Description:1.IntendedUse TheGenWayMycoplasmapneumoniaeIgAantibodyELISAkithasbeendesignedforthe detectionandthequantitativedeterminationofspecificIgAantibodiesagainstMycoplasma pneumoniaeinserumandplasma.FurtherapplicationsinotherbodyfluidsarepossIBLeandcanbe requestedfromtheTechnicalServiceofGenWay. Thisassayisintendedforresearchuseonly. 2.GeneralInformation MycoplasmsbelongtotheMollicutesclass.Commoncharacteristicsofthesixeubacterialgenerais thelackofabacterialcellwall,osmoticfragilityandsmalldimensions,whichallowapenetration througha0.45µmfilter.Alsothegenomewith 600kbpissignificantlysmallercomparedwith gram-positiveandgram-negativebacteria.Outofthisreasontheyhaveneverbeenfoundasfreely livingorganisms.InnatureMollicutesdependonahostcell,respectively,onahostorganismlikea parasite. Mycoplasmapneumoniaeisahumanpathogenicbacteriumcausingtracheobronchitisandprimary atypicalpneumonia.Associatedwiththehostcell,surfacecolonizationofhumanrespiratorytract epithelialcellstakesplace.Alsosecondary diseaseslikeinfarction,encephalitis,chronic neuropathyandtheGuillain-BarresyndromecaninsomecasesbeconnectedwithaM.pneumoniae infection. Inthelaboratory,M.pneumoniaecanbegrownwithoutahostcellinrichmediumsupplemented with10-20%horseserum.BesidesthecoldagglutinintestandcomplementfixationreactionCF, ELISAisthemethodofchoice,whichshowsanexcellentsensitivityandthepossibilityto differentiatebetweentheimmunogloblinclasses. SpecificIgAantibodiesweredevelopedmoreregularlyandmorerapidlythanIgMduringanacute infection.IgAtitresalsostartedtodecreaseearlierthanIgMorthelate-peakingIgGresponse. Itcouldbeshowninvariousstudies,thatthedeterminationofallthethreeimmunoglobulinclasses isnecessary,tomonitoreachstepofthecourse.3.PrincipleoftheTest TheGenWayMycoplasmapneumoniaeIgAantibodytestkitisbasedontheprincipleofthe enzymeimmunoassay(EIA).Mycoplasmapneumoniaeantigenisboundonthesurfaceofthe microtiterstrips.Dilutedsampleserumorready-to-usestandardsarePipettedintothewellsofthe microtiterplate.AbindingbetweentheIgAantibodiesoftheserumandtheimmobilized Mycoplasmapneumoniaeantigentakesplace.Afteraonehourincubationatroomtemperature,the plateisrinsedwithdilutedwashsolution,inordertoremoveunboundmaterial.Thenready-to-use anti-human-IgAperoxidaseconjugateisadded andincubatedfor30minutes.Afterafurther washingstep,thesubstrate(TMB)solutionispipettedandincubatedfor20minutes,inducingthe developmentofabluedyeinthewells.Thecolordevelopmentisterminatedbytheadditionofa stopsolution,whichchangesthecolorfrombluetoyellow.Theresultingdyeismeasuredspectrophotometricallyatthewavelengthof450nm.The concentrationofIgAantibodiesisdirectly proportionaltotheintensityofthecolor. AdditionalInformation:NameMycoplasmapneumoniaeIgAELISARelatedProductNamesMycoplasmapneumoniaeIgAELISAIntendedUseResearchUseOnly