论文部分内容阅读
目的鉴定日本血吸虫22.6 kDa膜蛋白(Sj22.6)的Th1型表位,为构建短肽疫苗奠定基础。方法采用合成肽Sj22.6-P4、无关肽(对照)及PBS免疫C57BL/6小鼠2次(间隔7 d),末次免疫后7~10 d取脾分离单个核细胞,用合成肽Sj22.6-P4、无关肽及PBS刺激培养,3H-TdR掺入法检测淋巴细胞的增殖效果,酶联免疫吸附试验(ELISA)检测其细胞培养上清中IFNγ-、IL-4及IL-2水平。运用流式细胞技术三色标记法检测经合成肽Sj22.6-P4、无关肽及PBS免疫2次的C57BL/6小鼠脾淋巴细胞内的Th1、Th2细胞因子。结果合成肽Sj22.6-P4可刺激经该抗原肽免疫2次的C57BL/6小鼠淋巴细胞增殖,与PBS组相比,增殖指数(SI)均>2。与无关肽对照组相比,细胞培养上清中IL-2、IFN-γ分泌水平增高,其中IL-2分泌水平差异有统计学意义(P<0.05),IFN-γ差异无统计学意义(P>0.05),而IL-4分泌水平明显降低(P<0.05)。合成肽Sj22.6-P4免疫组小鼠脾脏CD4+T细胞中分泌IFN-γ细胞的百分比显著增高,分泌IL-4细胞的百分比显著降低(P均<0.05)。结论合成肽Sj22.6-P4是C57BL/6小鼠特异的Th1型表位。
Objective To identify the Th1 epitope of Schistosoma japonicum 22.6 kDa membrane protein (Sj22.6) and lay the foundation for the construction of short peptide vaccine. Methods C57BL / 6 mice were immunized with synthetic peptide Sj22.6-P4, unrelated peptide (control) and PBS twice daily for 7 days. Mononuclear cells were isolated from the spleens 7-10 days after the last immunization and the synthetic peptide Sj22 was used. 6-P4, irrelevant peptide and PBS. 3H-TdR incorporation was used to detect the proliferation of lymphocytes. The levels of IFNγ-, IL-4 and IL-2 in the cell culture supernatants were detected by enzyme linked immunosorbent assay (ELISA) . The Th1 and Th2 cytokines in splenic lymphocytes of C57BL / 6 mice immunized twice with synthetic peptide Sj22.6-P4, unrelated peptide and PBS were detected by flow cytometry. Results The synthetic peptide Sj22.6-P4 stimulated the proliferation of lymphocytes of C57BL / 6 mice immunized twice with the antigenic peptide. The proliferation index (SI) was> 2 compared with PBS group. The levels of IL-2 and IFN-γ in the cell culture supernatant were higher than those in the non-peptide control group, with a significant difference in the level of IL-2 secretion (P <0.05) and no significant difference in IFN-γ P> 0.05), while IL-4 secretion was significantly decreased (P <0.05). The percentage of secreting IFN-γ cells in splenic CD4 + T cells and the percentage of IL-4 secreting cells in Sj22.6-P4 immunized mice were significantly decreased (all P <0.05). Conclusion The synthetic peptide Sj22.6-P4 is a Th1 type epitope specific to C57BL / 6 mice.