鸭坦布苏病毒E蛋白特异性抗原表位鉴定及Epitope-ELISA血清学诊断方法建立

发布时间：2018-05-14 13:39

本文选题：鸭坦布苏病毒 + 抗原表位　；参考：《畜牧兽医学报》2017年10期

【摘要】：旨在建立鸭坦布苏病毒(DTMUV)的特异性血清学诊断方法。本研究以纯化的单克隆抗体1A5为固相筛选分子,利用噬菌体展示技术鉴定DTMUV囊膜(E)蛋白的特异性抗原表位,并利用DNAStar分析抗原表位序列在DTMUV中保守性和与其他黄病毒的E蛋白相应位点的氨基酸序列相似性。结果成功筛选到DTMUV E蛋白的一个线性抗原表位_(87)YAEYI_(91),该抗原表位在DTMUV中具有高度的保守性,无氨基酸位点差异,而与其他黄病毒E蛋白相应位点的氨基酸序列不具有相似性。Dot-Blotting结果表明表位_(87)YAEYI_(91)与DTMUV阳性血清具有良好的亲和性,而与JEV、DENV、WNV等黄病毒阳性血清间不出现交叉反应,可见抗原表位_(87)YAEYI_(91)为DTMUV所特有的抗原表位。利用DTMUV特异性抗原表位87YAEYI_(91)建立Epitope-ELISA血清学诊断方法并对126份鸭血清样品进行检测,以中和试验作为标准进行比较,结果显示Epitope-ELISA检测方法的相对特异性为100%,相对敏感度为96%,并且Epitope-ELISA与其他黄病毒血清间不出现交叉反应性,表明本研究建立的Epitope-ELISA检测方法是一种特异的、灵敏的ELISA血清学诊断方法。
[Abstract]:The aim of this study was to establish a specific serological method for the diagnosis of duck tambusuvirus (DTMUV). In this study, the purified monoclonal antibody (1A5) was used as a solid phase screening molecule to identify the specific epitopes of DTMUV cystoma protein by phage display technique. DNAStar was used to analyze the conserved antigenic epitopes in DTMUV and the amino acid sequence similarity with the corresponding sites of E protein of other yellow viruses. Results A linear epitope of DTMUV E protein was successfully screened. The results of Dot-blotting showed that the epitope of tip87YAEYII-91) had good affinity with DTMUV positive serum, but had no cross reaction with other yellow virus positive sera, such as JEV / DENVWNV. It can be seen that the epitope of DTMUV is the epitope of DTMUV. The method of Epitope-ELISA serological diagnosis was established by using the epitope of DTMUV specific antigen 87YEYI. 126 duck serum samples were detected, and the neutralization test was used as the standard. The results showed that the relative specificity of Epitope-ELISA assay was 100 and the relative sensitivity was 96. There was no cross reactivity between Epitope-ELISA and other yellow virus sera, which indicated that the Epitope-ELISA detection method established in this study was a specific and sensitive method for the diagnosis of ELISA.
【作者单位】：中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室;
【基金】：十三五重点研发项目(2016YFD0500100) 国家自然科学基金面上项目(31670153) 中国农业科学院协同创新项目(CAASXTCX2016011-04-9)
【分类号】：S852.65

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