摘要
应用分子克隆技术将猪瘟病毒E2蛋白部分基因插入到载体pGEX-4T-1中构建重组质粒pGEX-4T-E(A),在大肠杆菌Rosetta中表达融合蛋白GST-E(A),以纯化的融合蛋白免疫BALB/c小鼠,取其脾细胞与骨髓瘤细胞SP2/0进行融合,经克隆化和间接ELISA筛选,获得了C3、A1两株稳定分泌抗猪瘟病毒E2蛋白单克隆抗体的杂交瘤细胞株,ELISA结果显示,其腹水效价为1∶128000和1∶256000。Western blotting结果证实,单克隆抗体C3和A1能与猪瘟病毒(classical swinefever virus,CSFV)发生特异性的反应,表明该单抗是针对猪瘟病毒E2蛋白的保守线性表位。
A truncated gene encoding the major antigenic domains of E2 protein of classical swine fever virus （CSFV） was amplified and cloned into pGEX-4T-1 expression vector to obtain recombinant pGEX-4T-E（A）. The recombinant GST-E（A） protein expressed in E. coli Rosetta. BALB/c mice was immunized with purified GST-E（A）protein as antigen and mouse splenic cells were fused with SP2/0 cells. Hybridoma cells were screened by ELISA and subcloned. Two hybridoma cells C3 and A1 secreting anti-CSFV monoclonal antibodies were obtained. The EI.ISA showed the antibody titres were 1 ： 128000 and 1 ： 256000. Western blotting indicated that C3 and A1 reacted with CSFV specially. These results demonstrated that the McAbs were specific reagent for CSFV, and recognized a linear conserved epitope on the E2 protein.
出处
《中国畜牧兽医》
CAS
北大核心
2009年第9期50-54,共5页
China Animal Husbandry & Veterinary Medicine
基金
安徽省"十五"科技攻关项目(01013003)资助
关键词
猪瘟病毒
E2蛋白
单克隆抗体
鉴定
classical swine fever virus
E2 protein
monoclonal antibody
identification
