猪δ冠状病毒胶体金试纸条检测方法的建立

为建立一种方便、快捷、实用的检测猪δ 冠状病毒(PDCoV)方法,本研究利用原核表达猪δ 冠状病毒N 蛋白,以纯化的N 蛋白免疫BALB/C小鼠,利用杂交瘤细胞技术研制2株分泌PDCoV N 蛋白单克隆抗体的杂交瘤细胞株,分别命名为E8、A10。2株杂交瘤细胞分泌单克隆抗体敏感性及特异性良好,E8亚类为IgG2a,A10亚类为IgG1。以E8作为金标抗体,A10作为检测抗体,兔抗鼠作为质控线抗体,制备猪δ 冠状病毒胶体金检测试纸条,检测猪δ 冠状病毒敏感性为100TCID50,检测猪流行性腹泻病毒、猪传染性胃肠炎病毒、猪瘟病毒、猪圆环病毒、猪δ 冠状病毒阴性对照均为阴性。本研究建立的胶体金试纸条检测方法敏感性及特异性良好。该检测方法的建立,为猪δ 冠状病毒(PDCoV)流行病学调查提供一种方法,同时为规模化养殖集团及基层兽医组织提供一种检测猪δ 冠状病毒产品。

Establishment of Colloidal Gold Strip Detection Method for Porcine Deltacoronavirus

In order to establish a rapid and effective method for the detection of porcine deltacoronavirus (PDCoV), the prokaryotic expression of PDCoV N protein was induced and BALB/c mice with purified N protein were immunized. Two monoclonal hybridioma cell lines (E8, A10) secreting PDCoV N protein antibody were developed by hybridioma cell technology. Subclass identification results showed that E8 was IgG2a, and A10 was IgG1. Using E8 as the colloidal gold labeled antibody, A10 as the detection antibody and rabbit anti-mouse as the quality control antibody, a colloidal gold strip for detecting PDCoV was prepared. The test line was visible with a detection limit of 100 TCID50 (PDCoV). And the specificity tests of dipsticks showed that porcine epidemic diarrhea virus, transmissible gastroenteritis virus, swine fever virus, porcine circovirus virus, and negative control of PDCoV were negative. The result showed that the colloidal gold strip detection method established in this study had good sensitivity and specificity, and could be used for PDCoV detection and epidemiological investigation.