丙型肝炎病毒E_2糖蛋白在哺乳动物细胞中的表达、纯化及初步应用  

Expression of glycoprotein hepatitis C virus in mammalian cell and application of Purified proton for detection of antibody against E2 in hepatitis C patients

作　　者：朱朝敏[1] 吴朝栋[2] 陶其敏[2] 冯百芳[2] 常锦红[2] 

机构地区：[1]重庆医科大学儿童医院 [2]北京医科大学人民医院肝病研究所,北京100044

出　　处：《中华肝脏病杂志》1999年第4期214-216,共3页Chinese Journal of Hepatology

基　　金：CMB及国家九五攻关项目基金

摘　　要：目的在哺乳动物细胞中表达丙型肝炎病毒（ＨＣＶ）Ｅ２糖蛋白，并对其进行纯化，研究该蛋白作为抗原检测丙型肝炎患者血清中抗Ｅ２抗体的可行性。方法将中国株ＨＣＶＥ２／ＮＳＩ基因片段克隆到含有６个连续组氨酸的真核表达载体上，转染哺乳动物细胞，表达Ｅ２糖蛋白，经金属和亲和层析（ＩＭＡＣ）纯化后用于ＥＩＡ检测丙型肝炎患者血清中抗Ｅ２抗体。结果哺乳动物细胞中表达的Ｅ２糖蛋白为７．０×１０４，ＩＭＡＣ纯化后纯度达９０．２％。３５份ＨＣＶＲＮＡ阳性标本中，２９份抗Ｅ２抗体阳性（８２．９％）。结论成功利用哺乳动物细胞中表达的Ｅ２糖蛋白建立ＥＩＡ检测抗Ｅ２抗体，并证实Ｅ２糖蛋白有较好的抗原性及敏感性，可用于建立新一代抗ＨＣＶ检测试剂。Objective E2 glycoprotein of hepatitis C virus was expressed in mammalian cell andpurified for detection of antibody against E2 in hepatitis C Patieme. Methods E2/NS1 gene derived fromHCV was inserted into expression vector containing six His tag. The recombinant plasmid was transfectedinto mammalian cells to express E2 glycoprotein expression. E2 glycoprotein was purified by affinitychromophotography. The purified protein was used to establish EIA method for detection of antibodiesagainst E2 in hepatitis C patients. Result Expressed E2 glycoprotein was 7.0 x 104. Purification of thepurified E2 protein was 90.2%. Twenty-nine patients were anti-E2 antibody positive(82.9%). CoIt was the first time to establish EIA method for detection of anti-E2 antibody by purified E2 glyCoprotein in China. E2 glycoprotein expressed in mammalian cells had good immunogrnity and could increasethe sensibility of anti-HCV detection. It snarests that E2 glycoprotein may be useful for development ofnew anti-HCV reagents.

关 键 词：丙型肝炎病毒 糖蛋白 血清学诊断 蛋白表达 

分 类 号：R373.21[医药卫生—病原生物学] R512.630.4[医药卫生—基础医学]