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1、Microbiol.Immunol.Vol.25(6),623-625,1981NeutralizingAntibodytoCalfDiarrheaCoronavirusinVariousAnimalSpeciesinJapanKunihikoSATO,YujiINABA,*YasuoMIURA,ShuichiTOKUHISA,HiroomiAKASHI,ITatsuhikoSHINOZAKI,andMinoruMATUMOTO*NationalInstituteofAnimalHealth,Tsukuba,Ibaraki305,De
2、partmentofPediatrics,TeikyoUniversitySchoolofMedicine,Itabashi-ku,Tokyo173,andKitasatoInstitute,Minato-ku,Tokyo108(Acceptedforpublication,March16,1981)Coronavirusinfectionsassociatedwithrespiratory,intestinalorotherdiseaseshavebeenreportedinanumberofanimalspecies,includ
3、ingman,cattle,pig,dog,cat,mouse,rat,turkey,andchicken(1-3,5,6,8,10,12,13).Recentlycorona?viruseshavebeendividedbyimmunofluorescenceintotwodistinctgroupsonthebasisofantigeniccross-reactivity(9),althoughcoronaviruses,asagroup,displaycomplexserologicvariability(5).Inthispa
4、perwereportonthepresenceofneu?tralizingantibodiesagainstcalfdiarrheacoronavirus(6,12)inserumspecimensfromvariousanimalspecies.Calfdiarrheacoronaviruspassagedinbovineembryonickidneycells(6)waskindlysuppliedbyDr.C.A.Mebus,UniversityofNebraska,U.S.A.Inourlabora?torytheviru
5、swasshowntoreplicatereadilyandinduceacytopathiceffectinculturesofacontinuouscellline,BEK-l,derivedfrombovineembryonickidney(4).Inthepresentstudytheviruswasusedatthe7thpassagelevelinBEK-lcells.TheneutralizationtestwascarriedoutbytheserumdilutionmethodinIIX100-mmtubecultu
6、resofBEK-lcells.ThegrowthmediumwasEagle'sminimumessentialmedium(MEM)containing10%tryptosephosphatebroth(TPB)and10%inactivatedcalfserum,andthemaintenancemediumwasMEMcontaining10%TPB,0.05%yeastextract,0.5%sodiumglutamateand0.1%glucose.Serialfourfolddilutionsoftheseruminac
7、tivatedat56Cfor30minweremadeinmainte?nancemediumandmixedwithequalvolumesofmaintenancemediumcontaining200TCIDsoofvirusper0.1ml.Theserum-virusmixtureswereincubatedat37Cfor1hrbeforeinoculationofO.l-mlvolumesintotwotubeculturesperserumdilution.Thetestswerereadafterincubatio
8、ninarollerdrumat37Cfor4days.Theantibodytiterwasexpressedasthereciprocalofthehighestserumdilutionwhichshowedcom
