文献类型：期刊文献

中文题名：抗猪PD-L1单克隆抗体的制备与其生物学特性鉴定

英文题名：Preparation of Anti-porcine PD-L1 Monoclonal Antibody and Identification of Its Biological Characteristics

作者：岳锋[1];杨健[1];周娟娟[1];李静[2];朱艳平[1];李鹏[1];孙国鹏[1];张艳芳[1];王选年[1]

第一作者：岳锋

机构：[1]新乡学院、生命科学技术学院,新乡453003;[2]河南省畜牧兽医服务中心,郑州450008

第一机构：新乡学院生命科学技术学院

年份：2018

卷号：37

期号：6

起止页码：2368-2373

中文期刊名：基因组学与应用生物学

外文期刊名：Genomics and Applied Biology

收录：CSTPCD;;北大核心:【北大核心2017】；CSCD:【CSCD2017_2018】；

基金：国家自然科学基金面上项目(31672540);河南省高等学校重点科研项目(17A230019);新乡学院博士启动科研项目(1366020053)和新乡学院科技创新团队项目(XXUTD20170106)共同资助

语种：中文

中文关键词：PD-L1;胞外区;单克隆抗体;效价

外文关键词：PD-L1;Extracellular domain;Monoclonal antibody;Titer

摘要：本研究旨在制备抗猪PD-L1单克隆抗体（monoclonal antibody,m Ab）,有助于阻断猪PD-1/PD-L1通路逆转免疫功能。免疫原为猪PD-L1胞外区重组蛋白,雌性BALB/c鼠为免疫动物,用淋巴细胞杂交瘤技术将鼠骨髓瘤细胞NS0和免疫BALB/c鼠脾细胞融合,ELISA法筛选及多次克隆化培养,筛选抗猪PD-L1 mAb的杂交瘤细胞株。成功制备1株能稳定分泌抗猪PD-L1的mAb的杂交瘤细胞株3B5,Ig亚型为IgG1,细胞上清和腹水效价分别为1：1×2^10和1：1.024×10^5,ELISA和Western-blotting结果表明该株单抗能特异性识别猪重组PD-L1蛋白,流式细胞术结果表明该单抗可以与猪PBMC上PD-L1蛋白有效结合,成功制备了1株分泌抗猪PD-L1单克隆抗体的杂交瘤细胞株3B5,为检测猪PD-L1蛋白表达水平及猪PD-1通路在猪传染性疾病中的致病机制提供有力的检测工具。
The aim of this study was to prepare anti-porcine PD-L1 monoclonal antibody（m Ab）, and to help block the porcine PD-1/PD-L1 pathway and reverse the immune function. Female BALB/c mice were immunized with the extracellular domain recombinant protein of porcine PD-L1. Murine myeloma cells NS0 and spleen cells of immunized BALB/c mice were fused by lymphocyte hybridoma technique, and hybridoma cell strains of antiporcine PD-L1 mAb were screened by ELISA method for multiple cloning culture. A hybridoma cell line with stable secretion of anti-porcine PD-L1 mAb was successfully obtained, named 3 B5. The Ig subtype was IgG1. The titers of cell supernatant and ascites were 1： 1 ×2^10 and 1： 1.024 ×10^5, respectively. The results of ELISA and Western-blotting showed that the m Ab could specifically recognize porcine PD-L1 recombinant protein. Flow cytometry showed that the mAb could effectively bind to the PD-L1 protein on porcine peripheral blood mononuclear cell（PBMC）. The hybridoma cell line 3B5 secreting anti-porcine PD-L1 mAb was successfully prepared, which would provide a powerful detection tool for detecting the expression level of porcine PD-L1 protein and the pathogenic mechanism of porcine PD-1 pathway in swine infectious diseases.