南京医科大学学报（自然科学版）2012，Vol.32Issue(3)：301-305,5.
人高尔基体蛋白GP73基因的克隆、表达、单克隆抗体的制备及鉴定
Cloning, expression and development of monoclonal antibody of human Golgi protein 73
摘要
Abstract
Objective;To clone,express human Golgi protein 73 (CP73),develop and characterize the monoclonal antibody(mAb) against GP73,a valuable serum marker for hepatocellular carcinoma(HCC). Methods: CP73 gene was amplified by PCR,and cloned into prokaryotic expression vector pComb3XSS. The recombinant construct was expressed in ToplOF' E. Coli. Host,and purified with their fusion partner by HisFF Trap affinity chromatography. The recombinant protein was used to immunize the BALB/c mice for mAb development. The titer of the mAb was detected by ELBA and its specificity was analyzed by Western blot. The sera level of GP73 in HCC patients and healthy people was measured by co-immunoprecipitation (IP). Results; The GP73-6xHis recombinant protein was successfully expressed and purified. Five hybridoma cell lines against GP73 were obtained. IP revealed that the mAb could combine with GP73 in human sera with high specificity. The level of GP73 in HCC patients is much higher than that of healthy people. Conclusion; The success in mouse a-GP73 mAb development provides the basis for further developing a sandwich ELISA for detection of human GP73.关键词
GP73/单克隆抗体/肝细胞性肝癌/肿瘤标志物Key words
GP73/ monoclonal antibody/ HCC/ tumor markers分类
医药卫生引用本文复制引用
徐言,曾晓燕,张晓,金秋,张建平,焦永军..人高尔基体蛋白GP73基因的克隆、表达、单克隆抗体的制备及鉴定[J].南京医科大学学报（自然科学版）,2012,32(3):301-305,5.基金项目
江苏省卫生厅科研项目(H200814) （H200814）
江苏省卫生厅医学重点人才课题(RC2011082) （RC2011082）
