抗癌胚抗原嵌合重链抗体与核心链霉亲和素融合基因在昆虫细胞中的表达  

Expression of the Fusion Gene Encoding Murine-Human Chimeric Heavy Chain to Carcinoembryonic Antigen with Core-streptavidin Gene in Insect Cells

作　　者：杨卫东[1] 李彪[2] 朱承谟[1] 杨冠珍[2] 吴祥甫[2] 

机构地区：[1]上海第二医科大学附属瑞金医院,上海200025 [2]中国科学院上海生物化学研究所,上海200031

出　　处：《上海免疫学杂志》2001年第1期11-13,24,共4页Shanghai Journal of Immunology

基　　金：国家自然科学基金!资助项目 (39770 2 31);上海市启明星计划资助!项目 (98Q14 0 11)

摘　　要：将抗癌胚抗原嵌合重链抗体与核心链霉亲和素融合基因插入昆虫杆状病毒供体质粒pFastBacHTa中 ,经大肠杆菌DH10Bac体内转座 ,产生重组杆状病毒pBacHTa VH Cr3 CS。将其转染粉纹夜蛾Tn 5B1 4细胞 ,经扩增后在细胞内进行表达。SDS PAGE分析结果表明 ,在粉纹夜蛾Tn 5B1 4细胞中都表达产生一条特异性蛋白质 ,其相对分子质量约为 75 0 0 0左右 ,以Westernblot分析表明 ,该特异条带即为VH Cr3 CS蛋白。SDS PAGE和Westernblot分析结果表明 ,以HRP标记的生物素作为抗体进行蛋白质印迹在相对分子质量 75 0 0 0处可见表达条带 ,表明融合蛋白能特异性的与生物素结合 ,RIA表明重组杆状病毒表达产生的VH Cr3 CS蛋白能与CEA有较高的结合力。The fusion gene encoding murine human chimeric heavy chain to carcinoembryonic antigen(CEA) with core streptavidin gene (V H C r3 CS) was inserted into the donor plasmids pFastBacHTa Then the recombinant plasmids were transformed to the E coli DH10Bac,in which transposition performed The recombinant bacmid DNAs,which contained the V H C r3 CS gene,were extracted from the E coli DH10Bac,and were transfected the Tn 5B1 4 cells The recombinant baculoviruses were obtained After amplified,the recombinant baculoviruses infected the Tn cells and the fusion gene of V H C r3 CS were highly expressed in Tn cells SDS PAGE analysis revealed that the molecular weight of V H C r3 CS were 75 000 approximately It was shown to possess the ability to bind to its specific antigen of CEA by RIA Using horseradish peroxidase (HRP) labeled biotin as antibody in Western blots,one band of 75 000 only was detected, demonstrating that the fusion protein not only had the ability to bind CEA,but also could bind biotin specifically

关 键 词：癌胚抗原 嵌合重链抗体 链霉亲和素 昆虫杆状病毒 表达 

分 类 号：R730.3[医药卫生—肿瘤]