抗犬瘟热病毒荧光标记单抗的制备和初步鉴定

Preparation and Preliminary Identification of Fluorescein Labeled Monoclonal Antibody Against Canine Distemper Virus

[目的]建立利用单克隆抗体检测犬瘟热病毒的直接免疫荧光诊断方法。[方法]通过搅拌法,用异硫氰酸荧光素标记G蛋白纯化的抗犬瘟热病毒单抗CE3,对荧光抗体进行纯化、鉴定,并确定其最适工作浓度。对61份临床可疑犬瘟热病料进行直接免疫荧光检测。[结果]吸收试验、阻断试验和特异性试验结果显示,标记抗体具有高度的敏感性和特异性,与犬细小病毒(CPV)、犬副流感病毒(CPIV)、犬腺病毒(CAV)、狂犬病毒(RV)无交叉反应,其最优化工作浓度为1∶80。对临床可疑犬瘟热病料的阳性检出率是48%。[结论]该研究建立的直接免疫荧光方法具有快速、特异和简便的优点,对犬瘟热疾病的早期诊断具有重要意义。

[ Objective ] The study aimed to devetope a direct immunofluorescence technique for the antigen detection of canine distemper using monoclonal antibody against canine distemper virus. [ Method] By stirring method, the monoclonal antibody CE3 purified by protein G was labeled with fluorescein isothiocyanate （FITC）. After purification and identification, the optimal working concentration of the fluorescent antibody was determined. Subsequently, 61 samples collected from suspected dogs were detected by direct immunofluorescence assay. [ Result] The absorption, blocking and specificity tests showed that the labeled antibody had high specificity and sensitivity, showing no cross reaction with canine parvovirus （ CPV）, canine parainfluenza virus （ CPIV）, rabies virus （RV） and canine adenovirus （CAV）. Its optimal working concentration was 1：80. The positive rate of clinical suspected samples was 48%. [ Conclusion ] The direct immunofluorescence method developed in this study possesses such advantages as rapidness, specificity and convenience, having an important significance to early diagnosis of canine distemper disease.