抗CEA嵌合小分子抗体的构建与表达  

Construction and expression of a small molecular chimeric antibody against human carcinoembryonic antigen

作　　者：赵泽国[1] 冉宇靓[1] 孔健[2] 孙立新[1] 遇垅[1] 刘军[1] 杨治华[1] 

机构地区：[1]中国医学科学院、中国协和医科大学肿瘤研究所细胞生物及分子生物学室,北京100021 [2]卫生部北京生物制品研究所,北京100024

出　　处：《中国生物学文摘》2006年第1期18-21,共4页Chinese Biological Abstracts

基　　金：国家九五攻关计划资助项目（96-906-01-23）

摘　　要：目的构建表达抗CEA小分子抗体，以利于放射免疫诊断及导向治疗。方法构建CEA单链抗体基因，克隆入载体pKpL-3a，在大肠杆菌中包涵体表达，ELISA检测活性。将轻重链基因克隆入分泌型表达载体pSCMH，在大肠杆菌中表达．ELISA极测活性。将CEA嵌合抗体的轻链和Fd基因克隆入杆状病毒表达载体pAcUW51中，在sf9细胞中分泌表达，ELISA检测活性及测定产量．竞争抑制ELISA测定其识别的抗原表位。结果多种方法复性的包涵体及原核分泌表达的单链扎体束测到活性。昆虫细胞中表达的小分子抗体具抗体活性，产量为1．7μg／mL，竞争抑制实验显示其与亲本鼠单抗C50识别相同的表位。结论该CEA抗体基因在大肠杆菌中不能表达出有功能的分子，而在昆虫细胞中表达了具活性抗体分子；某些抗体基因只有在真核细胞中才能表达出有功能的抗体分子。Objective A small molecular antibody against human carcinoembryonic antigen was constructed and expressed in order to be used for radioimmunoimage and target therapy. Methods The gene of single chain antibody against CEA was cloned into expression vector pKpL-3a and expressed in E. coli pop2136. The antigen binding activity was analysed by ELISA, The light and heavy chain variable region genes were cloned into secretory expression vector pSCMH whose gene II had been excised, and expressed in E, coli XL1-Blue, The antibody activity was assayed by ELISA, The light chain and Fd gene of CEA chimeric antibody were cloned into baculovirus expression vector pAcUWS1 and expressed in sf9 cell. The activity, yield and affinity were analysed by ELISA. Results After being renatured by various methods, the specific antigen binding activity of expression product in E. coli pop2136 was not detected, nor was the secretory expression product in E. coli XL1-Blue. The small molecular chimeric antibody expressed in sf9 cell can specifically bind human CEA antigen and had the yield of 1, 7 μg/mL. Competition ELISA showed that antigen epitope recognized by the chimeric antibody was the same as the parent McAb C50. Conclusion A small molecular chimeric antibody against CEA was successfully expressed in sf9 cell. Only in eucaryotic ceils can some antibody genes express functional molecules.

关 键 词：癌胚抗原 小分子抗体 抗体工程 

分 类 号：R392.11[医药卫生—免疫学]