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| 羊朊毒体单抗结合表位分析 | |
| 引用本文： | 张永强,吴晓东,赵永刚,鲍恩东,王清华,张维,刘雨田,王志亮.羊朊毒体单抗结合表位分析[J].生物工程学报,2009,25(3):348-353. |
| 作者姓名： | 张永强 吴晓东 赵永刚 鲍恩东 王清华 张维 刘雨田 王志亮 |
| 作者单位： | 1. 中国动物卫生与流行病学中心,国家外来动物疫病诊断中心,青岛,266032;南京农业大学动物医学院,南京,210095 2. 中国动物卫生与流行病学中心,国家外来动物疫病诊断中心,青岛,266032 3. 南京农业大学动物医学院,南京,210095 |
| 基金项目： | “十一五”国家科技支撑计划(No.2006BAD06A13);;现代农业产业(奶牛)技术体系建设(No.2060302)资助~~ |
| 摘 要： | 通过分段表达PrP核心片段和人工合成多肽,分析5株羊朊毒体单抗结合表位。分段表达PrP核心片段,通过PCR方法扩增目的片段,经酶切、连接后,将目的片段插入质粒pET32a,在大肠杆菌BL21中表达。将表达的系列融合蛋白与单抗进行免疫转印试验,根据反应情况确定单抗结合的大致部位,在此基础上设计合成多条针对性多肽,用ELISA方法进一步确定3株单抗的结合部位;通过与6段融合蛋白反应证明5株单抗的结合部位分别为:2H3在199aa～213aa之间,4C6、5F11和7F11在139aa～168aa之间,7F1在214aa～227aa之间,与3段人工合成多肽进行ELISA反应进一步得到4C6、5F11和7F11抗原结合表位在149aa～158aa之间;本研究确定了5株单抗在PrP分子上的结合部位,为羊痒病和牛海绵状脑病的检测、发病机制的研究奠定了基础。 |
| 关 键 词： | PrP核心片段 人工合成多肽 单抗 抗原结合表位 融合蛋白 羊痒病 牛海绵状脑病 |
| 收稿时间： | 2008/10/13 0:00:00 |
Analysis of monoclonal antibody binding sites in ovine prion protein |
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| Yongqiang Zhang,Xiaodong Wu,Yonggang Zhao,Endong Bao,Qinghua Wang,Wei Zhang,Yutian Liu and Zhiliang Wang.Analysis of monoclonal antibody binding sites in ovine prion protein[J].Chinese Journal of Biotechnology,2009,25(3):348-353. | |
| Authors: | Yongqiang Zhang Xiaodong Wu Yonggang Zhao Endong Bao Qinghua Wang Wei Zhang Yutian Liu and Zhiliang Wang |
| Institution: | National Diagnostic Center for Exotic Animal Diseases, China Animal Health and Epidemiology Center, Qingdao 266032, China; College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China;National Diagnostic Center for Exotic Animal Diseases, China Animal Health and Epidemiology Center, Qingdao 266032, China;National Diagnostic Center for Exotic Animal Diseases, China Animal Health and Epidemiology Center, Qingdao 266032, China;College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China;National Diagnostic Center for Exotic Animal Diseases, China Animal Health and Epidemiology Center, Qingdao 266032, China;National Diagnostic Center for Exotic Animal Diseases, China Animal Health and Epidemiology Center, Qingdao 266032, China;National Diagnostic Center for Exotic Animal Diseases, China Animal Health and Epidemiology Center, Qingdao 266032, China;National Diagnostic Center for Exotic Animal Diseases, China Animal Health and Epidemiology Center, Qingdao 266032, China |
| Abstract: | Binding sites of five monoclonal antibodies were obtained by reinforceable method of overlapping recombinant prion protein and synthetic peptide. Overlapping peptides of PrP core were expressed in Escherichia coli by insertion of serial PCR amplicons of ovine PrP gene fragments into pET32a. The expressed fusion peptides were then tested for the binding activity to PrP monoclonal antibodies in Western blotting. The binding sites of 5 monoclonal antibodies of ovine PrP were located respectively as follows: 2H... |
| Keywords: | binding site monoclonal antibody synthetic peptide PrP core fusion peptide scrapie bovine transmissible spongiform encephalopathy |
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