犬2型腺病毒纤突蛋白单克隆抗体的制备与鉴定  

Preparation and identification of monoclonal antibodies against FIBER protein of canine adenovirus-2

作　　者：王枞嵘 牛欣蕊 郭江涛 鲁维飞[1,2,3] 刘忠虎[1,2,3] 褚贝贝[1,2,3] 付朋飞 王江[1,2,3] WANG Congrong;NIU Xinrui;GUO Jiangtao;LU Weifei;LIU Zhonghu;CHU Beibei;FU Pengfei;WANG Jiang(College of Veterinary Medicine,Henan Agricultural University,Zhengzhou 450046,China;Key Laboratory of Animal Biochemistry and Nutrition,Ministry of Agriculture and Rural Affairs,Henan Agricultural University,Zhengzhou 450046,China;Key Laboratory of Animal Growth and Development of Henan,Henan Agricultural University,Zhengzhou 450046,China;College of Life Science and Engineering,Henan University of Urban Construction,Pingdingshan,Henan 467036,China)

机构地区：[1]河南农业大学动物医学院,河南郑州450046 [2]河南农业大学农业农村部动物生化与营养重点实验室,河南郑州450046 [3]河南农业大学河南省动物生长发育调控重点实验室,河南郑州450046 [4]河南城建学院生命科学与工程学院,河南平顶山467036

出　　处：《中国兽医学报》2023年第8期1635-1642,共8页Chinese Journal of Veterinary Science

基　　金：河南省自然科学基金资助项目(202300410213);河南省教学改革资助项目(2021SJGLX351)。

摘　　要：犬2型腺病毒(canine adenovirus-2,CAV-2)主要引起犬科动物的传染性呼吸道疾病和消化道疾病,且其传染性极强,全球范围内感染率非常高,但现有的临床防治措施难以完全阻断病毒的传播和快速特异性治疗患病动物。为了获得可用于CAV-2基础病毒学研究和临床诊治所需的单克隆抗体,本研究将浓缩纯化的CAV-HN45株灭活后免疫BALB/c小鼠,采用细胞融合技术将免疫小鼠的脾细胞与SP2/0细胞融合,经免疫过氧化物酶单层细胞试验和间接免疫荧光试验筛选出2株能分泌CAV FIBER蛋白抗体的单克隆杂交瘤细胞系:1-A12-C6、8-A12-B10,抗体亚类分别为IgG2a和IgG2b。腹水纯化后质量浓度为1 g/L抗体的间接免疫荧光效价为1-A12-C6:1∶8192;8-A12-B10:1∶2048,2株单克隆抗体均不能用于免疫印记试验检测。抗体具有体外中和病毒的活性,1-A12-C6和8-A12-B10抗体的CAV中和效价分别为1∶256和1∶64。本研究单克隆抗体的成功制备为建立快速、高效的CAV-2免疫学检测技术和CAV-2的防治提供了依据。Canine adenovirus-2(CAV-2)mainly causes infectious respiratory and digestive tract diseases in canids.It is highly infectious and has a very high infection rate worldwide.However,the existing clinical prevention and treatment measures are difficult to completely block the spread of the virus and quickly and specifically treat sick animals.In order to obtain monoclonal antibodies needed for basic virological research and clinical diagnosis and treatment of CAV-2,BALB/c mice were immunized with the concentrated and purified CAV-HN45 strain after inactivation.The spleen cells of the immunized mice were fused with SP2/0 cells by cell fusion technology.Two monoclonal hybridoma cell lines,1-A12-C6 and 8-A12-B10,were screened by immunoperoxidase monolayer assay(IPMA)and indirect immunofluorescence assay(IFA).The subclasses of antibodies were IgG2a and IgG2b,respectively.The titer of IFA was 1-A12-C6:1∶8192and 8-A12-B10:1∶2048at a concentration of 1g/L after purification of ascites.Both monoclonal antibodies could not be used for Western blot.The neutralization titers of 1-A12-C6and 8-A12-B10 antibodies were 1∶256 and 1∶64,respectively.The successful preparation of monoclonal antibodies in this study provides materials for the establishment of rapid and efficient immunological detection technology of CAV-2or the prevention and treatment of CAV-2.

关 键 词：CAV-2 Fiber蛋白 单克隆抗体 IGG2A IgG2b 

分 类 号：S852.65[农业科学—基础兽医学]