SARS冠状病毒S蛋白多价核酸疫苗的构建和免疫学原性

目的构建以SARS冠状病毒（severe acutere spiratory syndrome coronavlrus，SARs．c0V）S蛋白3个保守片段基因的组合为抗原基因DNA疫苗，采用肌肉注射的方法接种小鼠后观察机体产生免疫应答的情况。方法将人工合成的S蛋白3个保守片段基因组合（称为Sa）克隆至真核表达载体pcDNA3、1（-），随之将重组质粒进行小鼠肌肉免疫，定期检测血清中抗SARS—CoV的病毒特异性抗体水平，流式细胞仪观察其淋巴细胞表型变化，PCR法检测质粒分布，免疫组化检测抗原表达。结果疫苗注射后15d就能在血清中检测出病毒特异性抗体，并且疫苗能在机体中持续表达抗原引发机体持续的免疫应答，直到注射后45d免疫应答都不见减弱；以CTLT淋巴细胞为主的CD8＋T淋巴细胞的百分比含量增加极显著，引起强大的体液免疫和细胞免疫；而空载体质粒对照组未检测出明显的特异性免疫应答。结论以S蛋白3个保守片段基因组合为抗原基因的DNA疫苗通过肌注能诱导小鼠针对SARS病毒强大的免疫应答。

Immune characteristics of plasmid DNA and cloning of S gene conservative fragments in severe acute respiratory syndrome coronavirus

Objective To construct DNA vaccine plasmid against conservative gene of SARS coronavirus(SARS-CoV) S protein,and observe the immune response induced by DNA vaccine.Methods A recombinant DNA vaccine plasmid was constructed by cloning SARS-CoV S gene conservative fragments into pcDNA,which could induce SARS-CoV specific antibodies in mice.After mice immunized with the recombinant DNA vaccine by injection,the SARS-CoV antibodies in the sera of mice were detected by ELISA,The lymphocyte phenotypes were detected by FACS,distribution of the plasmids was detected by PCR,special antigen expressed in tissues were detected with immuno-histochemical analysis.Results 15days after injection special antibodies against SARS-CoV can be detected in the sera,and its quantity was directly increased as time went by;30days after injection it reached a plateau,it can induce potent humoral and cellular immune response in mice,there were dramatic leaps in the quantity of CD8+T cells;while no immune response could be observed in the control group.Conclusions Conservative regions of SARS-CoV S protein gene can be used as candidate antigens of DNA vaccine to induce SARS-CoV specific antibodies,it can induce potent humoral and cellular immune response in mice.