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1、42~tabBRhmentofanELISA.BasedPrimaryAntiviralScreenandPlaqueReduetlonBased~ryAssayforEvaluationdCompoundsagainst~eFilovirusEbolaUnderMaximumBiologicalContainmmztCondltlmm(BL-4).J.W.Huggins,Z.X.ZhangandT.P.Monath.DepartmentofAntiviralStudies,VirologyDivision,U.S.ArmyMedicalInstituteofInfectiousDisea
2、ses,FortDerrick,Frederick,Maryland,21701-5011,USAPrimaryEbolaoutbreakshavebeenassociatedwithhighmortality(40-90%)inman.Therequirementformaximumbiologicalcontainmentfacilities(BI~4)andlackofahigh-volumedrugscreenhaveinhibiteddrugdevelopment.Recently,infectionsassociatedwithimportedmacaquesinaU.S.pr
3、imatecolony,coupledwithEbola'spotentialforseriousdiseaseoutbreaks,emphasizedtheneedforapriorityefforttoidentifypotentialantiviralcompounds.ConfluentmonolayersofVeroE6cellsinmicroculturoplateswereinfectedwithEbolavirusatanMOIof10-2.Dilutions(1:2)oftestcompoundwereadded(intriplicate),incubatedat37oc
4、(Mayingastrain5days,Bonifacestrain4days),cellmonolayersfixedwithformalin-10%calciumchloride,anddecontaminatedwith106radsofTirradiationpriortoremovalfromBL-4containment.ViralantigenwasdetectedonfixedcellmonolayerswithamixtureofEbolamonocionesbyELISA.Theassaymeasuresviralreplicationandsubsequentinfe
5、ctionofadditionalcells.ToxicitywasdeterminedonmicrotiterplatesofconfluentmonolayersofVeroE6cellsinparallelwithanMTFassaybasedonmitochondrlalenzymereductionofcolorlessMTTtetrazoliumsalttothecoloredMTTformazanbyviablecells.ActivecompoundswereconfirmedinaplaquereductionassayinVeroE6cellsunderB1-4cond
6、itions.Twoactivecompoundshavebeenidentified.43Summaryc:.,fNatu:alProductsIsolatedfromMarineOrganismswithAntiviralActivityagainstMouseCoronavirus.S.CrossandL.Moritz.HarborBranchOceanographicInstitution,5600OldDixieHighway,Ft.Pierce,Florida,34946,U.S.A.Marineorganismsincludingalgae,sponges,softcoral
7、sandspeciesfromotherphylahavebeenscreenedforantiviralactivityagainstmousecoronavirusstrainA59.Thisstrainwasusedfortheantiviralscreen,becausesamplescouldbeprocessedandresultsobtainedin24hours.Anumberofnewandnovelc
