全人源抗结肠癌噬菌体单链抗体库的构建及筛选鉴定  

Construction and Screening of Human Anti-colon Carcinoma Single-chain Fv Fusion Phage Libraries

作　　者：谢平丽[1] 李官成[1] 李艳东[1] 周国华[1] 李跃辉[1] 郭锋杰[1] 王甲甲[1] 

机构地区：[1]中南大学肿瘤研究所,中国湖南长沙410078

出　　处：《生命科学研究》2010年第1期57-61,共5页Life Science Research

摘　　要：采用体外致敏法和EBV转化技术联合噬菌体展示技术构建噬菌体呈现型单链抗体库.从中筛选获得阳性克隆,并进行ELISA、免疫组化及测序鉴定.结果得到了库容为4.0×109的初级噬菌体抗体库,全长ScFv(Single-chain Fv)基因的插入率为90%.筛选获得两个克隆对HT-29呈强阳性反应,而与HFF等人正常细胞系呈弱阳性或阴性反应.免疫组织化学鉴定表明克隆F12与结肠癌组织和结肠癌旁组织阳性率的差别有统计学意义.由上述结果可见构建库容量达4.0×109的全人源抗结肠癌噬菌体单链抗体库是完全可行的,经筛选及鉴定获得了特异性较强的噬菌体克隆,为结肠癌的临床导向诊断和治疗奠定了基础.A fully human Single-chain Fv （ScFv） phage displaying library was constructed by using phage antibody library technique combination with in vitro sensitization and EBV transformation. Positive clones were obtained and identified by ELISA, immunohistochemistry, and sequencing. The result showed that a phage antibody library containing 4.0×10^9 TU （transduced unit） was obtained, and the percentage of full- length ScFv genes inserted into phage DNA was 90%. After testing by ELISA, 2 phage antibody clones reacting more strongly to HT-29 than to HFF were obtained. F12 reacted specifically to colon carcinoma cells in most human colon carcinoma tissue sections but in few human para-colon carcinoma tissue sections. The positive rate has statistical significance. It can be concluded that it is feasible to construct a fully human anti-colon carcinoma ScFv fusion phage library containing 4.0 x 109 TU. By cell ELISA and immuno- histochemistry with tissue sections, the clone F12 was confirmed to specifically bind with colon carcinoma cells. The ScFv fragment against colon carcinoma may be further developed and applied to clinical targeted diagnosis and therapy.

关 键 词：EB病毒转化 噬菌体抗体库 结肠癌 单链抗体 

分 类 号：R735.35[医药卫生—肿瘤]