流式细胞术检测小鼠中性粒细胞吞噬功能的方法学探讨

目的: 建立用流式细胞术检测小鼠中性粒细胞(PMN)吞噬功能的方法。方法: 用不同缓冲液制备荧光素(FITC)标记葡萄球菌或大肠埃希菌,流式细胞术检测细菌FITC标记率。小鼠全血与FITC标记细菌37℃共育10~20min,溶解红细胞后用流式细胞术检测小鼠PMN荧光强度的变化,计算PMN吞噬率。结果: 用PBS(pH7.2)标记金黄色葡萄球菌效果好,用碳酸盐缓冲液(pH9)标记大肠埃希菌效果好。小鼠PMN对金黄色葡萄球菌的吞噬率低,对大肠埃希菌的吞噬率高。结论: 用流式细胞术检测小鼠PMN吞噬FITC标记大肠埃希菌是测定小鼠PMN吞噬功能的简便、快速和重复性好的方法。

A methodology for detection of phagocytosis of murine neutrophilsby by flow cytometry

Objective: To establish the method for detection of phagocytosis of muirne neutrophils by flow cytometry.Methods: Staphylococcus aureus (S.aureus) and Escherichia coli (E.coli) were labeled with fluorescein isothiocyanate(FITC) in different buffers.The efficient rate of labeling was assessed by flow cytometry.Whole blood taken from mice were incubated with FITC labeled bacteria at 37℃ for 10-20 min,followed by lysing red blood cells.The fluorescent intensity of neutrophils was measured as phagocytotic rate by flow cytometry.Results: The good labeling of S.aureus and E.coli were obtained in PBS(pH 7.2) and carbonate buffer(pH 9),respectively.The percentage of phagocytosis of E.coli was higher than that of S.aureus by murine neutrophils.Conclusions: Detection of phagocytosis of FITC labeled E.coli by flow cytometry is simple,rapid and reproducible method for measuring phagocytotic activity of murine neutrophils.