利用优化DNA质粒制备大鼠CD138单克隆抗体  

Generation of Rat CD138 Antibody Using Optimized DNA Plasmid

作　　者：罗芳芳 钱峰[2] 梅芹 LUO Fangfang;QIAN Feng;MEI Qin(WuXi Biologics,Shanghai 200131;School of Life Sciences,Fudan University,Shanghai 200438)

机构地区：[1]上海药明生物技术有限公司,上海200131 [2]复旦大学生命科学学院,上海200438

出　　处：《生物化工》2020年第6期42-46,68,共6页Biological Chemical Engineering

摘　　要：目的:通过优化DNA质粒制备高同源性小鼠抗大鼠CD138抗体。方法:优化DNA质粒转录后调节元件和分子佐剂增强DNA免疫反应。通过将HPRE、WPRE、HTLV-1R转录后调节元件和鞭毛蛋白(FliC)佐剂分别克隆入pCAGGS,获得pCAGGSHPRE、pCAGGS-WPRE、pCAGGS-HTLV-1R和pCAGGS-FliC载体;将luciferase和大鼠CD138基因分别克隆入相应载体,进行体外表达检测;选用3个质粒(pCD138、pHCD138、pFCD138)分别免疫BALB/C小鼠,比较免疫反应强度和抗大鼠CD138抗体数量和质量。结果:与对照组(pCD138)相比,含FliC质粒显著地降低大鼠CD138表达(P<0.05);其他质粒无显著差异(P>0.05)。含FliC质粒明显提高小鼠免疫反应,其他质粒相对较弱;而且含FliC质粒组动物免疫反应较快,明显缩短免疫周期,获得高质量阳性杂交瘤单克隆抗体。结论:嵌合FliC和大鼠CD138明显提高抗原特异性免疫反应,最终获得两株高亲和力杂交瘤单克隆抗体(35-G11、39-D2)。Purpose:this study is to prepare highly homologous mouse anti-rat CD138 antibodies by optimizing DNA plasmids.Methods:include optimizing post-transcriptional regulatory elements and molecular adjuvants to enhance DNA immune response.First,by cloning HPRE,WPRE,HTLV-1R and flagellin FliC elements into pCAGGS,respectively,pCAGGS-HPRE,pCAGGS-WPRE,pCAGGS-HTLV-1R and pCAGGS-FliC vectors were obtained.Secondly,the luciferase and rat CD138 gene were cloned into corresponding vectors,and the expression was detected in vitro.Then,3 plasmids(pCD138,pHCD138,pFCD138)were used to immunize BALB/C mice respectively,and the quantity and quality of rat CD138 antibody produced by different plasmids were compared.Results:compared with the control group(pCD138 plasmid),the plasmid containing Flic significantly improved the immune response of mice,while other plasmids were relatively weak.In addition,the animals in the Flic plasmid group have a faster immune response,significantly shorten the immune cycle,and obtain more high-quality positive hybridoma cells.Disscussion:chimeric FliC and rat CD138 significantly improved the antigen-specific immune response,and finally obtained two high-affinity hybridoma monoclonal antibodies(35-G11,39-D2).

关 键 词：大鼠CD138 DNA免疫 鞭毛蛋白 pCAGGS 佐剂效应 

分 类 号：R392[医药卫生—免疫学]