前列腺干细胞抗原及其单链抗体-碱性磷酸酶融合蛋白的表达与检测  

Expression and detection of prostate stem cell antigen and fusion protein comprising a PSCA-specific single-chain antibody and alkaline phosphatase

作　　者：尚国富 刘江丽 于欢 唐开义 曾柱[1,2,3] 胡祖权 王赟[1,2] Shang Guofu;Liu Jiangli;Yu Huan(Immune Cells and Antibody Engineering Research Center of Guizhou Province,Key Laboratory of Biology and Medical Engineering,School of Biology and Engineering,Guizhou Medical University,Guiyang 550025;School of Basic Medical Sciences,Guizhou Medical University,Guiyang 550025)

机构地区：[1]贵州医科大学生物与医学工程重点实验室/贵州省免疫细胞与抗体工程研究中心,贵阳550025 [2]贵州医科大学基础医学院/生物与工程学院,贵阳550025 [3]贵州医科大学环境污染与疾病监控省部共建教育部重点实验室,贵阳550025

出　　处：《安徽医科大学学报》2020年第12期1968-1971,共4页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:31660258);贵州省科学技术基金(编号:黔科合支撑[2019]2787号、黔科合基础[2018]1412、黔科合平台人才[2016]5676);国家级大学生创新创业训练项目(编号:201810660020)。

摘　　要：扩增前列腺干细胞抗原(PSCA)及其特异单链抗体PaFv的编码基因,分别克隆到pET-32a和pDAP2/S载体中,转化大肠杆菌感受态细胞。融合蛋白TrxA-PSCA和PaFv-AP诱导表达后,SDS-PAGE和Westernblot分析其表达情况,碱性磷酸酶(AP)显色反应和ELISA检测蛋白的活性。结果显示构建的原核表达体系成功实现TrxA-PSCA和PaFv-AP融合蛋白的可溶性表达,而且PaFv-AP具有与PSCA抗原结合的抗体活性以及AP活性。The PSCA gene encoding prostate stem cell antigen(PSCA) and PaFv gene encoding PSCA-specific scFv antibody were amplified and cloned into pET-32 a and pDAP2/S vectors, respectively. After induced for expression of TrxA-PSCA and PaFv-AP fusion proteins, SDS-PAGE and Western blot were conducted to analyze their expression levels and forms, whereas alkaline phosphatase(AP) color-reaction and enzyme-linked immunosorbent assay(ELISA) were applied to detect the activity of PaFv-AP fusion. The results suggested that TrxA-PSCA and PaFv-AP fusion proteins were successfully expressed in soluble form in the constructed prokaryotic expression systems. Moreover, the PaFv-AP fusion retained the antigen-binding capacity of antibody and AP activity.

关 键 词：前列腺干细胞抗原 单链抗体 碱性磷酸酶 原核表达 免疫学检测 

分 类 号：R392.11[医药卫生—免疫学] Q786[医药卫生—基础医学]