检测猪繁殖与呼吸综合征抗体的重组N蛋白ELISA的建立.pdfVIP

维普资讯 第 19卷 3期 中 国 病 毒 学 19 (3)：250．254 2004年 6月 VIR0L0GICA SINICA June 检测猪繁殖与呼吸综合征抗体的重组N蛋白．ELISA 的建立 许立华 2，苏鑫铭 ，王 玲2，王志亮3，吴延功3，陈溥言 (1．南京农业大学农业部畜禽疫病诊断与免疫重点开放实验室，江苏南京 210095； 2．宁夏大学农学院动物科学系， 宁夏银川 750021；3．农业部动物检疫所 ，山东青岛 266032) EstabUshivientofRecombinantN ProteinBasedELISA f0rDetectionof AntibodyAgainstPorcineReproductiveandRespiratorySyndromeVirus XU Li．hual· SU Xin．mingl，WANG Ling ，WAN G Zhi．1iang W U Yan．gong ，CHEN Pu．yanl ， ． (1．KeyLaboratoryofAnimalDiseaseDiagnosisandImmunology,NnnjgAgriculturalUniversity,Nn内ing210095， China；2．DepartmentofAnima lScience，AgriculturalCollegeofNingxiaUniversiyt,Yinchuan750021，China 3．Anima l QuarantineInstituteofAgricultureMinistry,Qingdao266032，China) Abstract-TherecombinantplasmidpETNwastransformedlntoEcoliBL21(DE3)hostcellandthe expression product-recombinantnucleocapsid proteni ofPRRSV wasobtainedunderhteoptimized condition ofhostcellcultivation na d IPTG induction．Consequently,hte expression productwas purifiedbymena sofhis-bindingresin protein purification procedure．SDSna dWestern-blot wereused to detectthepurification effectand htespecificity ofpurifiedrecombinantnucleocapsid proteinofPRSSV．Th epurifiedNproteinwasusedtocoat96-wellplate，eachstepwasoptimi zed，such ascoatingconcentrationofrecombinnatnucleocapsidprotein，scampledilutim，chromogen(TMB)nad stopsolutionas wellasconcenrtation．Asaresult，na indirectELISA wasestablishedtodetectna tibody againstPRRSV．About200 serum sampleswere detected by htemehtod na d IDEXX ELISA kit， respectively．Th eagreementratiobetweenhtetwomehtodreachedat91％． Keywords：Recombinna