Six different secretory proteins of molecular weights (15, 26, 30, 41, 55 and 70 kDa) were isolated from 8-day-old culture filtrate of Mycobacterium tuberculosis H37Ra using different column chromatography techniques. These proteins were further examined for their ability to induce cell mediated (T-cell proliferation assay) and humoral immune response (ELISA) in mice immunized with total culture filtrate proteins. Out of six proteins, three proteins showed good reactivity. However, the activity was at a maximum with 30 kDa antigen. The immune response induced by 30 kDa antigen emulsified in Freund's incomplete adjuvant (FIA) was investigated and was found to be dose dependent. The T-cell response induced by this protein was skewed towards T-helper (Th1) cells as determined by the pronounced secretion of interleukin-2 (IL-2) and gamma-interferon (IFN-gamma). The protective activity of the 30 kDa protein was also evaluated and compared with reference to Bacillus Calmette Guerin (BCG) vaccine in the mice challenged with virulent M. tuberculosis H37Rv. The degree of protection afforded by the 30 kDa antigen on the basis of mortality and the significant decrease in c.f.u.'s recovered from different organs (lung, liver, spleen) after 30 days of challenge with LD50 of M. tuberculosis H37Rv was significantly higher in comparison to BCG vaccinated animals. However, the degree of immunity induced by this antigen decreased with time (when challenged 8 and 12 weeks post-immunization) but it was still comparable with BCG. These findings suggest that 30 kDa secretory protein of M. tuberculosis is the key immunoprotective antigen and may be a suitable candidate for the development of an alternative subunit vaccine against tuberculosis.

译文

:使用不同的柱色谱技术从结核分枝杆菌H37Ra的8天龄培养滤液中分离出六个分子量分别为15、26、30、41、55和70 kDa的分泌蛋白。在用总培养滤液蛋白免疫的小鼠中,进一步检查了这些蛋白诱导细胞介导的能力(T细胞增殖测定)和体液免疫应答(ELISA)。在六种蛋白质中,三种蛋白质显示出良好的反应性。但是,对于30kDa抗原,活性最大。研究了在弗氏不完全佐剂(FIA)中乳化的30 kDa抗原诱导的免疫反应,发现该反应是剂量依赖性的。该蛋白诱导的T细胞反应偏向T辅助(Th1)细胞,这由白介素2(IL-2)和γ-干扰素(IFN-γ)的明显分泌确定。还评估了30 kDa蛋白的保护活性,并与卡介苗芽孢杆菌(BCG)疫苗进行了比较,比较了在有毒结核分枝杆菌H37Rv攻击的小鼠中的行为。 30 kDa抗原提供的保护程度基于死亡率以及结核分枝杆菌H37Rv的LD50攻击30天后从不同器官(肺,肝,脾)回收的cfu显着降低。与接种卡介苗的动物比较。但是,这种抗原诱导的免疫度随时间降低(在免疫后8周和12周激发时),但仍可与BCG相提并论。这些发现表明,结核分枝杆菌的30kDa分泌蛋白是关键的免疫保护性抗原,并且可能是开发替代性抗结核亚单位疫苗的合适候选者。

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