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基于重组病毒蛋白2的一种形式开发用于检测传染性法氏囊病病毒抗体的酶联免疫吸附测定法。

Development of an enzyme-linked immunosorbent assay for detection of antibodies against infectious bursal disease virus based on a version of the recombinant viral protein 2.

作者信息

Keller Leticia, Romanutti Carina, Zanetti Flavia Adriana

机构信息

Instituto de Ciencia y Tecnología "Dr. Cesar Milstein", Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET). Saladillo 2468, C1440FFX, Ciudad Autónoma de Buenos Aires, Argentina.

Instituto de Ciencia y Tecnología "Dr. Cesar Milstein", Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET). Saladillo 2468, C1440FFX, Ciudad Autónoma de Buenos Aires, Argentina.

出版信息

Rev Argent Microbiol. 2025 Jul-Sep;57(3):208-216. doi: 10.1016/j.ram.2024.09.003. Epub 2025 Jan 22.

Abstract

Infectious bursal disease virus (IBDV) is the etiological agent of a highly contagious and immunosuppressive disease in chickens. In poultry farms, the level of anti-IBDV antibodies of numerous serum samples must be monitored using fast and simple methodologies. Therefore, the aim of this study was to develop an enzyme-linked immunosorbent assay (ELISA), in an indirect format, using a version of mature viral protein 2 (VP2) of IBDV as coating agent. This recombinant fusion protein (His-VP2) was expressed at high levels in Escherichia coli. Bacterial inclusion bodies containing His-VP2 were successfully recovered using a simple, inexpensive and efficient method, a further purification of recombinant protein by affinity chromatography using immobilized metal chelates being unnecessary. After the VP2-ELISA was optimized, its performance was evaluated using preanalyzed sera from uninfected specific pathogen-free chickens and broilers vaccinated against IBDV in poultry farms, using a commercial ELISA kit. Based on these results, the developed assay proved to be sensitive, specific and in high agreement with the kit available on the market. In addition, the in-house ELISA demonstrated to be reproducible by intra-assay and inter-assay variability studies. In conclusion, VP2-ELISA could be an efficient and low-cost alternative diagnostic method to detect antibodies to IBDV in the poultry industry.

摘要

传染性法氏囊病病毒(IBDV)是鸡的一种高度传染性和免疫抑制性疾病的病原体。在养禽场中,必须使用快速且简单的方法监测众多血清样本中的抗IBDV抗体水平。因此,本研究的目的是以间接形式开发一种酶联免疫吸附测定(ELISA),使用IBDV成熟病毒蛋白2(VP2)的一种形式作为包被剂。这种重组融合蛋白(His-VP2)在大肠杆菌中高水平表达。使用一种简单、廉价且高效的方法成功回收了含有His-VP2的细菌包涵体,无需使用固定化金属螯合物通过亲和色谱法对重组蛋白进行进一步纯化。在优化VP2-ELISA后,使用来自未感染的无特定病原体鸡的预先分析的血清以及养禽场中接种过IBDV疫苗的肉鸡血清,通过商业ELISA试剂盒评估其性能。基于这些结果,所开发的检测方法被证明是灵敏、特异的,并且与市场上现有的试剂盒高度一致。此外,通过批内和批间变异性研究表明自制ELISA具有可重复性。总之,VP2-ELISA可能是家禽业中检测IBDV抗体的一种高效且低成本的替代诊断方法。

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