人源抗寨卡病毒单克隆抗体Fab段基因的获得和表达  

Generation of human monoclonal antibody Fab fragment to Zika Virus by Phage Display Technology

作　　者：袁润余[1] 孙丽娜[4] 苏娟[1] 纪洵敏 梁均和 陈茂余 梁米芳[4] 柯昌文[1] Yuan Runyu;Sun Lina;Su Juan;Ji Xunmin;Liang Junhe;Chen Maoyu;Liang Mifang;Ke Changwen(Guangdong Provincial Center for Disease Control and Prevention,Guangzhou 511430,China;Guangdong Provincial Institute o f Biological Products and Materia Medica,Guangzhou 510440,China;Jiangmen City Center for Disease Control and Prevention,Jiangmen 529030,China;National Institute for Viral Disease Control and Prevention,Chinese Center for Disease Control and Prevention,Beijing 102206,China)

机构地区：[1]广东省疾病预防控制中心病原微生物检验所,广州511430 [2]广东省生物制品与药物研究所,广州510440 [3]江门市疾病预防控制中心,529030 [4]中国疾病预防控制中心病毒病预防控制所,北京102206

出　　处：《中华实验和临床病毒学杂志》2019年第6期626-631,共6页Chinese Journal of Experimental and Clinical Virology

基　　金：病毒性传染病病原谱和病毒基因变异变迁规律研究(2018ZX10713-002);国家重点研发计划(2016YFD0500303);广东省防控寨卡病毒病科技攻关专项(2016A020250001);科技部改革发展专项基金-zika病毒研究((2016)273);国家科技重大专项(2018ZX09739002-004);广东省医学科学基金(A2018187)。

摘　　要：目的 利用噬菌体表面展示技术进行人源抗寨卡病毒(Zika virus,ZIKV)噬菌体抗体库的构建及其基因工程单克隆抗体Fab段基因的获得和表达,掌握人源噬菌体抗体库的制备和针对特定病毒高度特异性的抗体筛选方法。方法 收集寨卡病毒病康复期患者的全血样本,分离淋巴细胞,采用RT-PCR方法成功地从淋巴细胞Ig mRNA中扩增出抗体轻链和重链Fab段基因,利用pComb3H系统成功构建具有基因多样性的人源抗ZIKV噬菌体抗体库制备。利用已纯化的ZIKV和已获得的ZIKV E蛋白抗原对已建立的抗体库进行富集筛选。结果 本研究成功地构建人源抗ZIKV噬菌体抗体库,并获得针对ZIKV E蛋白抗原的单克隆抗体Fab段基因,该基因能在大肠埃希菌中进行有效表达。结论 根据其序列分析研究表明,该单抗为一株新的抗ZIKV的人源基因工程抗体,这对建立ZIKV快速特异的早期诊断方法,获得特异性ZIKV人源治疗单抗奠定基础。Objective In this study,phage display technology was used to construct the human anti-Zika virus(ZIKV),phage antibody library and to obtain and express the monoclonal antibody.The aim was to master the preparation and expression of human phage antibody library screening method for highly specific antibodies.Methods The whole blood samples of Zika patients were collected and the lymphocytes were isolated.The RT-PCR method was used to amplify the antibody light chain and heavy chain Fab gene from lymphocyte Ig mRNA.The pComb3H system was used to construct the gene with genetic diversity Preparation of human anti-ZIKV phage antibody library.The purified antibody library was screened by using the purified ZIKV and the obtained ZIKV E protein antigen.Results The monoclonal antibody Fab fragment gene was successfully obtained for the ZIKV E protein antigen.The gene can be efficiently expressed in Escherichia coli.Conclusions According to the sequence analysis,this study showed that the monoclonal antibody was a new human genetically engineered antibody against ZIKV,which laid the foundation for the early diagnosis of ZIKV,and obtain a specific monoclonal antibody to ZIKV for human treatment of ZIKV infection.

关 键 词：人源基因工程抗体 寨卡病毒 噬菌体表面展示 

分 类 号：R373[医药卫生—病原生物学] Q78[医药卫生—基础医学]