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1、PurificationandFurtherCharacterizationofan“IBV-like”Virus(Coronavirus)(55074)HAROLDS.RAYE,JOHNC.HIERHOLZER,ANDWALTERR.DOWDLERespiratoryVirologyUnit,NationalCommunicableDiseaseCenter,HealthServicesandMentalHealthAdministration,PublicHealthService,U.S.DepartirtentofHealth,EducationandWeltare,Atla
2、nta,Georgia30333Theadaptationtogrowthinsuckling20%suspensionofinfectedSMBinsterilemousebrainofcertain“IBV-like”virusesphosphate-bufferedsaline(PBS,pH7.2).(coronaviruses)(1)previouslygrownonlyinAntigenictests.CFtestswereperformedorganculture,resultedintherecognitionofbythestandardizedmicrotitert
3、echniquebothcomplement-fixing(CF)andhemagglu-(3).HAtestswereperformedwithPBSdilu-tinating(HA)antigens(1,2).Neutraliza-entand0.5%adultchickenerythrocytes(4).tion,complement-fixation,andhemagglutina-Infectivitytitrations.Assaysofinfectivi-tion-inhibitiontestswerethusmadeavailabletywereperformedby
4、intracerebralinocula-forantigenicandseroepidemiologicstudies.tionof3-day-oldmice.CalculationsofLD50PreliminaryfindingssuggestedthattheCFweremadebythemethodofKarber(5).andHAantigenswerecloselyrelatedandAntisera.Immuneserumwaspreparedinprobablyassociatedwiththevirisn(2).weanlingmicebytwointracere
5、bralinocula-Whethertheantigensmeasuredbytheseteststions21daysapart,followedbybleeding7alsooccurredassolublecomponentsordayslater(1).whethertheyconsistedofmorethanoneUltracentrifugation.Whereindicated,anti-structuralcomponentwasnotdetermined.genpreparationswerecentrifugedat23,800gThepresentstudy
6、describtesasimplemethod(Pi=4.7Xlo8)for30minand/or6,600gforobtaininghighlypurifiedhumancoro.navi-(Pi=1.3Xlos)for20mininaSpinco2No.rusesfrominfectedmousebrainandattempts40rotor.tofurtherdefinethenatureoftheCFandHAantigens.Densitygradientcentrifugation.Equilibsi-MaterialsandMethods.Virus.“IBV-like
7、”umcentrifugationwasperformedinlinearstrainOC43wasisolatedonhumanembry-25-55%sucrosegradients.Amilliliterofonictrachealorganculture(HET5)andsub-crudeorpurifiedviruswaslayeredonthesequentlyadaptedtosucklingmousebrainsurfaceandbande
