幽门螺杆菌vacA基因重组表达的包涵体复性及ELISA方法的建立  

Renaturation of Helicobacter pylori vacA gene recombined expressive inclusion body and construction of ELISA method

作　　者：党双锁[1] 王宏仓[1] 贾晓黎[1] 袁利超[1] 王宝峰[1] 张欣[1] 张正国[1] 程延安[1] 

机构地区：[1]西安交通大学第二医院感染科,陕西省西安市710004

出　　处：《世界华人消化杂志》2005年第20期2500-2503,共4页World Chinese Journal of Digestology

摘　　要：目的:通过对vacA重组工程菌表达产物包涵体的变性、复性,提高表达产物的生物学活性,建立ELISA方法,讨论VacA抗原规模生产的可行性.方法:利用已经构建的表达VacA抗原的工程菌,用IPTG诱导,表达的包涵体通过系列条件的变性、复性、透析,并经活性鉴定.所制抗原用以包被ELISA板,建立ELISA方法并确定抗原的生物学活性.结果:重组工程菌可表达Mr3300的目的蛋白,表达形式为包涵体,条件优化后的包涵体经SDS-PAGE显示纯度达95%以上,活性经ELISA法测定正常人126例和blot电泳测定结果符合率为97.1%.结论:包涵体变性复性后生物活性大幅度提高,并且具有良好的重复性和稳定性,可作为幽门螺杆菌疫苗制备的候选抗原及临床检测的诸多方法的抗原原料.AIM： To increase the biological activity of the expressive product through denaturation and renaturation of H pylori vacA gene recombined expressive inclusion body, and to discuss the feasibility of mass production of VacA antigen. METHODS： The VacA antigen expressed engineering bacterium was induced by Isopropyl-β-D-thiogalactopyranoside （IPTG）, The expressed inclusion body was denatured, renatured and dialyzed, and its activity was identified, The plates of ELISA were folded by the produced antigen. Then the ELISA method was constructed to determine the biological activity of the antigen RESULTS： The recombined engineering bacterium expressed M, 3 300 target protein in the form of in- clusion body. SDS-PAGE showed the purity of the optimized inclusion body was above 95%, The ac- cordant rate was 97.1% between ELISA and blot electrophoresis after the activity examination was performed in 126 healthy people. CONCLUSION： The biological activity of Hpylori vacA gene recombined expressive inclusion body can be significantly and stably improved after denaturation and renaturation. So the inclusion body can be used as a candidate for the production of H pylori vaccine.

关 键 词：幽门螺杆菌 VACA基因 包涵体 变性 复性 ELISA方法 VACA 重复性 基因重组表达 acA抗原 

分 类 号：R573[医药卫生—消化系统]