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Rv2628蛋白是结核分枝杆菌Mycobacterium tuberculosis(M.tb)DosR调控的潜伏感染相关抗原。本研究对Rv2628蛋白进行了原核表达和纯化,并以巨噬细胞系和小鼠为研究模型,对其免疫生物学特性进行了分析。SDS-PAGE及Western blotting鉴定结果表明,Rv2628-His融合蛋白以包涵体形式表达,能与兔抗H37Rv多抗血清发生特异性反应,具有较好的免疫反应性。与巨噬细胞系RAW264.7的互作实验结果表明,在1–12 h内Rv2628蛋白能诱导前炎性因子IL-6的上调表达。将纯化的Rv2628融合蛋白皮下免疫BALB/c小鼠,夹心ELISA的测定结果表明,Rv2628蛋白免疫组诱导产生的特异性IFN-γ水平显著高于IL-4的水平(P<0.000 1),呈现Th1型细胞免疫应答趋势;以Rv262811-30多肽作为包被抗原,通过间接ELISA测定的血清抗体效价能达到11 600,表明Rv2628也能诱导体液免疫应答。总之,Rv2628能促进巨噬细胞炎症反应的发生,激发小鼠产生强烈的Th1型细胞免疫应答和较好的体液免疫应答,具有作为亚单位疫苗的潜力,为M.tb与宿主之间的相互作用奠定了一定的理论基础。
Rv2628 protein is a latent infection-associated antigen regulated by Mycobacterium tuberculosis (M.tb) DosR. In this study, Rv2628 protein was prokaryotic expressed and purified. The macrophage cell line and mouse model were used to analyze the immunological characteristics of Rv2628 protein. The results of SDS-PAGE and Western blotting showed that Rv2628-His fusion protein was expressed in inclusion bodies and could react specifically with rabbit anti-H37Rv polyclonal antiserum with good immunoreactivity. The interaction with macrophage cell line RAW264.7 showed that Rv2628 protein induced the upregulation of pro-inflammatory factor IL-6 within 1-12 h. The purified Rv2628 fusion protein was immunized subcutaneously in BALB / c mice. The results of sandwich ELISA showed that the specific IFN-γ level induced by Rv2628 protein immunization group was significantly higher than that of IL-4 (P <0.000 1) Th1 type immune response trend; Rv262811-30 polypeptide as coating antigen, by indirect ELISA measured serum antibody titer can reach 1 1600, indicating that Rv2628 can induce humoral immune response. In summary, Rv2628 promotes the macrophage inflammatory response, stimulates the mouse to produce a strong Th1-type cellular immune response and a better humoral immune response, has the potential as a subunit vaccine, and serves as a The role has laid a theoretical foundation.