双色法和三色法检测NK细胞杀伤功能的实用性比较  

Comparison of two-color method and three-color method to detect the killing function of NK cell

作　　者：陈晓燕[1] 阳莉[1] 蔡惠宁[1] 李翠平[1] 卢建溪[1] CHEN Xiaoyan;YANG Li;CAI Huining;LI Cuiping;LU Jianxi(The Third Affiliated Hospital Of Sun Yat-sen University,Biological Treatment Center,Guangzhou,Guangdong,China,510000)

机构地区：[1]中山大学附属第三医院生物治疗中心科,广东广州510000

出　　处：《分子诊断与治疗杂志》2022年第11期1899-1904,共6页Journal of Molecular Diagnostics and Therapy

基　　金：广东省药品监督管理局科技创新项目(2021TDB22)。

摘　　要：目的 比较双色法和三色法检测自然杀伤细胞(NK cell)的体外杀伤功能。方法 选取20例自2019年至2021年于中山大学附属第三医院献血者外周血,体外诱导扩增NK细胞(效应细胞),在培养第14天按10∶1、20∶1效靶比分别与K562细胞(靶细胞)混合共培养,且在1 h、2 h、3 h、4 h共4个时间组取样,分别采用CFSE/PI双色法和CFSE/Annexin-V/7-AAD三色法进行流式染色,检测K562细胞的凋亡。结果 在不同效靶比实验组,两种染色方法都能检测出NK细胞在效靶比20∶1杀伤功能显著高于效靶比10∶1,差异有统计学意义(P<0.05),在4个时间点实验组中,效靶细胞共培养4 h时NK细胞杀伤功能最强。其中,三色法检测出细胞杀伤功能显著高于双色法,差异有统计学意义(P<0.05)。结论三色荧光染色法比双色法能更灵敏更特异地检测NK细胞杀伤功能,结果更为可靠。Objective To compare the killing function of natural killer cells(NK cells)in vitro by two-color method and three-color method. Methods Peripheral of 20 blood donors in our hospital from 2019 to2021was selected to induce and expand NK cells(effector cells)in vitro. On the 14th day of culture,NK cells(effector cells)were mixed and co cultured with K562 cells(target cells)according to the efficiency target ratio of 10∶1 and 20∶1,and samples were taken in four time groups of 1 h,2 h,3 h and 4 h. CFSE/PI two-color method and CFSE/annexin-V/7-AAD three-color method were used to detect the apoptosis of K562 cells.Results In the experimental groups with different effect target ratio,both staining methods can detect that the killing function of NK cells in the effect target ratio of 20∶1 is significantly higher than that in the effect target ratio of 10∶1(P<0.05). In the experimental groups at four time points,the killing function of NK cells is the strongest when the effect target cells are co cultured for 4H. Among them,the cell killing function detected by the three color method is significantly higher than that by the two-color method,and the difference is statistically significant(P<0.05). Conclusion Under the experimental conditions of effect target ratio of 20∶1and killing time of 3 hours,trichromatic fluorescence staining method can detect NK cell injury function more sensitively and specifically than bichromatic method,and the result is more reliable.

关 键 词：双色法 三色法 NK细胞 杀伤功能 

分 类 号：R363[医药卫生—病理学]