幽门螺杆菌脂蛋白Lpp20的表达纯化及其临床应用的初步研究  

Expression and purification of Lpp20 from Helicobacter pylori and primary study of its clinical application

作　　者：靖吉芳[1] 张艳[2] 

机构地区：[1]韶关学院医学院,广东韶关512026 [2]南华大学病原生物研究所,湖南衡阳421001

出　　处：《热带医学杂志》2013年第5期631-633,667,共4页Journal of Tropical Medicine

基　　金：湖南省自然科学基金优秀面上项目(06JJ2093);湖南省教育厅优秀青年项目(08B067)

摘　　要：目的获得重组幽门螺杆菌(H.pylori)脂蛋白(rLpp20),并研究其在血清学诊断中的应用价值。方法应用重组质粒pGEX-6P-2/Lpp20在大肠杆菌(E.coli)BL21中表达,并进行GST亲和层析纯化,用Western-blot检测纯化产物(rLpp20)的免疫反应性;以rLpp20为包被抗原建立ELISA法对200份患者血清标本进行特异性IgG抗体检测,并与幽门螺杆菌-IgG抗体检测试剂盒法进行比较。结果成功表达的rLpp20(43000Mr),纯化产物可被幽门螺杆菌多克隆抗体及阳性血清识别;成功建立了以rLpp20为包被抗原的间接ELISA法,其灵敏度为91.0%,特异度为100%。与试剂盒法比较,两者的符合率为95.5%。结论 Lpp20具有良好的免疫反应性,能与幽门螺杆菌阳性血清特异性结合,有望应用于幽门螺杆菌血清学诊断。Objective To obtain the recombinant lipoprotein20 （rLpp20） of Helicobacter pylori （H.pylori） and analyse its application value in serodiagnosis to Helicobacter pylori infection. Methods rLpp20 was expressed in Escherichia coli （E.coli） BL21, and purified by GST affinity chromatograph. The immunoreactivity of rLpp20 was analyzed by Western- blot.ELISA were used for the quantitative detection of the Lpp20 specific IgG antibody in the sera of two hundred patients, and compared with the detection results using H.pylori specific total IgG detection kit. Results A soluble recombinant fusion protein with molecular weight about 43 000 was obtsined after expression and purification. The result of Western-blot showed that rLpp20 could be recognized by the polyelonal anti-H.pylori antibody and H.pylori positive sera. Indirect ELISA was successfully developed to detect the anti-H.pylori antibodies in human sera, with the sensitivity of 91.0% when compared with the results of the kit test, and with the specificity of 100%. The concordance of results between the indirect ELISA test and the kit test was 95.5%. Conelution The expressed recombinant protein showed better immunoreactivity and could specifically react with H.pylori positive sera. This recombinant Lpp20 can be used for development of quick diagnostic kit for H.pylori detection.

关 键 词：幽门螺杆菌 重组蛋白 LPP20 血清学诊断 

分 类 号：R378.2[医药卫生—病原生物学]