chapter6AntibodyAffinityIsaQuantitativeMeasureofBindingStrengthThecombinedstrengthofthenoncovalentinteractionsbe-tweenasingleantigen-bindingsiteonanantibodyandasin-gleepitopeistheaffinityoftheantibodyforthatepitope.Low-affinityantibodiesbindantigenweaklyandtendtodis-sociatereadily,whereashigh-affinityantibodiesbindantigenmoretightlyandremainboundlonger.Theassociationbe-tweenabindingsiteonanantibody(Ab)withamonovalentantigen(Ag)canbedescribedbytheequationk1Ag�Ab34Ag-Abk�1IStrengthofAntigen-AntibodyInteractionsICross-ReactivityIPrecipitationReactionsIAgglutinationReactionsIRadioimmunoassayIEnzyme-LinkedImmunosorbentAssayIWesternBlottingIImmunoprecipitationIImmunofluorescenceIFlowCytometryandFluorescenceIAlternativestoAntigen-AntibodyReactionsIImmunoelectronMicroscopyAntigen-AntibodyInteractions:PrinciplesandApplicationsT--lecularassociationsimilartoanenzyme-substrateinteraction,withanimportantdistinction:itdoesnotleadtoanirreversiblechemicalalterationineithertheantibodyortheantigen.Theassociationbetweenananti-bodyandanantigeninvolvesvariousnoncovalentinterac-tionsbetweentheantigenicdeterminant,orepitope,oftheantigenandthevariable-region(VH/VL)domainofthean-tibodymolecule,particularlythehypervariableregions,orcomplementarity-determiningregions(CDRs).Theexquis-itespecificityofantigen-antibodyinteractionshasledtothedevelopmentofavarietyofimmunologicassays,whichcanbeusedtodetectthepresenceofeitherantibodyorantigen.Immunoassayshaveplayedvitalrolesindiagnosingdiseases,monitoringthelevelofthehumoralimmuneresponse,andidentifyingmoleculesofbiologicalormedicalinterest.Theseassaysdifferintheirspeedandsensitivity;somearestrictlyqualitative,othersarequantitative.Thischapterex-aminesthenatureoftheantigen-antibodyinteraction,anditdescribesvariousimmunologicassaysthatmeasureorex-ploitthisinteraction.StrengthofAntigen-AntibodyInteractionsThenoncovalentinteractionsthatformthebasisofantigen-antibody(Ag-Ab)binding...
