猪流行性腹泻病毒IgA抗体ELISA检测方法的建立和初步应用
Establishment and preliminary application of an ELISA method for detection of porcine epidemic diarrhea virus IgA antibody
王婉莹 1石正旺 2罗俊聪 2廖焕程 2冯露 2郑海学 2田宏 2蔺国珍3
作者信息
- 1. 西北民族大学生命科学与工程学院,甘肃兰州 730030;中国农业科学院 兰州兽医研究所 兰州大学动物医学与生物安全学院 动物疫病防控全国重点实验室,甘肃兰州 730000
- 2. 中国农业科学院 兰州兽医研究所 兰州大学动物医学与生物安全学院 动物疫病防控全国重点实验室,甘肃兰州 730000
- 3. 西北民族大学生命科学与工程学院,甘肃兰州 730030
- 折叠
摘要
为检测猪流行性腹泻病毒(PEDV)IgA抗体,以PEDV重组S蛋白作为包被抗原,辣根过氧化物酶标记的小鼠抗猪IgA单克隆抗体为检测抗体,建立并优化了PEDV IgA抗体的ELISA检测方法,并评估了其特异性、敏感性、重复性及临床样品检测性能.结果显示:该方法抗原最佳包被质量浓度为2 µg/mL,酶标抗体的最佳工作浓度为1∶25000,血清最佳稀释度为1∶50;与猪伪狂犬病病毒、非洲猪瘟病毒、猪瘟病毒、塞内卡病毒、口蹄疫病毒阳性血清均无交叉反应;灵敏度为1∶800;批内、批间变异系数均小于10％;与IDEXX商品化试剂盒的检测结果具有高度一致性.结果证明,本研究中建立的PEDV IgA抗体检测方法特异性强、敏感性高、重复性好,适用于猪流行性腹泻的临床诊断与防控、流行病学调查以及免疫效果评估.
Abstract
To identify the IgA antibody against porcine epidemic diarrhea virus(PEDV),an ELISA technique was developed and refined using PEDV recombinant S protein as the coating antigen and horseradish peroxidase-conjugated murine anti-porcine IgA monoclonal antibody as the identifying an-tibody,and its specificity,sensitivity,reproducibility,and performance of clinical samples were evaluated.The findings indicated that the optimal antigen encapsulation mass concentration was 2 μg/mL,the best enzyme-labeled antibody working concentration was 1∶25 000,and the most effective serum di-lution was 1∶50.There was no cross-reactivity with positive serums of porcine pseudorabies virus,African swine fever virus,classical swine fever virus,Senecavirus A and foot-and-mouth disease virus.The sensitivity was determined to be 1∶800.The coefficients of variation were less than 10％within and among different batches,and the consistency rate with IDEXX commercial kit was very high in testing result.These results prove that the detection method for PEDV IgA antibody developed in this research has good specificity,sensitivity and reproducibility,making it suitable for clinical diagnosis,di-sease prevention,and control of porcine epidemic diarrhea,as well as for epidemiological studies and evaluation of vaccination efficacy.
关键词
猪流行性腹泻病毒/S蛋白/IgA抗体/ELISAKey words
porcine epidemic diarrhea virus/S protein/IgA antibody/ELISA引用本文复制引用
基金项目
甘肃省科技重大专项(22ZD6NA012)
甘肃省科技重大专项(22ZD6NA001)
甘肃省自然科学基金项目(21JR1RA203)
中央高校基本科研业务费专项(lzujbky-2022-ct02)
国家生猪技术创新中心先导科技项目(NCTIP-XD/C03)
出版年
2024
国家科技期刊平台
NSTL
万方数据