基础医学与临床 ›› 2010, Vol. 30 ›› Issue (12): 1303-1308.

• 研究论文 • 上一篇    下一篇

SARS-CoV相关重组B细胞线性多表位肽的表达及鉴定

周建华1,曾玉红1,崔莲仙2,康宁1,何维2   

  1. 1. 中国医学科学院 基础医学研究所 北京协和医学院 基础学院2. 中国医学科学院基础医学研究所 北京协和医学院基础学院
  • 收稿日期:2010-03-24 修回日期:2010-04-30 出版日期:2010-12-05 发布日期:2010-12-05
  • 通讯作者: 何维

Preparation and characterization of recombinant poly-epitope peptides based on SARS-CoV B cell recognized linear epitopes

ZHOU Jian-hua 1,ZENG Yu-hong 1,CUI Lian-xian 1,KANG Ning 1,HE Wei 2   

  1. 1. Institute of Basic Medical Sciences, CAMS & PUMC2. IBMS & School of Basic Medicine, CAMS & PUMC
  • Received:2010-03-24 Revised:2010-04-30 Online:2010-12-05 Published:2010-12-05
  • Contact: HE Wei

摘要: 目的 利用已筛选到的线性B细胞表位组合得到重组多表位肽,并验证其用作表位肽疫苗的可行性。方法 用搭桥PCR和人工合成DNA方法获得多表位肽编码基因片段,构建原核表达载体表达多表位肽。ELISA法测其与SARS病人血清的交叉反应性及其用于免疫动物获得的多克隆抗血清的效价。Western blot检测抗原抗体结合特异性。中和试验检测中和SARS-CoV效果。结果 成功获得PEP1和PEP2编码基因片段并表达重组多表位肽rPEP1和rPEP2。纯化的rPEP1和rPEP2与病人血清的交叉反应率分别为88.7%和97.2%;与免疫动物血清具有结合特异性,结合效价分别为1×106和5×105;中和SARS-CoV的效率分别为70%和80%。结论 成功获得交叉反应性好、免疫原性强的重组多表位肽rPEP1和rPEP2,为制备多表位肽疫苗提供了实验依据。

关键词: SARS, 疫苗, 重组多表位肽, 表位疫苗

Abstract: Objective To obtain recombinant poly-epitope peptides based on selected linear B cell epitope and evaluate its probability as epitope vaccines. Methods Poly-epitope peptides were expressed by prokaryotic expression vector with multi-epitope peptides coding gene fragments, which were obtained through gene engineering technology. The cross-reactivity of tested peptides with the SARS patients sera and titers to serum from rabbits immunized with recombinant poly-epitope peptides were determined by ELISA. Their specificity was measured by Western blotting, while SARS-CoV-neutralizing response was identified by neutralization technology. Results PEP1 and PEP2 encoding gene fragments were successfully obtained and the recombinant poly-epitope peptides rPEP1 and rPEP2 were successfully expressed. Cross-reaction rate of purified rPEP1 and rPEP2 with the patient serum were 88.7% and 97.2%, respectively. They can be recognized by immunized rabbits serum, binding titers were 1×106 and 5×105, and SARS-CoV neutralizing efficiency were 70% and 80% respectively. Conclusions Recombinant poly-epitope peptides rPEP1 and rPEP2 with high cross-reactivity, strong immunogenicity were successfully obtained. It provided experimental basis for the preparation of poly-epitope peptide vaccine.

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