猪A型口蹄疫病毒特异性IgA抗体间接ELISA检测方法的建立与初步应用

发布时间：2018-07-06 11:32

本文选题：口蹄疫 + 黏膜免疫　；参考：《中国畜牧兽医》2017年06期

【摘要】：为建立一种检测口蹄疫病毒(foot-and-mouth disease virus,FMDV)特异性IgA抗体的检测方法,本研究以原核表达系统表达纯化的FMDV结构蛋白VP1作为包被抗原,以鼠抗猪IgA单克隆抗体为二抗,辣根过氧化酶标记的羊抗鼠IgG抗体为三抗,建立猪A型FMDV特异性IgA抗体间接ELISA检测方法。确定抗原包被浓度为3.50μg/mL,二抗与三抗的最佳稀释度为1∶10 000,二抗和三抗作用时间均为30min。所建立的方法与抗猪瘟病毒、猪繁殖与呼吸道综合征病毒等病原的特异性IgA抗体间无交叉反应,A型口蹄疫感染样品的阳性检出率在90%以上,批内和批间重复性试验的变异系数介于3.16%~9.76%。该方法为监测FMDV特异性IgA抗体水平变化规律及猪的黏膜免疫效果评价及口蹄疫的早期诊断提供了一种新方法。
[Abstract]:In order to establish a method for the detection of foot-and-mouth disease virus (FMDV) specific IgA antibody, the purified FMDV structural protein VP1 was expressed in prokaryotic expression system as the coating antigen, and the mouse anti porcine IgA monoclonal antibody was used as the second antibody. The goat anti-mouse IgG antibody labeled by horseradish peroxidase (HRP) was a triple antibody. An indirect Elisa method for detection of swine FMDV specific IgA antibody was established. The concentration of antigen coating was 3.50 渭 g / mL, the best dilution of the second and third antibodies was 1:10, and the time of the second and third antibodies was 30 min. The positive rate of the method and the specific IgA antibody against swine fever virus, porcine reproductive and respiratory syndrome virus were over 90%. The coefficient of variation of intra-and inter-assay reproducibility tests ranged from 3.16 to 9.76. This method provides a new method for monitoring the changes of FMDV specific IgA antibody level, the evaluation of mucosal immune effect of pigs and the early diagnosis of foot-and-mouth disease.
【作者单位】：中国农业科学院兰州兽医研究所国家口蹄疫参考实验室家畜疫病病原生物学国家重点实验室;
【基金】：“十三五”国家重点研发计划(2016YFD0501503) 甘肃省科技支撑计划(1604NKCA045-2) 国家生猪现代产业技术体系项目(CARS-36-06B)
【分类号】：S852.651

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