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1、lournalofImmunologicalMethods,154(1992)195-204195©1992ElsevierSciencePublishersB.V.Allrightsreserved0022-1759/92/$05.00JIM06433Effectoffixationonthedetectionoftransmissiblegastroenteritiscoronavirusantigensbythefixed-cellimmunoperoxidasetechniqueLong-ThhnhT6aa
2、ndSergeBernardbaNationalInstituteofVeterinaryResearch,DepartmentofVirology,Hanoi,VietNam,and~INRA,LaboratoiredePathologicInfectieuseetImmunologic,37380Nouzilly,France(Received5December1991,revisedreceived20February1992,accepted16April1992)Theeffectofvariousfix
3、ativesanddetergentsontheinvitrodetectionoftheviraldeterminantswhichareexpressedinswinetestiscellsinfectedwiththetransmissiblegastroenteritiscoronavirus(TGEV)wasstudiedusingamicrowellimmunoperoxidasetechnique.Whencomparedwithglutaraldehydeandformaldehyde,0.1%pa
4、raformaldehydewasfoundtobethefixativeofchoiceforthedetectionofthesedeterminantsonthemembranesofinfectedcells.Amongdehydratingfixatives,80%acetoneoramixtureofacetoneandethanolorofacetone,methanolandethanolwerefoundtobethebestfixativesforthedetectionoftheseviral
5、determinantswh;chareexpressedininfectedcells.Inthecaseofacetone,thetemperatureoffixationanditsconcentrationinthefixativepreparationwerefoundtobeimportant.Thetreatmentof0.05%glutaraldehyde-fixed,infectedcellswith0.1%saponinor0.1%paraformalde-hyde-fixed,infected
6、cellswithl%NP-40ledtosatisfactorydetectionofviraldeterminants.UsingTritonX-100torendercellspermeable,thequanqtiesofNandMantigendetectedinTGEV-infectedcellsprefixedwitheither0.05%glutaraldehydeor0.1%paraformaldehydewereequaltothoseof80%acetone-fixed,TGEV-infect
7、edcellswhilethequantityofSantigendetectedwasdiminished.Theeffectofotherdetergentssuchaszwittergent,emoigenBB,ChapsandN-lauroylsarcosineonthedetectionofviraldeterminantswasalsostudied.Keywords:Viraldeterminantdetection;Transmissiblegastroenteritiscoronavirus;Fi
8、xative;lmmunoperoxidaseIntroductionmajorstructuralproteins:thenucleocapsid(N),whichisassociatedwiththeRNAgenome;theTransmissiblegastroenteritis(TGE)isahighlypeplomerglycoprotein(S)
