| 猪瘟病毒E2基因在真核细胞中表达及抗E2蛋白单克隆抗体的制备 |
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| 引用本文： | 许信刚,胡建和,张彦明,张海棠,陈永耀. 猪瘟病毒E2基因在真核细胞中表达及抗E2蛋白单克隆抗体的制备[J]. 畜牧兽医学报, 2005, 36(12): 1318-1322 |
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| 作者姓名： | 许信刚 胡建和 张彦明 张海棠 陈永耀 |
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| 作者单位： | 1. 西北农林科技大学动物科技学院,杨凌,712100
2. 河南科技学院动物科学系,新乡,453003 |
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| 基金项目： | 基金项目：国家自然科学基金项目（30270988） |
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| 摘 要： | 利用DNA重组技术将猪瘟病毒（CSFV）石门株囊膜蛋白E2基因插入逆转录病毒载体pBABE-puro中构建重组逆转录病毒载体pBABE-puro-E2,该重组逆转录病毒载体与pVSVg质粒经磷酸钙共转染法转入293T细胞中包装逆转录病毒假病毒.用包装的假病毒感染SP2/0细胞,经嘌呤霉素筛选阳性细胞后进行流式细胞术（FACS）分析,结果表明CSFV E2基因在SP2/0细胞膜上成功表达.将表达E2蛋白的SP2/0细胞腹腔免疫BALB/c小鼠,成功诱导小鼠产生了抗E2蛋白的抗体.取免疫小鼠脾细胞与SP2/0骨髓瘤细胞融合,经克隆和筛选获得了4株稳定分泌抗猪瘟病毒E2蛋白单克隆抗体的杂交瘤细胞株,所分泌的单抗可与CSFV产生特异性反应并具有中和活性.
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| 关 键 词： | 猪瘟病毒 E2囊膜蛋白 逆转录病毒载体 假病毒 FACS 单克隆抗体 |
| 文章编号： | 0366-6964（2005）12-1318-05 |
| 收稿时间： | 2004-07-02 |
| 修稿时间： | 2004-07-02 |
Expression of CSFV E2 Envelope Protein Gene in Eukaryotic Cells and Preparation of Monoclonal Antibodies against CSFV E2 Envelope Protein |
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| XU Xin-gang,HU Jian-he,ZHANG Yan-ming,ZHANG Hai-tang,CHEN Yong-yao. Expression of CSFV E2 Envelope Protein Gene in Eukaryotic Cells and Preparation of Monoclonal Antibodies against CSFV E2 Envelope Protein[J]. Chinese Journal of Animal and Veterinary Sciences, 2005, 36(12): 1318-1322 |
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| Authors: | XU Xin-gang HU Jian-he ZHANG Yan-ming ZHANG Hai-tang CHEN Yong-yao |
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| Affiliation: | 1. College of Animal Science and Technology , Northwest Sci-Tech University of Agriculture and Forestry, Yangling 712100, China;2. Department of Animal Science, Henan Institute of Science and Technology , Xinxiang 453003, China |
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| Abstract: | The recombinant retroviral vector pBABE-puro-E2 was constructed by inserting E2 gene full-length cDNA of CSFV Shimen strain into pBABE-puro.Both the recombinant retroviral vector and pVSVg plasmid were transfected into 293T cells by calcium phosphate transfection method,thus the pseudovirus were produced.The pseudovirus infected eukaryotic cells SP2/0 and expression E2 protein was determined by puromycin-resistant and FACS analysis.BALB/c mice were injected in abdomen with expressing E2 protein SP2/0 cells.Anti-CSFV E2 antibody was screened by FACS.The results showed that CSFV E2 protein was expressed in SP2/0 cells' envelope protein successfully.FACS could detect specific anti-E2 antibody in immune mouse serum.Mouse spleen cells were fused with myeloma SP2/0.Culture supernatants of hybridoms were screened by FACS and four cell lines which could secrete against E2 protein of CSFV McAbs stably were obtained.The McAbs not only react with(CSFV) specially but also had neutralization activity.The McAbs type belongs to IgM. |
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| Keywords: | CSFV E2 envelope protein retroviral vector pseudovirus FACS McAbs |
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