抗淋巴细胞激活基因3(LAG-3)全人源抗体的制备  

Preparation of anti-lymphocyte activation gene 3 (LAG-3) fully human antibodies

作　　者：陈秀秀 于晓杰 刘剑飞 吴成林 钦可为 周丽君 CHEN Xiuxiu;YU Xiaojie;LIU Jianfei;WU Chenglin;QIN Kewei;ZHOU Lijun(Naval Clinical College,Anhui Medical University,Hefei 230032;Central Laboratory,Sixth Medical Center,Chinese People’s Liberation Army General Hospital,Beijing 100048,China)

机构地区：[1]安徽医科大学海军临床学院,安徽合肥230032 [2]中国人民解放军总医院第六医学中心中心实验室,北京100048

出　　处：《细胞与分子免疫学杂志》2020年第5期451-456,共6页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金(31470897);中国人民解放军总医院第六医学中心创新培育基金(CXPY201817)。

摘　　要：目的淋巴细胞激活基因3(LAG-3)是一个极具潜力的肿瘤治疗靶点。本研究旨在从大容量全人源噬菌体抗体库中筛选抗LAG-3的单链抗体(scFv),将其转化为完整的全人源抗体。方法从全人源噬菌体抗体库,筛选抗LAG-3全人源噬菌体scFv,ELISA鉴定噬菌体scFv的特异性,采用表面等离子共振技术(SPR)测定其亲和力,再通过真核表达系统获得抗LAG-3全人源完整抗体。结果通过3轮筛选富集,获得了4个具有特异结合活性的scFv。亲和力测定最高的达到3.48×10-10。再通过构建完整的抗LAG-3全人源抗体表达载体,经真核细胞表达、纯化后,获得了3株特异性抗LAG-3全人源抗体。结论利用大容量全人源噬菌体抗体库,成功筛选并表达出3种高亲和力和特异性的抗LAG-3全人源抗体。Objective Lymphocyte activation gene 3(LAG-3)is a potential therapeutic target of tumor.This work aims to screen for anti-LAG-3 human scFvs from a human phage antibody library,and transform them into fully human antibodies.Methods The human phage antibody library was screened for anti-LAG-3 human scFvs.The specificity and affinity of phage scFvs were determined by ELISA and surface plasmon resonance(SPR),respectively.The anti-LAG-3 full human antibodies were obtained using eukaryotic expression system.Results Through 3 rounds of screening,we obtained 4 phage scFv clones that could bind to human LAG-3 recombinant protein.Affinity determination showed that the highest affinity reached 3.48×10-10.Then through the construction of whole human antibody expression vector,expression in eukaryotic cells,and purification,3 clones of anti-LAG-3 full human antibodies were harvested finally.Conclusion Using a large-capacity human antibody phage display library,we obtained three anti-LAG-3 full human antibodies with high affinity.

关 键 词：噬菌体展示技术 淋巴细胞激活基因3(LAG-3) 人源抗体 

分 类 号：R730.51[医药卫生—肿瘤] R392.1[医药卫生—临床医学]