Prokaryote expression of HPeV-1 VP1 protein, production of VP1 polyclonal antibody and the development of an ELISA

文献类型： 外文期刊

第一作者： Yu, Ying

作者： Yu, Ying;Shan, Tongling;Zhu, Caixia;Hua, Xiuguo;Shan, Tongling;Wang, Chunmei;Zhu, Shaofeng;Wang, Chunmei

作者机构：

关键词： Human parechovirus;VP1 protein;Expression;Hyper-immune serum;Indirect ELISA;Serological survey

期刊名称：JOURNAL OF VIROLOGICAL METHODS （ 影响因子：2.014； 五年影响因子：2.001 ）

ISSN：

年卷期：

页码：

收录情况： SCI

摘要： The VP1 protein of human parechovirus (HPeV) plays a critical role in receptor binding based on its functional arginine-glycine-aspartic acid (RGD) motif region. Currently, only the neutralisation assay is used for seroepidemiological surveys of HPeVs. In the present study, the VP1 gene of HPeV-1 was cloned into the vector pET28a(+) to express the His-tagged VP1 protein in the bacterium Escherichia coil Rosetta. The recombinant protein was purified from inclusion bodies by Ni+-NTA affinity chromatography under denaturing conditions, followed by a refolding process in gradient urea. The identity and antigenicity of the His-tagged protein was confirmed by Western blotting using an anti-His monoclonal antibody and human HPeV-1-positive serum respectively. Polyclonal antibodies against the His-tagged VP1 protein were raised in rabbits by standard procedures, and the reactivity and specificity were tested by enzyme-linked immunosorbent assay (ELISA) and Western blot analysis. An indirect ELISA was developed based on the fusion protein VP1, and evaluated in order to facilitate the detection of antibodies in persons who had been infected naturally with HPeVs. A serological survey was performed using the assay amongst children in the Shanghai region of China; the seropositivity rate was found to be about 73%

分类号： R37

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