Medical Microbiology

terminantsarelocalizedonthesame,orondifferent,antigens;orwhether

differentantibodiescanbindtothesameantigen(Fig.2. 21 ).

RadialimmunodiffusionaccordingtoMancini.Thisisaquantitativeantigen

assaybasedonapredeterminedstandardcurve(Fig.2. 22 ).

Nephelometry.Thismethodmeasurestheamountoflightscatterasaquan-

tificationofprecipitationturbidity.

ImmunoprecipitationCombinedwithElectrophoresis.Antigensaresepa-

ratedinanagarosegelbyapplyinganelectriccurrent.Theantibodiesreactby

migratinginthegel,eitherwithoutanelectricfield,orsimultaneouslywithin

theelectricfield;andeitherinthesamedimensionastheantigensorina

secondverticalstep(“rocket”electrophoresis).

ImmunoelectrophoresisaccordingtoGrabarandWilliams.Inthefirstin-

stanceserumproteinsareelectrophoreticallyseparatedwithinathinagarose

gellayer.Atroughisthencutintotheagar,nexttotheseparatedsampleand

paralleltothedirectionofmigrationalongtheentiremigrationdistance,and

anti-serumisappliedtothetrough.Theantibodiesdiffuseintothegel,and

precipitationlinesareformedwherevertheyencountertheirantigens.The

122 2 BasicPrinciplesofImmunology

DoubleDiffusionAccordingtoOuchterlony

Fig.2. 21 Thistechniquefacilitatesassignmentofantigens(violet)toacertain

testantibody(yellow),orviceversa.Theantigensandantibodiesarepipetted

intotroughswithinthegelanddiffusethroughthismedium(thenumbersdes-

ignatetheepitopespresent).Wheretheymeetlinesofprecipitation(knownas

precipitinbands)develop,indicatingimmunecomplexformation.aTheanti-

bodiesprecipitateidenticalepitopes(epitope1)ofbothantigens,resultingin

formationofprecipitinbandswhichflowtogethertoformanarch,mutuallyin-

hibitingtheirmigration.Inb,threeindependentprecipitinbandsform,indicating

thattheantibodiesdifferentiatethreedifferentepitopesonthreedifferentanti-

gens.cEpitope 1 ofbothantigensamplesformsprecipitinbandswhichflowto-

gether.Anti-2migratesbeyondthelineofconfluenceintotheareainwhichit

precipitateswithfreeantigen 1 , 2 andformsaspur.

2

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Kayser, Medical Microbiology © 2005 Thieme

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