乙型脑炎病毒E蛋白抗原表位E39特异性单克隆抗体的制备  

Preparation of monoclonal antibody specific to envelope protein epitope E39 of Japanese encephalitis virus

作　　者：华荣虹[1] 王斌[2] 陈娜莎[2] 于海[2] 刘天强[2] 马红[3] 刘娣[1] 童光志[3] 

机构地区：[1]东北林业大学博士后流动站 [2]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室,黑龙江哈尔滨150001 [3]黑龙江省农业科学院畜牧研究中心博士后工作站,黑龙江哈尔滨150086

出　　处：《中国兽医科学》2008年第2期133-136,共4页Chinese Veterinary Science

基　　金：国家自然科学基金项目(30700027);中国农业科学院哈尔滨兽医研究所所长基金项目(2006,2007);黑龙江省博士后基金项目(LBH-Z05213)

摘　　要：为了制备乙型脑炎病毒E蛋白抗原表位特异性单克隆抗体,将编码乙型脑炎病毒E蛋白抗原表位E39的DNA序列进行人工合成,随后插入表达载体pGEX-6p-1的限制性酶切位点BamHⅠ与XhoⅠ之间,构建了表位肽与GST的融合表达质粒。该表位融合蛋白经亲和层析纯化后免疫BALB/c小鼠,按常规方法进行细胞融合。以化学合成的E39表位多肽为抗原,对融合细胞上清进行间接ELISA筛选。结果,筛选出1株分泌E39表位特异性抗体的杂交瘤细胞株,该杂交瘤细胞分泌抗体能力稳定。经鉴定,该单克隆抗体亚型为IgG2b,轻链类型为κ链。结果表明,用表位融合蛋白为抗原可以制备表位特异性单克隆抗体。The DNA sequence encoding envelope protein epitope E39 of Japanese encephalitis virus was synthesized and inserted into prokaryotic expression vector pGEX-6p-1 between sites BarnH Ⅰ and Xho Ⅰ . BALB/c mice were immunized with the purified epitope fusion protein GST-E39 expressed in Escherichia coli BL21. The myeloma cells SP2/0 were fused with splenocyte of the immunized BALB/c mice. Hybridoma cell lines were screened and selected by an indirect ELISA using the synthesized peptide E39 as antigen. One hybridoma cell line 5G7 was identified and selected for stably secreting epitope E39-specific antibody. The monoclonal antibody was identified to be IgG2b subtype,and its light chain was κ chain. The results confirmed that the epitope-specific monoclonal antibody could be successfully prepared with the epitope fusion protein as antigen.

关 键 词：乙型脑炎病毒 E蛋白 单克隆抗体 表位 

分 类 号：S852.659.6[农业科学—基础兽医学]