摘要
目的:对含有HEV主要抗原表位的嵌合蛋白进行表达与纯化,并对其免疫原性进行研究.方法:将含有一段HEV ORF2基因片段(394～660 aa)和HEVORF3全基因的表达质粒pHEV23在大肠杆菌中进行异丙基硫代半乳糖苷(IPTG)诱导表达,其表达产物经Ni2+-NTA树脂亲和纯化,透析复性.纯化蛋白免疫小鼠,用ELISA检测小鼠血清IgG抗体,T细胞增殖反应检测细胞免疫应答.以Western-blot检测纯化蛋白对患者阳性血清的免疫反应性.结果:纯化蛋白的分子量为 55 KDa,纯度达 90% 以上;实验组特异性抗体水平明显高于对照组;T细胞增殖反应实验组与对照组比较有极显著性差异.纯化蛋白能与患者阳性血清呈特异反应. 结论:嵌合蛋白具有良好的免疫原性和免疫反应性,为进一步研究开发HEV诊断试剂盒及基因工程疫苗提供了基础.
Objective：To explore the immunogenicity of a HEV recombinant chimeric protein expressed in E. coli, Methods： Recombinant plasmids pHEV23 containing a fragment of HEV ORF2 gene and a full length of ORF3 gene was transformed into E-coli B1 -21. Expressed chimeric protein was analyzed by SDS- PAGE and Western-blot. Blab/c mice were immunized intramuscularly with purified protein twice. Antibody responses and T cell proliferation were then detected. Results：The SDS-PAGE showed a dominant additional protein with molecular weight of 55 KDa which could specially react with the sera of patients with hepatitis E. Expressed chimeric protein could induced high specific anti HEV IgG and T lymphocyte proliferation. Conclusion： Expressed chimeric protein have a good immunogenicity and may be the promising candidate for a HEV vaccine.
出处
《中国卫生检验杂志》
CAS
2005年第9期1037-1039,共3页
Chinese Journal of Health Laboratory Technology
基金
浙江省自然基金青年人才培养资金项目(RC01054)
浙江省科技厅项目(2004F11G136001)
