抗犬瘟热病毒核蛋白单克隆抗体的制备及其线性抗原表位鉴定  

Preparation and identification of linear epitope of monoclonal antibodies against Canine distemper virus N protein

作　　者：曹智刚[1] 易立[1] 程悦宁[1] 仝明薇[1] 李爽[1] 王建科[1] 林鹏[1] 孙亚茹[1] 程世鹏[1] 

机构地区：[1]中国农业科学院特产研究所特种经济动物分子生物学国家重点实验室,吉林长春130112

出　　处：《中国预防兽医学报》2017年第7期578-582,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：吉林省自然科学基金(20150101104JC)

摘　　要：为制备犬瘟热病毒(CDV)单克隆抗体(MAb),本研究用腺病毒表达的CDV截短N蛋白(aa401~aa523)免疫BALB/c小鼠,取免疫小鼠脾细胞与SP2/0骨髓瘤细胞进行融合并筛选到2株能够稳定分泌抗CDV N蛋白的MAb,命名为N1-C8和N1-C41。经间接ELISA测定N1-C8和N1-C41培养上清液及小鼠腹水效价分别为1∶1 024、1∶106和1∶512、1∶105。通过肽扫描的方法筛选出N1-C8的抗原表位为线性表位^(440)ENQGGDKYPIHFNDE454,但并未能够筛选出N1-C41的抗原表位,推测可能是因为其抗原表位为空间构象型。Western blot结果显示N1-C8和N1-C41两株MAb在识别CDV的不同毒力病毒株上有差异,可能与CDV强弱病毒株在N蛋白的差异变化有关。本研究两株MAb的制备为CDV不同毒力株的鉴别诊断提供了可能。To prepare the monoclonal antibodies （MAb） against canine distemper virus （CDV）, two hybridoma cell lines, named NI-C8 and N1-C41, secreting MAb against the CDV was prepared by fusion of the SP2/0 myeloma cells with spleen cells of BALB/c mice immunized by purified recombinant nucleocapsid （aa401-aa523） protein of CDV prepared from adenovirus expression system. These culture supernatant and mouse ascites titer of the MAbs were 1：1024, 1：106 and 1：512, 1：105 for N1-C8 and NI-C41 respectively by indrect ELISA. The linear epitope 440ENQGGDKYPIHFNDE454 is the epitope of N1-C8, which was selected by peptide scanning, but the epitope of N1-C41 was not screened, and it was speculated that the epitope might be space conformation. Western blot results showed that there were differences of MAbs N1-C8 and N1-C41 in identifying different virulence strains of CDV, this phenomenon may be associated with virulent and avirulent CDV strains in N protein changes. These MAbs could have potential for developing differential diagnosis for different virulent strains of CDV.

关 键 词：犬瘟热病毒 单克隆抗体 免疫荧光 蛋白质免疫印迹 

分 类 号：S852.65[农业科学—基础兽医学]