抗幽门螺杆菌rHpaA鸡蛋黄抗体的研制  

Preparation of chicken egg yolk immunoglubin Y(IgY) against recombinant Helicobacter pylori adhesin A

作　　者：黄进[1] 杨致邦[1] 黄伟[2] 毛小琴[1] 邓颖[1] 叶翠莲[1] 

机构地区：[1]重庆医科大学微生物学教研究室,重庆400016 [2]西南农业大学荣昌校区蚕学与生物技术学院生物技术系,重庆402460

出　　处：《中国免疫学杂志》2006年第10期937-939,943,共4页Chinese Journal of Immunology

基　　金：重庆市教委渝教科2003(7号文)资助

摘　　要：目的:采用基因工程技术表达重组幽门螺杆菌黏附素rHpaA,以此蛋白为抗原制备抗rHpaA的高特异性鸡蛋黄抗体。方法:诱导重组质粒pQE30-HpaA在大肠杆菌中大量表达rHpaA蛋白,用rHpaA免疫鸡,以水稀释法联合盐析法提取纯化IgY,采用Bradford法测定含量,SDS-PAGE电泳分析纯度,Western blot鉴定抗原特异性,ELISA测定效价和巴氏消毒后的活性。结果:IgY提纯后含量为24·6mg/ml,纯度约为90%,Western blot显示在相对分子量30000处出现单一条带,ELISA效价为1∶12800,巴氏消毒后未见效价损失。结论:成功获得了高浓度、高纯度、高效价、耐巴氏消毒的特异性IgY,为进一步制备预防幽门螺杆菌感染的特异性IgY制剂奠定了基础。Objective：To prepare a highly specific egg yolk immunoglubin Y（IgY） against recombinant HpaA of Helicobacter py/or from the yolk of hen＇s eggs immunized with recombinant HpaA protein made by gene-engineering. Methods： Recombinant plasmid pQE30-HpaA was expressed in large numbers in E. coli to get rHpaA protein. The recombinant rHpaA was used to immunize the hens. The rHpaA IgY was extracted from the yolk of hen＇ s eggs by water-dilution and salt-dialysis methods. The purity of IgY was analyzed by SDS-PAGE,and protein content of IgY was checked by Bradford method,and the specificity of IgY to antigen of HpaA was assayed by Western blot method. The titer and activity of post-pasteurized of IgY were measured by ELISA respectively. Results：The content and purity of IgY were 24. 6 mg/ml and 90% respectively after purified. Western blot exhibited only one protein band with molecular weight of 30 000. The titer of HpaA IgY was 1：12 800,and the titer of post-pasteurized did not decrease. Conclusion：The specific rHpaA IgY with high concentration, high purity, high titer, pasteurized-stability and specificity was successfully preparated. It might play an important role in further preparing product to prevent the infection of Helicobacter pylori.

关 键 词：蛋黄抗体 幽门螺杆菌 黏附素 

分 类 号：R392.1[医药卫生—免疫学]