抗HBsAg人源单链可变区抗体的筛选与可溶性抗体的表达  

Identification and Expression of Human ScFv Against Surface Antigen of Hepatitis B Virus in E.coli

作　　者：钟彦伟[1] 成军[1] 施双双[1] 夏小兵[1] 李克[1] 董菁[1] 刘妍[1] 杨继珍[1] 

机构地区：[1]解放军三○二医院传染病研究所基因治疗研究中心,北京100039

出　　处：《中国病毒学》2001年第2期105-108,共4页Virologica Sinica

基　　金：国家自然科学基金!资助 (3990 0 130 )

摘　　要：采用噬菌体表面展示技术 ,以从乙型肝炎病毒 (HBV)表面抗原 (HBsAg)阳性血清超速离心纯化的HBsAg为固相抗原 ,从噬菌体单链可变区半合成抗体库中经过 5轮“吸附 洗脱 扩增”筛选过程 ,获得特异性较强的HBsAg人源单链可变区抗体 (ScFv)克隆并提取质粒 ,经SfiⅠ /NotⅠ酶切鉴定后 ,亚克隆到 pCANTAB5E表达载体中 ,转化大肠杆菌XL1 Blue。经IPTG诱导后 ,表达的可溶性HBsAg特异性ScFv以 5 0 %硫酸胺沉淀 ,经SDS PAGE电泳表明 ,XL1 Blue中表达的HBsAg可溶性ScFv的分子量约 2 8kD。免疫活性检测结果表明 ,该单链抗体具有较强的抗原结合活性和特异性。HBsAg人源单链抗体的筛选和表达成功 ,为今后HBsAg人源抗体的研究和应用奠定了基础。Using HBsAg antigen purified from HBsAg positive serum as the coating antigen, the single chain variable fragment (ScFv) antibody to HBsAg has been screened and identified from a semi synthetic phage library by phage display technique. The SfiI/NotI ScFv DNA fragment was harvested from pHEN1 ScFv and inserted into pCANTAB5E, and the recombinant plasmid DNA was used to transform competent host XL1 Blue. After induction with IPTG for 20 hrs, the expressed HBsAg ScFv in the supernantant of XL1 Blue was precipitated with 50% ammonium sulfated and demonstrated the molecular weight is about 28kDa by SDS PAGE. The reactivity and specificity have been confirmed by enzyme linked immuno sorbent assay (ELISA). The identification and expression of HBsAg ScFv from E.coli were successfully achieved.

关 键 词：HBSAG 噬菌体 单链抗体 筛选 表达 HBV 

分 类 号：R446.6[医药卫生—诊断学]