犬瘟热病毒单克隆抗体的研制及胶体金试纸条检测方法的建立

Preparation of Monoclonal Antibody against Canine Distemper Virus and Development of Colloidal Gold Strip

关键词:  犬瘟热病毒  N 蛋白  单克隆抗体  胶体金试纸条

Key words:  Canine distemper virus  N protein  monoclonal antibody  colloidal gold strip

摘要:

本研究利用原核表达、纯化的犬瘟热病毒(Canine distemper virus,CDV)N 蛋白免疫BALB/C小鼠,成功筛选到两株分泌CDV N 蛋白单克隆抗体的杂交瘤细胞株,分别命名为D3、D6。2株杂交瘤细胞分泌单克隆抗体敏感性、特异性良好,能识别不同来源的犬瘟热病毒株,单抗亚类均为IgG1。以D3作为金标抗体,D6作为检测抗体,兔抗鼠IgG作为质控线抗体,制备犬瘟热病毒胶体金检测试纸条,检测犬瘟热病毒敏感性为1000TCID50,能有效检测区分犬瘟热病毒、犬细小病毒(Canine parvovirus,CPV)、犬腺病毒2型(Canine adenovirus type 2,CAV2)、狂犬病病毒(Rabies virus,RV)。综上,本研究建立的胶体金试纸条检测方法,灵敏度高、特异性强,适用于犬瘟热病毒临床快速诊断。

Abstract:

In this study, we induced the prokaryotic expression of canine distemper virus (CDV) N protein and immunized BALB/C mice with purified N protein. Two monoclonal hybridioma cell lines (D3, D6) secreting CDV N protein antibody were developed by hybridioma cell technology. The monoclonal antibodies secreted by the two hybridoma cells were sensitive and specific, and could recognize different canine distemper virus strains. The monoclonal antibody subclass both was IgG1. Using D3 as the colloidal gold labeled antibody, D6 as the detection antibody and rabbit anti-mouse as the quality control antibody, a colloidal gold strip for detecting porcine coronavirus was prepared. The test line was visible with a detection limit of 1000 TCID50 (CDV). And the specificity tests of dipsticks showed that could distinguish CDV, canine parvovirus(CPV), canine adenovirus type 2 (CAV2) and rabies virus (RV). The colloidal gold strip detection method established in this study has good sensitivity and specificity, and could be suitable for rapid clinical diagnosis of CDV.