抗番鴨呼腸孤病毒σB蛋白單克隆抗體的制備.pdf

1、東北農(nóng)業(yè)大學碩士學位論文抗番鴨呼腸孤病毒σB蛋白單克隆抗體的制備姓名：陳曉丹申請學位級別：碩士專業(yè)：臨床獸醫(yī)學指導(dǎo)教師：葛銘20090620AbstractIIIThePrepareofMonoclonalAntibodiesagainstProteinσBofMuscovyDuckReovirusAbstractMuscovyduckreovirus(MDRV)diseaseisonekindofhighincidencehighmt

2、alitymuscovyduckinfectivedisease.Ithascausedseriouseconomiclossesinthemuscovyduckindustrysinceithappenedbecameoneofthemainepidemicdiseaseswhichrestrictthehealthydevelopmentofthemuscovyduckindustryinourcountry.Muscovyduck

3、reovirusisatypeofRNAvirus.TheparticlesaresphericalnearlysphericalIcosahedronwitha60nm~73nmlengthindiameter.ItisSymmetricthreedimensionalnoncapsularhavevisibledoublecapsidstructure.Muscovyduckreovirusiscomposedbysubsegmen

4、tsof10genefragmentswhichcodef10proteinsincluding8proteinsfthecapsidstructuralproteins2nonstructuralproteinsdividingintothreegroupsλσ.AmongthemthestudyofproteinσCisthemesophisticatedwhichisthemajstructuralproteinreculting

5、bytheimmunogenicity.WhileitwasalsofoundinrecentyearsnonstructuralproteinsσBUnitingσCimmunecanplayedbetterrecults.IthasnotbeenreptedabouttheroleofthemechanismofproteinσBinthedomestic.InthisstudypET30σBhavingHistagwastrans

6、fmedintoE.coliBL21(DE3)expressedtoσBwhichis45kDaasexpectedbyRecombinantplassrecombinantedinlabaty.TheHisBindPurificationKitwasusedtopurifytheproteinwhichwastheantigen.TherecultsofWesternblotsanalysisMuscovyDuckReoviruspo

7、sitiveserumfirstantibodyshowedthat.AspecificreactionocceredbetweenσBpositiveserumofMuscovyduckreoviruswhichindicatethatσBhasgoodimmunogenicity.Immunizing8weedsfemalemousewiththedepurantedrenaturatedrenaturatproteintheres

8、earcherpreparedfσBMonoclonalantibodiesofMuscovyduckreovirus.Throughaseriesofbasicidentification4stsofmonoclonalhybridomawereconfirmednamedA3FB2BC3ED2G.MeoverthenaturalstructureoftheproteinswereexpressedinSf9cellsbybaculo

9、virustheimmunohistochemistyrecultsofthe4stsofmonoclonalhybridomashowedthattheyallhavereactionwithnatruelConfmationproteinswhichmeanstheyaregreatvaluable.AlsothepreliminaryepitopeanalysisoftheMonoclonalantibodiesshowedtha