基于p30-gp90融合蛋白的禽网状内皮组织增生症病毒抗体间接ELISA方法的建立
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S855.3

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“十三五”国家重点研发计划项目(2018YFD0501404)


Development of an Indirect ELISA for Detection of Antibodies against Avian Reticuloendotheliosis Virus Based on a Recombinant p30-gp90 Fusion Protein
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    摘要:

    禽网状内皮组织增生症病毒(reticuloendotheliosis virus,REV)既可以诱发禽的肿瘤性疾病,又能引起感染禽群免疫抑制,给养禽业带来重大经济损失。REV抗体监测对于该病的诊断、预防和控制具有重要意义。本试验首先通过大肠杆菌表达系统表达了重组REV p30-gp90融合蛋白,随后建立了基于该融合蛋白的REV抗体间接酶联免疫吸附试验(ELISA),并对反应条件进行了优化。结果显示,p30-gp90间接ELISA的灵敏度(1:51 200)是间接免疫荧光法(IFA)(1:6 400)的8倍,与其他常见禽类病毒无交叉反应,且具有良好的批内和批间重复性。对从山东省部分地区养殖场采集的690份血清样品,同时利用p30-gp90间接ELISA和IFA进行检测,发现20.87%的血清样品经间接ELISA检测为阳性,间接ELISA和IFA的总体符合率为96.96%。结果表明,本研究建立的间接ELISA检测方法具有特异性强、敏感性高和重复性好的优势,为REV的快速诊断、流行病学调查和血清学监测提供了技术支持。

    Abstract:

    Reticuloendotheliosis virus(REV)could cause not only avian neoplastic diseases but also immunosuppressive syndromes in infected poultry,leading to significant economic loss to poultry industry,so monitoring of REV antibodies would be vital to diagnosis,prevention and control of the disease. In the paper,an indirect enzyme-linked immunosorbent assay(ELISA)for detection of REV antibodies that was based on a recombinant p30-gp90 fusion protein expressed in E. coli expression system was established,then its conditions of reaction were optimized. The results showed that the sensitivity of p30-gp90 indirect ELISA(1:51 200)was eight times that of an indirect immunofluorescence assay(IFA)(1:6 400),but the method failed to react with other common avian viruses,with good intra-assay and inter-assay repeatability. 690 serum samples collected from farms in some regions in the province were detected respectively by p30-gp90 indirect ELISA and IFA,it was found that 20.87% samples were positive by indirect ELISA,and the overall concordance rate of indirect ELISA and IFA was 96.96%. It was concluded that the method established was with good specificity,sensitivity and repeatability,which technically supported rapid diagnosis,epidemiological investigation and serological monitoring for REV.

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栾晓宁,郭龙宗,马广斌,李玉燕,王一新,巩新民.基于p30-gp90融合蛋白的禽网状内皮组织增生症病毒抗体间接ELISA方法的建立[J].《中国动物检疫》编辑部,2021,38(10):107-114.

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