【摘要】 目的:纯化可刺激人γδ+ T细胞增殖的结核杆菌耐热多肽抗 原。方法:采用快速蛋白液相色谱仪(FPLC)S-100分子筛层析柱,对结核杆菌耐热抗原(Mtb-Ag)初步分离。将获得的结核杆菌耐热Mr低的多肽抗原(Mtb-LW-Ag)洗脱峰,分别再经FPLC Mono Q离子交换层析柱进一步纯化,并且采用流式细胞仪对获得的Mtb-LW-Ag进行刺激γδ+ T细胞增殖活性的测定。结果:Mtb-Ag经FPLC S-100柱可分离出1个大分子蛋白峰A和3个M,低的多肽峰(B,C,D)。Mr低的多肽峰分别经Mono Q柱层析,鉴定出B峰含有6个主峰,C峰含有1个主峰,D峰含有8个主峰。对Mtb-LW-Ag及其纯化多肽进行活性检测,发现多肽峰B和C以及纯化多肽B-Ⅲ和C-主肽均可显著刺激γδ+ T细胞扩增。结论:利用FPLC法可快速高效地从Mtb-Ag中纯化出多种Mr低的多肽,而且其中的B-Ⅲ多肽和C-主肽可能是促进γδ+ T细胞活化增殖的主要多肽。更多还原

【Abstract】 AIM: To purify Mycobacterium tuberculosis heat-resistant peptide antigen ( Mtb-Ag) that can stimulate γδ8+ T cells. METHODS: Mtb-Ag was first separated by fast performance liquid chromatography (FPLC) with S-100 column, and then the components including low molecular weight peptide antigens (Mtb-LW-Ag) peaks were purified by FPLC with Mono Q column. Activity of the purified Mtb-LW-Ag that stimulates human γδ8+ T cell proliferation was examined by flow cytometry. RESULTS: Mtb-Ag was separated into four peaks(A, B, C and D) by FPLC S-100 column and the peak B, C and D contained Mtb-LW-Ag. The peak B was further separated into six main peaks( B- I ~ VI), peak C only one main peak (C-main), and peak D eight main peaks(D- I -VIII) by FPLC Mono Q column. Furthermore, peak B, C, B-III and C-main peptide could significantly stimulate human γδ8 + T cell proliferation in a dose of 0.1 mg/L. CONCLUSION; Varied Mtb-LW-Ag peptides are purified from Mtb-Ag by FPLC, and the B-III and C-main peptides may be the main components of stimulating human γδ8+ T cell proliferation.更多还原