空肠弯曲菌表面蛋白特异性多表位抗原的设计、表达与鉴定

构建空肠弯曲菌表面蛋白特异性多表位抗原原核表达载体,并在大肠杆菌中诱导表达,免疫动物后检测其抗血清与空肠弯曲菌菌体的反应性。从空肠弯曲菌的6个表面蛋白中分析到特异性抗原表位,人工合成其串联基因片段,将其插入原核表达载体,获得重组质粒pET-32a(+)-CJMEA-A,经IPTG诱导后表达出25k的目标蛋白,纯化后免疫新西兰大白兔,Western blot和间接ELISA检测重组蛋白抗原性和抗血清反应性。结果表明目标蛋白具有良好的反应原和免疫原性,抗血清能与菌体发生反应。其抗体可用于C.jejuni免疫磁珠和免疫胶体金等检测方法的建立。

Design,Expression and Identification of Specific Multi-Epitope Antigen of Campylobacter Jejuni Surface Proteins

To construct and to express the gene of specific multi-epitope antigen from Campylobacter jejuni and to detect the reactivity of its antibody to this bacteria.Firstly,some specific antigenic determinants were obtained after 6 surface proteins of Campylobacter jejuni were analyzed.Secondly these epitopes were linked in series and its gene was synthesized.Thirdly,the recombinant plasmid pET-32a(+)-CJMEA-A was obtained when the gene was inserted into pET-32a(+) vector.Then,after being induced by IPTG,the 25ku of fusion protein was expressed,purified and used for the immunization of rabbits.Finally,SDS-PAGE and indirect ELISA were used to detect the antigenicity of the protein and the reactivity of the antiserum.The results showed that this protein was expressed successfully;it possesses good immunogenicity and reactogenicity;its antiserum can react to thalli of C.jejuni.A conclusion can be made that the antibody of this protein could be used to detect C.jejuni by immunomagnetic beads or immune colloidal gold technique.