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| 猪戊型肝炎病毒Ⅳ型衣壳蛋白单克隆抗体的制备与鉴定 | |
| 引用本文： | 赵菲菲,赵钦,胡守彬,陈福勇,肖一红,周恩民.猪戊型肝炎病毒Ⅳ型衣壳蛋白单克隆抗体的制备与鉴定[J].畜牧兽医学报,2012,43(6):950-955. |
| 作者姓名： | 赵菲菲 赵钦 胡守彬 陈福勇 肖一红 周恩民 |
| 作者单位： | 1. 山东农业大学动物科技学院免疫生物学实验室,泰安,271018 2. 西北农林科技大学兽医免疫学研究所,西北农林科技大学动物医学院,杨凌712100 3. 中国农业大学动物医学院,北京,100193 |
| 基金项目： | 山东省“泰山学者”建设工程项目 |
| 摘 要： | 为获得猪戊型肝炎病毒(Hepatitis E virus,HEV)Ⅳ型衣壳蛋白单克隆抗体,将猪HEV衣壳蛋白的C端267(408—675)个氨基酸基因序列克隆入原核表达载体pET-28a(+),构建重组质粒pET-28a-ORF2-C,转化E.coli Rosetta(BL21)感受态细胞进行诱导表达,SDS-PAGE和Western blot鉴定,纯化后免疫小鼠。取免疫小鼠的脾脏与鼠骨髓瘤细胞SP2/0融合制备单克隆抗体。通过间接ELISA和竞争ELISA方法筛选并鉴定单抗。结果表明蛋白得到正确、高效表达,获得3株识别不同的抗原表位区的单克隆抗体,分别命名为Mab-1E4(IgG1)、Mab-2C7(IgG1)和Mab-2G9(IgG2b),其中1E4和2G9能阻断临床阳性猪血清,提示该2株单克隆抗体识别的抗原表位是猪HEVⅣ型衣壳蛋白上重要的抗原表位区,而单抗Mab-2C7不能阻断。本研究为猪HEVⅣ型的诊断及研究提供重要工具。 |
| 关 键 词： | 猪戊型肝炎病毒 衣壳蛋白 单克隆抗体 鉴定 |
Preparation and Characterization of Monoclonal Antibodies Specific for Capsid Protein of Swine Hepatitis E Virus Genotype Ⅳ |
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| ZHAO Fei-fei , ZHAO Qin , HU Shou-bin , CHEN Fu-yong , XIAO Yi-hong , ZHOU En-min.Preparation and Characterization of Monoclonal Antibodies Specific for Capsid Protein of Swine Hepatitis E Virus Genotype Ⅳ[J].Acta Veterinaria et Zootechnica Sinica,2012,43(6):950-955. | |
| Authors: | ZHAO Fei-fei ZHAO Qin HU Shou-bin CHEN Fu-yong XIAO Yi-hong ZHOU En-min |
| Institution: | 1.College of Animal Science and Veterinary Medicine,Shandong Agricultural University,TaiAn 271018,China;2.Veterinary Immunology Research Institute of Northwest A & F University, College of Veterinary Medicine,Northwest A & F University,Yangling 712100,China; 3.College of Veterinary Medicine,China Agricultyral University,Beijing 100193,China) |
| Abstract: | To prepare monoclonal antibodies(Mabs) against capsid protein of swine hepatitis E virus Ⅳ(HEV),the gene of swine HEV encoding C-terminal 267(408-675) amino acids of capsid protein was cloned into pET-28a(+) vector to construct recombinant plasmid pET-28a-ORF2-C.The protein was expressed in E.coli Rosetta(BL21),and identified by SDS-PAGE and Western blot.Spleens of BALB/c mice immunized with the purified protein were used to produce monoclonal antibodies(Mabs).The Mabs were selected and characterized with the indirect ELISA and competitive ELISA.Protein was expressed correctly.Three Mabs,designated Mab-2C7,2G9 and 1E4,were selected and recognized three independent epitopes.Mab-2G9 is IgG2b and Mab-2C7 and Mab-1E4 are IgG1.The positive HEV swine sera could be blocked by Mab-1E4 and Mab-2G9 indicating the epitopes recognized by Mab-1E4 and Mab-2G9 are predominant epitopes in capsid protein of swine HEV genotype Ⅳwhile Mab-2C7 can not.The results suggest that the Mab-1E4 and Mab-2G9 are potential diagnostic reagents for detection of swine HEV infection genotype Ⅳ. |
| Keywords: | swine hepatitis E virus capsid protein monoclonal antibodies identification |
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