抗肝癌细胞单链抗体的筛选和鉴定  

Screening and Identifi cation of scFv Against Hepatoma Carcinoma Cell

作　　者：吴晓军[1,2] 张娟[2] 陈卫[2] 栾立标[1] 王旻[2] 

机构地区：[1]中国药科大学药剂学教研室 [2]中国药科大学分子生物学教研室,江苏南京210009

出　　处：《中国医药工业杂志》2009年第6期412-416,共5页Chinese Journal of Pharmaceuticals

基　　金：国家自然科学基金资助项目(30672547)

摘　　要：以人正常肝细胞系HL-7702为阴性筛选细胞,肝癌细胞系HepG-2为阳性筛选细胞,从全人源噬菌体抗体库中筛选与HepG-2细胞系特异性结合的单链抗体。通过PCR及ELISA鉴定,将阳性克隆转入pET22b载体中构建表达质粒。所得的基因工程菌经测序,以IPTG诱导可溶性单链抗体表达,并通过SDS-PAGE、Westernblot和ELISA检测,最终得到能与HepG-2特异性结合的单链抗体。A humanized scFv phage library was used for the bio-panning of the genes coding for specific antibodies, with hepatocellular carcinomas cell line HepG-2 as positive selecting cells and a liver cell line HL-7702 as negative selecting cells. After four rounds of bio-panning, PCR and ELISA were carried out for the identification of the positive clones which could bind to HepG-2. The gene from positive clones was inserted into the expression vector pET22b. After sequencing, the expression of the positive scFv was induced by IPTG and the antibody was confirmed by SDS-PAGE, Western blot and ELISA. The scFv which could bind to HepG-2 specifically was obtained.

关 键 词：噬菌体展示 单链抗体 肝癌细胞 可溶性表达 

分 类 号：Q786[生物学—分子生物学]