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目的:制备和初步鉴定抗结核分枝杆菌抗原优势肽段的单克隆抗体。方法:以两种优势肽段的融合抗原PGEX-4T-2/38kD-ESAT6-CFP10和PGEX-4T-2/Mtb8.4-MPT64-Mtb16.3-Mtb8作为免疫原免疫BALB/c小鼠,取其脾细胞与SP2/0小鼠骨髓瘤细胞融合后,通过有限稀释法筛选阳性克隆,经ELISA和Western blotting分析单抗的特性和特异性。结果:获得17株结核分枝杆菌抗原优势肽段特异单克隆抗体杂交瘤细胞株,经间接ELISA方法测定,杂交瘤细胞培养上清效价为28～212,接种小鼠的腹水效价为216～220。Western blotting结果显示,每株单抗均能特异性的识别融合蛋白的优势肽段。结论:制备了针对抗结核分枝杆菌抗原优势肽段的单克隆抗体,为结核分枝杆菌的快速诊断和抗原性分析奠定了基础。
OBJECTIVE: To prepare and preliminarily identify monoclonal antibodies against Mycobacterium tuberculosis antigens. METHODS: BALB / c mice were immunized with the two dominant peptides, PGEX-4T-2 / 38kD-ESAT6-CFP10 and PGEX-4T-2 / Mtb8.4-MPT64-Mtb16.3- After spleen cells were fused with SP2 / 0 mouse myeloma cells, the positive clones were screened by limiting dilution, and the specificity and specificity of McAbs were analyzed by ELISA and Western blotting. Results: 17 hybridoma cell lines with specific monoclonal antibody against Mycobacterium tuberculosis antigen were obtained. The titer of hybridoma cell culture supernatant was 28 ~ 212 as determined by indirect ELISA. The titer of ascites in inoculated mice was 216 ~ 220. Western blotting showed that each monoclonal antibody could specifically recognize the dominant peptide of the fusion protein. Conclusion: Monoclonal antibodies against Mycobacterium tuberculosis antigens have been prepared for the rapid diagnosis and antigenic analysis of Mycobacterium tuberculosis.