猪圆环病毒Ⅱ型Cap蛋白单克隆抗体的制备及鉴定  

Preparation and Characterization of Monoclonal Antibodies against PCV2-Cap Recombiant Protein

作　　者：元永平[1] 丛丽媛[1] 陈德坤[1] 蒋智勇[2] 张春红[2] 宋长绪[2] 

机构地区：[1]西北农林科技大学,陕西杨凌712100 [2]广东省农科院兽医研究所,广东广州510640

出　　处：《西北农业学报》2008年第1期15-19,共5页Acta Agriculturae Boreali-occidentalis Sinica

基　　金：国家食品安全重大专项(2001BA804A31);广东省农业科技攻关(2003B21404、20052490100和2006B20301013);广东省自然科学基金(04002083);广东省自然基金重点项目(06105311)资助

摘　　要：以纯化复性的猪圆环病毒Ⅱ型（PCV2）Cap重组蛋白作为免疫原，按常规方法免疫BALB／c小鼠，取其脾细胞与SP2／0细胞融合，经间接ELISA筛选，获得了3株分泌PCV2-Cap蛋白单克隆抗体的杂交瘤细胞，分别命名为1C7A4、4F3F5和4G8F7，其染色体平均数为98～103，间接ELISA检测3株细胞培养上清效价分别为1：3125、1：3125、1：15625，小鼠腹水效价分别为1：390000、1：390000、1：1950000。ELISA结果显示，1C7A4、4F3F5和4G87F仅与融合表达的PCV2-Cap蛋白反应，而与PET-32a载体表达的蛋白不反应。相加ELISA结果显示3株单克隆抗体对应两个不同的病毒抗原位点。间接免疫荧光结果显示，1C7A4、4F3F5和4G8F7能与PCV2发生反应，而不与PCV1发生交叉反应。结果表明所获得的3株单抗是PCV2型特异性的，为PCV1和PCV2鉴别诊断方法的建立奠定了基础。BALB/C mice were regularly immunized with redenatured and purified PCV2-Cap recomnbinant protein. Spleen cells were collected and infused with SP2/0 cell. Three hybridoma cell strains against PCV2-ORF2 recombinant protein named 1C7A4, 4F3F5 and 4GSF7 were obtained by indirect enzyme-linked immunosorbent assay method. The average number of chromosome of the hybridoma cell lines was 98-103. The ELISA titer for Cap antobodies in culture supernatant were 1 ： 3125, 1 ： 3125, 1 ： 15625 and ascites were 1 ： 390000, 1 ： 390000, 1 ： 1950000 respectively. The results of ELISA indicated that the three monoclonal antibodies were only against PCV cap recombinant protein but not reacted with PET-32a protein. The results of IFA indicated that the three monoclonal antibodies could only react with PCV2 virus rather than the PCV1 virus, suggesting that these monoclony antibodies were specific to PCV2.

关 键 词：猪圆环病毒Ⅱ型 ORF2重组蛋白 单克隆抗体 

分 类 号：S852.5[农业科学—基础兽医学]