抗犬细小病毒单克隆抗体的制备与鉴定  

Preparation and identification of monoclonal antibodies against canine parvovirus

作　　者：杨凯越 宋彩玲 李彤彤 王文静 陈慧宇 赵爽爽 刘明[1] YANG Kaiyue;SONG Cailing;LI Tongtong;WANG Wenjing;CHEN Huiyu;ZHAO Shuangshuang;LIU Ming(State Key Laboratory of Veterinary Biotechnology/Harbin Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Harbin 150069,China)

机构地区：[1]中国农业科学院哈尔滨兽医研究所/兽医生物技术国家重点实验室,黑龙江哈尔滨150069

出　　处：《畜牧与兽医》2020年第10期54-58,共5页Animal Husbandry & Veterinary Medicine

基　　金：国家重点研发计划(2016YFD0501001);动物基因工程疫苗国家重点实验室开放课题(AGVSKL-ZY-201804)。

摘　　要：为制备针对犬细小病毒(CPV)的单克隆抗体,以纯化的CPV免疫BALB/c小鼠,3次免疫后,取其脾细胞与SP2/0细胞进行细胞融合。利用超速离心纯化的病毒作为包被抗原,采用间接ELISA方法筛选阳性杂交瘤细胞,获得3株能稳定分泌单克隆抗体的杂交瘤细胞株,分别命名为1D9、3G9和4F6。单克隆抗体细胞培养上清效价为1∶64、1∶64、1∶128,腹水效价为1∶10^4、1∶10^4、1∶10^5。3株杂交瘤细胞的染色体数分别为92、94、98。亚类鉴定结果显示1D9和3G9重链为IgG1,4F6重链为IgG2b,轻链均为kappa链。病毒中和试验表明,单克隆抗体4F6对CPV具有明显的病毒中和活性。交叉试验表明,3株单克隆抗体均可与CPV特异性结合,与其他犬类常见病毒无交叉反应。本研究制备的单克隆抗体为CPV的检测及其感染动物的治疗提供了物质基础。In order to prepare monoclonal antibodies against canine parvovirus(CPV),BALB/c mice were immunized with purified canine parvovirus.After three immunizations,spleen cells were fused with SP2/0 cells for cell fusion.The purified virus was used as a coating antigen,and an indirect ELISA method was established to screen positive hybridoma cells.Three hybridoma cell lines capable of stably secreting monoclonal antibodies were obtained,and were named 1 D9,3 G9,and 4 F6,respectively.The antibody titers in the supernatants of the three hybridoma cells were 1:64,1:64,and 1:128;and the titers in the ascites were 1:10^4,1:10^4,and 1:10^5.The chromosome numbers of the three hybridoma cells were 92,94,and 98,respectively.Subclass identification results showed that the heavy chain of 1 D9 and 3 G9 was IgG1,and that of 4 F6 was IgG2 b,and their light chains were both the kappa chain.Virus neutralization experiments showed that this study successfully screened 4 F6 with neutralizing activity against CPV.Cross-over experiments showed that all the three monoclonal antibodies were able to specifically bind to CPV but did not cross-react with other common viruses in dogs.The monoclonal antibodies prepared in this study were the basis for diagnostic detection of CPV and therapeutic application to CPV infected animals.

关 键 词：犬细小病毒 单克隆抗体 中和活性 

分 类 号：S855.3[农业科学—临床兽医学]