猫IL-1β阻断单克隆抗体的筛选与猫源化改造  

Felinized development of blocking McAbs against cat IL-1β

作　　者：王定一 王阅 张晨曦 李秋燕 周登元 司有辉 曹胜波[1] 叶静[1] WANG Dingyi;WANG Yue;ZHANG Chenxi;LI Qiuyan;ZHOU Dengyuan;SI Youhui;CAO Shengbo;YE Jing(College of Veterinary Medicine,Huazhong Agricultural University,Wuhan 430070,China)

机构地区：[1]华中农业大学动物医学院,湖北武汉430070

出　　处：《中国兽医科学》2024年第3期364-372,共9页Chinese Veterinary Science

基　　金：湖北省自然科学基金项目(2019CFA010)。

摘　　要：为给猫的炎症性疾病治疗提供具有与猫白细胞介素-1β(IL-1β)高亲和力的单克隆抗体,将表达后纯化的重组猫IL-1β蛋白接种小鼠后制备单克隆抗体,经验证单抗对猫IL-1β蛋白的阻断效果并分析其靶向位点后,将小鼠单克隆抗体的轻/重链可变区基因片段连接到表达猫抗体的真核质粒中,转染真核细胞后检测猫源化改造抗体效果。结果表明,成功扩增了猫IL-1β基因,并成功构建了原核表达质粒pET-30a-IL-1β并表达猫IL-1β重组蛋白;在使用猫IL-1β重组蛋白免疫小鼠以及细胞融合、杂交瘤细胞筛选后,成功得到2株分别能够与猫IL-1β重组蛋白第二、第三结构域反应的小鼠源单克隆抗体C4C、H10E;将单克隆抗体C4C、H10E猫源化基因改造后的重组抗体仍具有阻断效果。本试验研究结果为开发针对猫IL-1β的阻断单克隆抗体药物奠定了基础,也为猫炎症性疾病的治疗提供了一种新的方案。In order to develop monoclonal antibody(Mc Ab) with binding cat interleukin-1β(IL-1β) to block cat inflammatory disease as a new drug product,the blocking mouse Mc Ab to cat IL-1β protein was detected,and the action site of the antibody was analyzed. Then the light/heavy chain variable region genes of the blocking Mc Ab were amplified and linked to the constant region of cat antibody gene to construct eukaryotic plasmid transfection to detect the effect of cat source modified antibody.The results showed that the cat IL-1 β gene was successfully amplified,and the prokaryotic expression plasmid p ET-30a-IL-1 β was successfully constructed,and the recombinant cat IL-1β protein was expressed.After immunizing mice with recombinant cat IL-1β protein, cell fusion and hybridoma cell screening, two murine monoclonal antibodies C4C and H10E which can react with the second and third domain of cat IL-1β recombinant protein were successfully obtained.The recombinant antibody genetically modified by monoclonal antibodies C4C and H10E still had blocking effect.The results of this study laid a foundation for the development of blocking monoclonal antibody drugs against IL-1β in cats,and also provided a new scheme for the treatment of inflammatory diseases in cats.

关 键 词：白介素1Β 单克隆抗体 阻断效果 猫源化抗体改造 

分 类 号：S852.42[农业科学—基础兽医学]