犬PD-1单克隆抗体的筛选与鉴定
Development and Characterization of Mouse Anti-canine PD-1 Monoclonal Antibodies
王卓 1徐司雨 2夏兴霞 2李志敏 2毕振威 2钱晶 2莫菲 2诸玉梅 2王永山 2孙树民 3谭业平2
作者信息
- 1. 内蒙古民族大学,内蒙古通辽 028000;江苏省农业科学院兽医研究所,农业农村部兽用生物制品工程技术重点实验室,国家兽用生物制品工程技术研究中心,南京 210014
- 2. 江苏省农业科学院兽医研究所,农业农村部兽用生物制品工程技术重点实验室,国家兽用生物制品工程技术研究中心,南京 210014
- 3. 内蒙古民族大学,内蒙古通辽 028000
- 折叠
摘要
为筛选犬程序性死亡受体1(cPD-1)单克隆抗体,以原核表达并纯化复性的cPD-1 胞外区蛋白为靶抗原免疫BALB/c小鼠,应用单克隆抗体杂交瘤细胞技术和间接ELISA方法,经3 轮轮筛选和克隆化,获得1 株能稳定分泌抗cPD-1 单克隆抗体的杂交瘤细胞株1F9.间接ELISA检测显示1F9 杂交瘤细胞株连续传代培养能稳定分泌单克隆抗体,效价达到1∶2048,亚型鉴定重链为IgG1,轻链为κ;以HEK293 细胞表达cPD-1 胞外区蛋白为检测抗原,Western-blot和间接免疫荧光试验进行鉴定,结果显示1F9 单克隆抗体能特异性结合cPD-1 胞外区蛋白.本研究通过小鼠杂交瘤技术成功筛选到1 株鼠源cPD-1 单克隆抗体.
Abstract
In order to screen the monoclonal antibody against canine programmed death receptor 1(cPD-1),BALB/c mice were immunized with prokaryotically expressed and purified recombinant cPD-1 extracellular region protein as the target antigen,and a hybridoma cell line 1F9,which could stably secrete anti-cPD-1 monoclonal antibody,was obtained after 3 rounds of screening and clonation by applying monoclonal antibody hybridoma cell technology and indirect ELISA method.Indirect ELISA showed that the 1F9 hybridoma cell line was able to stably secrete monoclonal antibodies with a potency of 1∶2048 in continuous passaging culture,and the heavy chain was identified as IgG1 and the light chain as κ by subtype identification;the cPD-1 extracellular region protein expressed by HEK293 cells was used as the detection antigen,and Western-blot and indirect immunofluorescence assays were used for identification;the results showed that 1F9 monoclonal antibody could specifically bind to cPD-1 extracellular region protein.In this study,a mouse-derived cPD-1 monoclonal antibody was successfully screened by mouse hybridoma technique.
关键词
犬/PD-1/单克隆抗体/肿瘤Key words
cannie/PD-1/monoclonal antibody/cancer引用本文复制引用
出版年
2024
国家科技期刊平台
NSTL
万方数据