Medical Microbiology

TetramertestfordetectionofspecificTcells(Fig.2. 32 ,p. 1 34):recombinant
MHCclassIantigencoupledtobiotin,labeledwithavidin,andcorrectly
foldedtogetherwithpeptideandb 2 microglobulin,formstetramers;which
arerecognizedbyspecificTCRs.Subsequentanalysisoftetramerbinding
usingFACSequipmentisbasedonthecolorindicatoroftheavidin(fluores-
cein,phycoerythrin,etc.).TetramersspecificforMHCclassIIantigensplus

ImmunologicalTestMethods 131

RadioallergosorbentTest(RAST)

Addition

of antigen

(solid phase)

Wash

Patient serum with

IgE?

Wash

Addition of

labeled Anti-IgE

Cellulose plate

(solid phase)

Antigen

Anti-IgE

IgE

Fig.2. 29 Thistestisahighlysensitivedetectionmethodforthepresenceofspe-

cificIgEinpatientserum.Antigenisboundcovalentlytoacelluloseplate(solid

phase).AnyIgEintheserumthatbindstotheantigenisthendetectedusingradi-

olabeledanti-IgEantibodies.

ELISPOTAssay

Fig.2. 30 IntheELISPOTassayanti-

gens,orspecificanti-ILantibodies,

areappliedtotheplasticsurface.It

isthenpossibletodeterminethenum-

berofimmunecellsreleasingantibo-

diesspecificfortheappliedantigen,

orreleasinginterleukinsthatarere-

cognizedbytheappliedanti-ILantibo-

dies.Followingincubationat 378 C,

theimmunecomplexeswhichform

aroundthesecellscanbevisualized

usingacoveringagaroselayerwhich

includes an enzyme-coupled anti-

body.Theseenzymescatalyzeacolor

reaction,resultingintheformationof

colorspots,eachofwhichwillcorre-

spondtoasinglecellproducingthe

specificantibodyorinterleukin.

2

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Kayser, Medical Microbiology © 2005 Thieme

All rights reserved. Usage subject to terms and conditions of license.