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1、ArchivesofVirologyArchivesofVirology80,23--31(1984)©bySpringer-Verlag1984DetectionofBovineCoronavirusinFecesbyReversedPassiveHemagglutinationByK.SAgol,Y.INABAI,S.TOKUIIISAI,Y.MIVI~AI,N.KA~EKOI,M.ASAGI2,andM.MATUMOTO3iNationalInstituteofAnimalI-Iealth,Tsukuba,Ibaraki,Japan
2、2ZEN-NOHInstituteofAnimalHealth,Sakura,Chiba,Japan3KitasatoInstitute,Minato-ku,Tokyo,JapanAcceptedOctober6,1983SummaryAreversedpassivehemagglutination(I~PHA)methodwasdevelopedforthedetectionofbovineeoronavirusinfecalspecimens.Sheeperythro-cytesfixedwithglutaraldehyde,andt
3、hentreatedwithtannicacidwerecoatedwithanti-bovinecoronavirusrabbitantibodiespurifiedbyaffinitychromatographyusingbovineeoronaviruslinkedtoSepharose4B.TheRPHAtestwascarriedoutbyamicrotitermethod.Erythrocytescoatedwithpurifiedspecificantibodieswereagglutinatedbybovinecorona
4、virus,butnotbybovinerotavirusorenterovirus.Thereactionwasinhibitedbyantiserumtobovinecoronavirus,confirmingthespecificityofthereaction.TheI~PIIAtestdetectedbovinecoronavirusin13of22fecalspecimens(59percent),fromnaturalcasesofdiarrhea,whilethepositiverateswereonly14percent
5、(3/22)and22percent(5/22)forimmunofluorescentstainingofprimaryculturesofcalfkidneycellsinfectedwiththespecimens,andimmuneelectronmicroscopyrespectively.TheadvantagesoftheI~PHAmethodareitssimplicity,highsensitivityandrapidity.IntroductionMEBUSandhisassociates(9,Ii,17)havede
6、monstratedanagentwiththemorphologicfeaturesofacoronavirusbyelectronmicroscopyofthefecesofcalveswithneonataldiarrhea,andhaveprovedittobeacausativeagentofthedisease.Thevirusmultipliedinbovineembryonickidneycellcultures,butfailedtoinduceareadilyrecognizablecytopathiceffect.T
7、hereforealthoughthevirusmaybeassayedinthesecultures,themethod24K.SATOetal.:israthercumbersome,asthepresenceofvirusisdetectedeitherbythemicroscopicexaminationofstainedcultures,orbyimmunofluorescencewithaspecificantiserum(10).I~ABAetal.(7)havedemonstratedthatthevirusreplica
8、tesreadilyproducingamarkedcytopathiceffectinculturesofthecontinuouscelllines.BEK-1,derivedfrombo
