抗人结缔组织因子人源单链抗体的筛选、表达及活性鉴定  

Screening,expression and activity identification of anti-CTGF human single-chain Fv

作　　者：周云[1] 包林[2] 金秋[2] 张黎[2] 蔡雪婷[3] 卢悟广[3] 曹鹏[3] 焦永军[2] 张建平[1] 

机构地区：[1]南京医科大学第二附属医院,南京210011 [2]卫生部肠道病原微生物重点实验室,江苏省疾病预防控制中心病原微生物研究所 [3]江苏省中医药研究院细胞与分子生物学实验室

出　　处：《山东医药》2013年第20期15-17,21,I0002,共5页Shandong Medical Journal

基　　金：江苏省卫生厅科研项目(H200814);江苏省医学重点人才课题(RC2011082)

摘　　要：目的从人源噬菌体抗体文库中筛选抗人结缔组织生长因子(CTGF)单链抗体(ScFv),原核表达并鉴定其活性。方法以重组人源CTGF(rhCTGF)为靶标,对人源噬菌体抗体文库进行筛选;用Phage-ELISA筛选阳性克隆并进行原核表达、纯化,获得了高纯度的ScFv蛋白,通过ELISA和Millicell迁移实验鉴定其活性。结果经过3轮筛选,获得1株特异性抗人CTGF ScFv 1C5,ELISA显示1C5能与rhCTGF特异性结合;在Millicell迁移实验中,1C5能抑制CTGF的生物学功能,具有中和活性。结论成功筛选到1株抗人CTGF的ScFv,有望在CTGF相关疾病的治疗上发挥作用。Objective To screen anti-connective tissue growth factor （CTGF） human single-chain variable fragment （ScFv） from human phage antibody library, prokaryotic expression and to identify its activity. Methods The human phage antibody library was screened by using rhCTGF as the target, and the positive clones were selected by phage-ELISA to induce its prokaryotic expression and to purify the protein, and the high purity ScFv protein was obtained. Its activity was identified by ELISA and Millicell migration assay, respectively. Results After three rounds of screening, only 1 clone, designated 1 C5, with good reactivity to rhCTGF was selected. 1 C5 was found to bind human CTGF specifically in ELISA, and inhibit CTGF＇s function in Millicell migration assay, indicating its neutralizing activity. Conclusion An anti-CTGF human single- chain Fv clone was successfully identified, which could be used as therapeutic agent candidate for CTGF-related disease.

关 键 词：结缔组织生长因子 噬菌体展示技术 单链抗体 靶向治疗 

分 类 号：R392[医药卫生—免疫学]