJlmm. He pmently hradr the un mid-infra"d "hennt "U"".

. .

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