中华实验和临床病毒学杂志2024,Vol.38Issue(6) :702-708.DOI:10.3760/cma.j.cn112866-20240829-00125

人H组轮状病毒VP7蛋白表达及其多克隆抗体制备

Expression and generation of the polyclonal antibody of human group H rotavirus VP7 protein

刘晓钰 赵颖 张璐琦 欧阳瑄泽 章青 李丹地 李金松 李利利 孙晓曼 段招军
中华实验和临床病毒学杂志2024,Vol.38Issue(6) :702-708.DOI:10.3760/cma.j.cn112866-20240829-00125

人H组轮状病毒VP7蛋白表达及其多克隆抗体制备

Expression and generation of the polyclonal antibody of human group H rotavirus VP7 protein

刘晓钰 1赵颖 1张璐琦 1欧阳瑄泽 1章青 2李丹地 2李金松 2李利利 2孙晓曼 3段招军3
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作者信息

  • 1. 甘肃中医药大学公共卫生学院,兰州 730000
  • 2. 中国疾病预防控制中心病毒病预防控制所传染病溯源预警与智能决策全国重点实验室 国家卫生健康委员会医学病毒和病毒病重点实验室,北京 102206
  • 3. 甘肃中医药大学公共卫生学院,兰州 730000;中国疾病预防控制中心病毒病预防控制所传染病溯源预警与智能决策全国重点实验室 国家卫生健康委员会医学病毒和病毒病重点实验室,北京 102206
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摘要

目的 表达和纯化人H组轮状病毒(group H rotavirus,RVH)J19株的VP7蛋白,制备VP7的多克隆抗体(多抗)并对其多抗功能进行鉴定.方法 通过杆状病毒系统表达RVH J19株的VP7蛋白,通过亲和层析和凝胶过滤层析方法进行蛋白纯化.将纯化得到的RVH J19株的VP7蛋白免疫兔子以制备VP7多抗血清,通过酶联免疫吸附测定(enzyme-linked immunosorbent assay,ELISA)、蛋白质免疫印迹法(Western blot,WB)、免疫荧光荧光试验(Immunofluorescence,IF)等对VP7多抗血清进行鉴定.结果 纯化得到可溶的J19 VP7蛋白;制备获得J19株VP7兔多抗;ELISA显示VP7多抗具有较好的结合效价,在稀释2 048 000倍时与人RVH VP7蛋白仍有结合;WB表明多抗能够结合J19 VP7蛋白;IF显示VP7多抗可以特异识别人H组轮状病毒J19株.结论 本研究成功得到了人H组轮状病毒J19 VP7蛋白,制备的VP7多抗可以特异性识别人H组轮状病毒,为人H组轮状病毒的检测及病毒衣壳结构功能研究提供了基础.

Abstract

Objective To express and block the VP7 protein of human group H rotavirus(group H rotavirus,RVH)J19,and to prepare the polyclonal adsorbent polyantibody function of VP7 for identification.Methods The VP7 protein of human H rotavirus J19 strain was expressed by baculovirus system,and the protein was blocked by affinity chromatography and hydrophobic chromatography.The VP7 polyantiserum was prepared by immunizing the VP7 protein of human H group H rotavirus J19,and the VP7 polyantiserum was identified by ELISA,Western blot(WB)and immunofluorescence.Results Soluble J19 VP7 protein was obtained.J19 VP7 rabbit polyclonal antibody was prepared.ELISA showed that VP7 polyclonal antibody had a good binding effect,binding to human RVH VP7 protein at 2 048 000-fold.WB showed that the polyclonal antibody could bind to J19 VP7 protein;Immunofluorescence assay showed that VP7 polyclonal antibody could specifically recognize human group H rotavirus J19.Conclusions The J19 VP7 protein of human H rotavirus was successfully obtained,and the prepared VP7 polyclonal antibody could recognize human H rotavirus,which provided a basis for the detection of human H rotavirus and the study of the structure and function of the viral capsid.

关键词

人H组轮状病毒/杆状病毒表达系统/VP7蛋白/多克隆抗体

Key words

Human group H rotavirus(RVH)/Baculovirus expression system/VP7 protein/Polyclonal antibody

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出版年

2024
中华实验和临床病毒学杂志
中华医学会

中华实验和临床病毒学杂志

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影响因子:0.718
ISSN:1003-9279
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