EnzymeimmunoassayforthequalitativedeterminationofIgM-classantibodiesagainstBorreliaburgdorferiinhumanserumorplasma(citrate)andCSF(cerebrospinalfluid).SpiRochetesaremotilebacteriawithaperiplasmaticaxialfilament.AllpathogenicspeciesbelongtothefamilyTreponemataceae,whichincludesthethreegenera:Treponema,Borrelia,andLeptospira.TheTreponemaeareextremelylong,flexIBLe,filamentouscellsthatareusuallyheldinacharacteristicspiral,orcoiled-springshape.BorreliaearethelargestTreponemataceaewithverycoarseandirregularspirals.Borreliaburgdorferi,thecausativeagentofLymedisease,istransmittedmainlybyticksbutprobablyalsobyotherblood-suckinginsects.HABItatsarethewooded,humidandtemperateregionsofNorthAmerica,Europe,NorthAfrica,AustraliaandJapan;foresters,farmers,andanybodyenteringinfestedforestsmaybeaffected.
Thedegreeofcontaminationofticksamountsto3-60%dependentonseasonalandregionaldifferences;upto30%ofthepopulationmaybeinfected(about1500casesannuallyinUSA,severalhundredinEurope).ThemajorconstituentofB.burgdorferiflagellaisflagellin(41kDa,p41).WhereasthelipoproteinOspC(22kDa,p22)withintheoutermembraneofthespirocheteinducesanearlyantibodyformationintheECMphaseofLymedisease.Recombinantp41iisincludedinthepresentedantigenstoavoidcrossreactivitywithantibodiesofsyphiliticsera.TheBorreliaburgdorferiIgM-ELISAcontainsthefollowingrecombinantepitopes:OspCofthephylumPKo(B.afzelii)and20047(B.garinii)andp41iofthephylumPBi(B.garinii).Microtiterstripwellsarepre-coatedwithrecombinantBorreliaburgdorferiantigens.DilutedpatientspecimensandreadytousecontrolsareaddedtothesewellsandantibodiesrecognizingtheimmobilizedB.burgdorferiantigenbindduringthefirstincubation.AfterwashingthewellstoremoveallunboundsampleandcontrolmaterialhorserADIshperoxidaselabelledanti-humanIgMconjugateisadded.Duringasecondincubationthisconjugatebindstothecapturedantibodies,andtheexcessunboundconjugateisremovedbyafurtherwashstep.TheimmunecomplexformedbytheboundconjugateisvisualizedwithTMBSubstrateSolutionwhichgivesabluereactionproduct,theintensityofwhichisproportionaltotheamountofB.burgdorferi-specificIgMantibodyinthepatientspecimen.Sulphuricacidisaddedtoeachwelltostopthereaction.Thisproducesayellowendpointcolour.Absorbanceat450nmisreadusinganELISAmicrowellplatereader.