新城疫病毒HN特异性多肽抗体间接ELISA方法的建立
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摘要
为建立快速检测新城疫病毒抗体的血清学方法,通过合成NDV主要保护性抗原HN蛋白和F蛋白上的特异性表位多肽,将其与BSA载体蛋白偶联,作为包被抗原,建立检测NDV抗体的间接ELISA方法。并对其进行敏感性、特异性、与HI方法阴阳性符合率检测。结果显示:基于"PDEQDYQIRMAKSSYKPGRFGGKRIQQAILS"多肽的该ELISA方法与其他鸡主要传染病阳性血清无交叉反应,具有良好的特异性。通过对112份临床鸡血清样品进行检测,该方法的敏感性为94.0%、特异性为100%,与HI方法检测的阴阳性符合率为96.4%。研究表明,建立的ELISA方法对血清中NDV的抗体水平检测具有较高的灵敏度和很好的特异性,可为ND疫苗免疫后抗体水平的检测和监测提供可靠工具。
Specific epitope peptides on NDV major protective antigens(HN protein and F protein) were synthesized and conjugated with BSA carrier protein to form an indirect ELISA method for detecting antibodies against NDV. And the sensitivity, specificity and positive detection rate of HI method were tested. The results showed that the ELISA method based on the "PDEQDYQIRMAKSSYKPGRFGGKRIQQAILS" peptide had no cross-reaction with other chicken-positive infectious sera, and had good specificity.By measuring 112 clinical chicken serum samples, the sensitivity of the method was 94.0%, the specificity was 100%, and the negative and positive coincidence rate with the HI method was 96.4%.The established ELISA method has high sensitivity and good specificity for detecting the antibody level of NDV in serum,and can provide a reliable tool for detecting and monitoring the antibody level of ND vaccine after immunization.
Specific epitope peptides on NDV major protective antigens(HN protein and F protein) were synthesized and conjugated with BSA carrier protein to form an indirect ELISA method for detecting antibodies against NDV. And the sensitivity, specificity and positive detection rate of HI method were tested. The results showed that the ELISA method based on the "PDEQDYQIRMAKSSYKPGRFGGKRIQQAILS" peptide had no cross-reaction with other chicken-positive infectious sera, and had good specificity.By measuring 112 clinical chicken serum samples, the sensitivity of the method was 94.0%, the specificity was 100%, and the negative and positive coincidence rate with the HI method was 96.4%.The established ELISA method has high sensitivity and good specificity for detecting the antibody level of NDV in serum,and can provide a reliable tool for detecting and monitoring the antibody level of ND vaccine after immunization.
引文
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[7]孟艳,黄梅粤,王松林.鸡新城疫检测技术研究概述[J].贵州畜牧兽医,2001(3):12-14.
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[9]郑其升,徐公豹,邓毕华,等.禽流感病毒M2特异性多肽ELISA检测方法的建立[J].畜牧与兽医,2010,42(3):77-81.
[10]杨苏珍,杨继飞,鲍登克,等.以合成肽为抗原检测O型口蹄疫病毒抗体ELISA方法的建立及评价[J].畜牧兽医学报,2009,40(12):1826-1830.
[11]陈善真,曹仁祺,刘博奇,等.以偶联多肽为抗原建立检测O型猪口蹄疫病毒抗体的ELISA方法[J].广东农业科学,2014(4):136-139.
[2]陈溥言.兽医传染病学[M].北京:中国农业出版社,2015.
[3] YIN H S, WEN X, PATERSON R G, et al. Structure of the parainfluenza virus 5 F protein in its metastable, prefusion conformation[J]. Nature, 2006, 439(7072):38-44.
[4] GRAVEL K A, MCGINNES L W, REITTIER J, et al. The transmembrane domain sequence affects the structure and function of the Newcastle disease virus fusion proteins[J].J Virol, 2011, 85(7):3486-3497.
[5]殷震,刘景华主编.动物病毒学[M]. 2版.北京:科学技术出版社,1997.
[6]邢广旭.鸡新城疫病毒HN表位抗原多肽的免疫原性鉴定及应用研究[D].郑州:河南农业大学,2007.
[7]孟艳,黄梅粤,王松林.鸡新城疫检测技术研究概述[J].贵州畜牧兽医,2001(3):12-14.
[8]王乐义.新城疫病毒间接ELISA抗体检测试剂盒的研究[D].北京:中国农业大学,2004.
[9]郑其升,徐公豹,邓毕华,等.禽流感病毒M2特异性多肽ELISA检测方法的建立[J].畜牧与兽医,2010,42(3):77-81.
[10]杨苏珍,杨继飞,鲍登克,等.以合成肽为抗原检测O型口蹄疫病毒抗体ELISA方法的建立及评价[J].畜牧兽医学报,2009,40(12):1826-1830.
[11]陈善真,曹仁祺,刘博奇,等.以偶联多肽为抗原建立检测O型猪口蹄疫病毒抗体的ELISA方法[J].广东农业科学,2014(4):136-139.