血清Ⅲ型鸭甲型肝炎病毒VP0蛋白单克隆抗体的制备与鉴定  

Preparation and Characterization of a Monoclonal Antibody against Protein VP0 of Duck Hepatitis Virus TypeⅢ

作　　者：黄军生 宋翠萍[1] 孙英杰[1] 仇旭升[1] 孟春春[1] 谭磊[1] 廖瑛[1] 刘炜玮 王桂军[2] 丁铲[1] HUANG Junsheng;SONG Cuiping;SUN Yingjie;QIU Xusheng;MENG Chunchun;TAN Lei;LIAO Ying;LIU Weiwei;WANG Guijun;DING Chan(Shanghai Veterinary Research Institute,CAAS,Shanghai 200241,China;College of Animal Science and Technology,Anhui Agricultural University,Hefei 230036,China)

机构地区：[1]中国农业科学院上海兽医研究所,上海200241 [2]安徽农业大学动物科技学院,合肥230036

出　　处：《中国动物传染病学报》2022年第3期146-151,共6页Chinese Journal of Animal Infectious Diseases

基　　金：国家重点研发计划项目(2017YFD0500800)。

摘　　要：血清Ⅲ型鸭甲型肝炎病毒(DHAV-3)属于小RNA病毒科,主要引起雏鸭以肝脏肿胀、出血为主的急性、烈性传染病。DHAV-3有3个主要结构蛋白,其中VP0蛋白具有极佳的抗原性及宿主保护位点。为研制鸭肝炎病毒VP0蛋白的单克隆抗体,本研究将鸭肝炎病毒的VP0基因进行原核表达,用纯化后的重组蛋白作为抗原免疫BALB/c小鼠,通过细胞融合、IFA筛选以及亚克隆后,获得了1株能够稳定分泌抗VP0蛋白单克隆抗体的杂交瘤细胞3A5,亚类鉴定为IgG1,IFA检测腹水效价为1∶12800。Western blot、IFA结果表明,制备的单克隆抗体效价高,反应原性强。本研究为后续建研制血清Ⅲ型鸭甲型肝炎病毒的快检试剂盒,及建立病原及抗体检测方法奠定基础。Duck hepatitis A virus typeⅢ(DHAV-3)belongs to the family Picornavirus and can cause acute and severe infectious diseases of ducks,which is mainly characterized by liver swelling and bleeding.Among the structural proteins of DHAV-3,VP0 protein possesses excellent antigenicity and host protection site,which plays an important role in diagnosis.In order to prepare a mAb against DHAV-3 VP0 protein,the recombinant pCold-VP0 was induced with IPTG for protein expression and then BALB/c mice were immunized with the purified VP0.After cell fusion,IFA screening and three rounds of subcloning,a monoclonal cell line 3A5 was obtained for stably secreting anti-VP0 antibody.The mAb 3A5 was identified as a subtype IgG1 and the IFA titer of ascites was 1:12800.The high titer and good immunoreactivity of this mAb as examined in Western blot and IFA suggested that it might be useful for further research ofDHAV-3.

关 键 词：血清Ⅲ型鸭甲型肝炎病毒 VP0基因 原核表达 单克隆抗体 

分 类 号：S852.4[农业科学—基础兽医学]