流式细胞术检测人和小鼠Th细胞亚群

Flow cytometric detection of human and murine Th1/Th2 cells

目的 建立人和小鼠Th1和Th2细胞的检测方法 ,探讨两者在流式细胞仪检测方面的不同。方法 采用PMA和离子霉素共同刺激外周血单个核细胞 ,观察在不同时间和不同剂量的PMA刺激下外周血T淋巴细胞CD3、CD4、CD8表达的变化 ;采用多色荧光共同标记分析技术检测外周血单个核细胞IL 4和IFN γ的表达。结果 在PMA和离子霉素共同刺激后 4h ,人外周血T淋巴细胞表面CD4表达迅速下降 ,而小鼠T淋巴细胞表面CD4表达仅轻度下降 ;采用多色分析技术可以有效的检测Th1细胞和Th2细胞 ;采用CD3、CD8设门分析可以有效的区分人Th细胞 ;采用CD4设门可以有效区分小鼠Th细胞。结论 人Th细胞检测宜采用 4色染色分析技术 ,小鼠Th细胞检测采用 3色染色技术即可。

Objective To establish the method for flow cytometric detection of human and murine Th1/Th2 cells. Methods Peripheral mononuclear cells (PMNCs) were stimulated with phorbolmyristateacetate (PMA) and ionomycin. The expressions of CD3, CD4, and CD8 were monitored after stimulation with different concentrations of PMA for different time. A multiple color staining of the cells was used to detect the intracelluar expressions of IL-4 and IFN-γ. Results Expression of CD4 in human PMNCs decreased dramatically at 4 h after stimulation with PMA and ionomycin and Th cells could be distinguished from other cells using CD3 and CD8 dot plot gating, but expression of CD4 in mice PMNCs decreased slightly after stimulation with PMA and ionomycin and Th cells could be distinguished from other cells using CD4 histogram gating. Conclusion Our results suggest human Th1/Th2 cells should be detected by four-color staining method and murine Th1/Th2 cells by three-color staining method.