利用重组系统构建大容量噬菌体抗体库  

Construction of Large Nave Phage Display Antibody Library Using Recombinant System

作　　者：关武祥[1] 孙丽娜[1] 刘峰[1] 李川[1] 王琰[2] 李德新[1] 梁米芳[1] 

机构地区：[1]中国疾病预防控制中心病毒病预防控制所传染病预防控制国家重点实验室,北京100052 [2]解放军海军总医院,北京100037

出　　处：《病毒学报》2006年第3期203-208,共6页Chinese Journal of Virology

基　　金：国家自然科学基金项目(30371269)

摘　　要：噬菌体抗体库技术是获得人源抗体的重要方法,此文探讨大容量天然噬菌体抗体库在筛选人源抗体中的实际应用价值。从正常人外周血分离淋巴细胞,通过基因工程方法获得抗体基因并且构建到含有重组位点的载体pDF中获得初级噬菌体抗体库,初级库在重组系统中重组之后获得了容量为9×1010的天然Fab噬菌体抗体库,通过序列测定和重组蛋白筛选来对抗体库的质量进行初步鉴定。对随机挑取的96个克隆测序序列分析表明各种型别比例基本合适;用四种抗原筛选有明显的富集,并且获得了针对其中一种抗原的阳性抗体克隆。Phage display antibody library especially large naive antibody library is a reliable technology for the se lection of antibody from human. Diverse antibody genes were amplified from lymphocytes collected from normal adult peripheral blood, the antibody genes were inserted into the vector which contains Cre-Loxp recombinant sites to obtain a primary phage display library . A large naive Fab phage display library containing 9 ×10^10 clones was constructed after the recombination of primary library completed in the recombinant system. The quality of the library was tested by sequence analysis and selection against several recombinant proteins. Sequence analysis of the randomly picked 96 Ig clones showed the library contains various antibody gene types. Panning of four recombinant proteins was successful and specific clones against one antigen were obtained.

关 键 词：噬菌体抗体库 Cre-Loxp重组 FAB抗体 

分 类 号：R373.32[医药卫生—病原生物学] Q78[医药卫生—基础医学]