圣路易斯脑炎病毒样颗粒的真核表达、纯化与鉴定

目的 利用293T细胞表达、纯化圣路易斯脑炎病毒样颗粒（Virus like particles,VLPs）,为研制圣路易斯脑炎病毒免疫诊断试剂奠定基础.方法 构建含圣路易斯脑炎病毒PrM和E基因的重组质粒pcDNA5/FRT-SJME,瞬时转染293T细胞,表达并纯化重组蛋白,通过透射电镜、间接免疫荧光试验、Western-Blot和ELISA对其进行鉴定.结果 表达的重组蛋白形成50 nm的球型颗粒,能与抗圣路易斯脑炎病毒抗体产生特异性反应,与部分黄病毒属阳性血清发生交叉反应.结论 已成功在哺乳动物细胞中表达并纯化了圣路易斯脑炎病毒样颗粒,其具有良好的抗原性和完整的颗粒形态,为进一步研制圣路易斯脑炎病毒感染的免疫诊断试剂奠定了基础.

Expression, purification and identification of St. Louis encephalitis virus-like particles in eukaryotic cells

Objective To express St.Louis encephalitis virus-like particles in mammalian cells,it will provide a prerequisite for further immunological diagnostic studies.Methods 293T-cell were transiently transfected with recombinant PreM-E plasmid.Expression and antigenicity of the purified protein were determined by transmission electron microscope (TEM),Western-Blot,immunofluorescence assay and ELISA.Results Recombinant subviral particles,about 50 nm in diameter,were observed by TEM in the supernatant of transfected cells.The results of Westeru-Blot,IFA and ELISA showed the recombinant proteins retained immunoreactivity similar to those of native virus proteins.Conclusion St.Louis encephalitis viruslike particles has good antigenicity and physical appearance,It will provide a prerequisite for further immune diagnostic reagent.