拟态弧菌OmpU蛋白B细胞表位的筛选鉴定及其免疫活性研究

李槿年

摘 要：拟态弧菌是一种人和水产动物共患病病原菌，不仅引起水产动物严重的腹水病，而且可经水体和水产品感染人类，引起腹泻和食物中毒。拟态弧菌OmpU蛋白是一种具有良好抗原性和保守性的首选诊断抗原和疫苗候选分子，但其抗原表位仍不清楚。对此，该项目拟开展“拟态弧菌OmpU蛋白B细胞表位及其免疫活性研究”。应用生物信息软件预测OmpU蛋白B细胞线性表位，进而利用合成肽ELISA鉴定预测表位，确定优势B细胞线性表位。再以OmpU多克隆抗体和单克隆抗体为靶标从噬菌体随机肽库中筛选鉴定优势B细胞模拟表位。最后，对表（拟）位噬菌体克隆进行免疫活性检测，分析不同表位的免疫活性差异。本项目预期研究成果不仅为建立以表位为基础的特异快速的诊断方法和研制表位疫苗奠定基础。

关键词：拟态弧菌 OmpU蛋白 B细胞表位 免疫活性

Abstract： Vibrio mimicus （V. mimicus） is a zoonisis pathogens. It not only cause ascitic fluid disease of aquatic animals， but also cause human gastroenteritis and food poisoning through contacting aquatic environment and aquatic products. The out membrance protein U （OmpU） of V. mimicus with higher antigenicity and conservative property is taken as a primary diagnostic antigen and good candidate protein for developing vaccine， but B-cell epitopes of OmpU protein are not clear now. In order to design new vaccine and diagnosing reagent in the future， the B-cell eptopes of OmpU protein of V. mimicus and their immune characteristics were researched. Firstly， B-cell epitopes of OmpU protein were analyzed with bio-softwares Biosun and Hopp-wood. The putative epitopes peptides were synthesized and were defined by peptide enzyme-linked immunosorbent assay. Secondly， in order to define B-cell mimotopes of OmpU protein， the phage clones binding to purified polyclonal antibody and monoclonal antibody against OmpU protein were selected from the 12-mer random peptide library. Then positive monoclonal phages were sequenced and analyzed. Finally， the immunogenicity and immunoreactivity of differnt phage clones containing B-cell epitopes were determined with immunoassay to compare their immune characteristics.

Key Words： Vibrio mimicus； OmpU protien； B cells epitopes； Immune characteristics

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