HPV58型E1重组蛋白的表达纯化及其多克隆抗体的制备  

Expression and Purification of HPV58 E1 Protein and Preparation of Polyclonal Antibodies against HPV 58 E1 Protein

作　　者：高锦阳[1] 程一航[1] 唐海平[2] 谢毅[1] 李元春[2] 徐万祥[2] 顾少华[1] 

机构地区：[1]复旦大学生命科学学院遗、传学研究所、遗传工程国家重点实验室,上海200433 [2]国家卫生与计划生育委员会计划生育药具重点实验室、上海市计划生育科学研究所,上海200032

出　　处：《生殖与避孕》2013年第8期505-510,共6页Reproduction and Contraception

摘　　要：目的：原核表达重组人类乳头瘤病毒（human papillomavirus，HPV）58型E1蛋白，并制备兔抗HPV58型El蛋白多克隆抗体。方法：分别构建含编码E1蛋白分子中1～326aafEl却的cDNA片段的表达质粒pET－28b－E1－N，以及含编码E1蛋白分子中319～644aa（E1．c）cDNA片段的表达质粒pET－28b－E1-C原核表达载体，分别转化感受态BL21（DE3）细胞，经诱导表达后，利用镍柱亲和层析方法，分离纯化目的蛋白E1-N和E1-C作为免疫原免疫新西兰大白兔，制备抗El多克隆抗体，Western blotting检测抗体的特异性。结果：制备获得高纯度的El－N和E1－C蛋白，以及高滴度的兔抗E1多克隆抗体，Western blotting结果示，制备的多克隆抗体具有较好的抗原特异性。结论：成功获得兔抗HPV58型E1蛋白多克隆抗体，为E1蛋白分子中线性B细胞表位的鉴定提供了物质基础。Objective： To express and purify human papillomavirus （HPV） 58 E1 protein and to obtain rabbit polyclonal antibodies against HPV58 E1 protein. Methods： pET-28b-El-N and pET-28b-E1-C vectors were consturcted and transformed into BL21 （DE3） competent cells. Target proteins were expressed after IPTG induction and purified through Ni affinity chromatography and then used as antigens to immunize New Zealand rabbits to obtain polyclonal antibodies. Results： Highly purified target proteins were obtained and Western blotting results showed that the prepared polyclonal antibodies had high specificity to HPV58 E1 protein. Conclusion： The polyclonal antibodies against HPV58 E1 protein were obtained successfully, which is helpful for further research on identification of linear B-cell epitopes of HPV58 E1 protein.

关 键 词：HPV58型E1重组蛋白 蛋白表达 包涵体纯化 抗体制备 

分 类 号：R392-33[医药卫生—免疫学]