猪圆环病毒4型Cap蛋白单克隆抗体的制备与鉴定  

Preparation and identification of monoclonal antibodies against Cap protein of porcine circovirus 4

作　　者：姚晓慧 连拯民 袁丽莉 胡丹鹤 刘盼娆 朱振邦 李向东 YAO Xiaohui;LIAN Zhengmin;YUAN Lili;HU Danhe;LIU Panrao;ZHU Zhengbang;LI Xiangdong(College of Veterinary Medicine,Yangzhou University,Yangzhou 225009,China;Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses,Yangzhou University,Yangzhou 225009,China;Joint International Research Laboratory of Agriculture and Agri-Product Safety,the Ministry of Education of China,Yangzhou University,Yangzhou 225009,China)

机构地区：[1]扬州大学兽医学院,江苏扬州225009 [2]江苏高校动物重要疫病与人兽共患病防控协同创新中心,江苏扬州225009 [3]扬州大学教育部农业与农产品安全国际合作联合实验室,江苏扬州225009

出　　处：《畜牧与兽医》2022年第4期67-71,共5页Animal Husbandry & Veterinary Medicine

基　　金：江苏现代农业(生猪)产业技术体系疾病防控创新团队(JATS[2021]358)。

摘　　要：为制备猪圆环病毒4型(PCV4)Cap蛋白单克隆抗体,以原核表达的重组Cap蛋白免疫6~8周龄小鼠,三次免疫后取免疫小鼠脾细胞与SP2/0骨髓瘤细胞进行融合。利用亚克隆技术和间接免疫荧光(IFA)方法筛选到2株阳性单克隆杂交瘤细胞株,分别命名为5H9和2F3。间接免疫荧光和Western blot试验表明,2株单抗均能特异性识别293T细胞中特异表达的Cap蛋白,且5H9和2F3的IFA效价分别为1∶12 800和1∶6 400。亚类鉴定结果表明,两株单抗重链均属于IgG1,轻链类型为kappa型。本研究制备的抗PCV4 Cap特异性单抗,为PCV4检测方法的建立和流行病学调查提供了有效的生物材料,也为该病毒的分离鉴定与Cap蛋白功能的探究奠定了基础。To prepare monoclonal antibodies against the Cap protein of porcine circovirus 4(PCV4), 6-8 week-old BALB/c mice were immunized with recombinant Cap proteins expressed in a prokaryotic system. After immunization for three times, spleen cells of the mice were obtained and were fused with SP2/0 cells. By subcloning and indirect immunofluorescence assay(IFA) screening, two positive monoclonal hybridoma cell lines were obtained and were designated as 5 H9 and 2 F3, respectively. Western blot and IFA showed that both monoclonal antibodies specifically recognized the Cap protein transiently expressed in 293 T cells, and the IFA titers of 5 H9 and 2 F3 were 1∶12 800 and 1∶6 400, respectively. Subtype identification results showed that the heavy chains of these two monoclonal antibodies belonged to IgG1, and the light chains were of the κ type. The monoclonal antibodies generated in this study not only provided important biological materials for the establishment of detection methods and for epidemiological investigation for PCV4, but also laid the foundation for virus isolation and functional study of Cap proteins.

关 键 词：猪圆环病毒4型 衣壳蛋白 原核表达 单克隆抗体 

分 类 号：S811.6[农业科学—畜牧学]