摘要
目的流式细胞术和乳酸脱氢酶(LDH)释放法检测自然杀伤细胞(natural killer cell,NK细胞)体外杀伤功能的比较和优化。方法以人外周血PBMCS经体外诱导扩增的NK细胞为效应细胞,K562细胞作为靶细胞,按5:1、10:1、20:1、40:1效靶细胞比共培养,分别采用LDH法和流式细胞术检测靶细胞死亡率。结果在4个效靶细胞比实验组中,5:1效靶细胞比条件下LDH释放法检测的靶细胞死亡率显著低于CFSE/PI标记法,其他3个效靶细胞比组的两种方法检测结果无显著性差异。高效靶比、细胞毒作用较强时,LDH释放法的结果误差较大,可重复性不强。结论流式细胞术和LDH释放法检测NK细胞体外杀伤效能的最佳效靶细胞比为10:1。流式细胞术比LDH法灵敏度更高,稳定性更好;在效靶细胞比为10:1的实验条件下,LDH法可作为流式细胞术的替代方法。
Objective To compare and optimize the detection process of cytotoxicity of natural killer cell(NK cell)in vitro by flow cytometry and lactate dehydrogenase(LDH)release assay.Methods NK cells induced by human peripheral blood PBMCS in vitro were used as effector cells,and K562 cells were used as target cells.The two cell lines were co-cultured with E/T ratios of 5:1,10:1,20:1 and 40:1,then the death rate of target cells was detected by LDH release assay and flow cytometry respectively.Results The mortality rate of target cells detected by LDH release method was significantly lower than that by flow cytometry under the E/T ratio of 5:1,while there was no significant difference between results of these two methods in the other three groups with E/T ratio of 10:1,20:1 and 40:1.When the E/T ratio is higher and the cytotoxicity is stronger,the results of LDH release method are not repeatable.Conclusion The optimal E/T ratio of flow cytometry and LDH release assay was 10:1.The sensitivity and consistency of flow cytometry is higher than that of LDH release assay,and LDH release assay can be used as an alternative for flow cytometry under the E/T ratio of 10:1.
作者
阳莉
丘秀生
胡元
陈晓燕
陈伟材
卢建溪
Yang Li;Qiu Xiusheng;Hu Yuan;Chen Xiaoyan;Chen Weicai;Lu Jianxi(Biotherapy Center,The Third Affiliated Hospital of Sun Yat-sen University,Guangzhou 510000,China;Vaccine Research Institute,The Third Affiliated Hospital of Sun Yat-sen University,Guangzhou 510000,China;The Department of Internal Medicine,Dongguan Chang’an Harbor Hospital,Dongguan 523000,China)
出处
《中国组织化学与细胞化学杂志》
CAS
CSCD
2021年第1期27-32,共6页
Chinese Journal of Histochemistry and Cytochemistry
