基于实时无标记技术研究流式分选后细胞活性的检测方法  

Study on a Cell Viability Detection Method after Flow Cytometric Sorting Based on Real-time Labeling-free Technique

作　　者：李艳伟[1] 王佳佳[1] 邢月婷 郭春 黄莹莹[1] 宋兴辉[1] 黄琼[1] LI Yan-wei;WANG Jia-jia;XING Yue-ting;GUO Chun;HUANG Ying-ying;SONG Xing-hui;HUANG Qiong(Core Facilities,School of Medicine,Zhejiang University,Hangzhou 310058,China)

机构地区：[1]浙江大学医学院公共技术平台,浙江杭州310058

出　　处：《分析测试学报》2022年第7期1084-1089,共6页Journal of Instrumental Analysis

基　　金：浙江省教育厅项目(Y201941524);浙江大学实验技术研究项目(SYB202130)。

摘　　要：利用流式细胞法、细胞计数试剂盒(CCK-8)法、实时细胞功能分析(RTCA)法分别检测分选前后细胞的活性,综合比较了3种方法的检测原理、操作特点等。结果显示,流式细胞法测得的分选后细胞凋亡率为(3.85±0.008)%,分选前细胞凋亡率为(14.09±0.021)%,分选后细胞凋亡率低于分选前,具有统计学差异(P<0.05);分选前后的活细胞比例显示,分选后活细胞比例高于分选前,具有显著性统计学差异(P<0.001)。CCK-8法测得的分选后细胞活性高于分选前,具有显著性统计学差异(P<0.001);RTCA法测得的分选后细胞连续增殖活性明显高于分选前,具有极显著性统计学差异(P<0.0001)。结果显示,相较于CCK-8法和流式细胞法,RTCA法的细胞用量少、可回收,操作简便,无需标记,检测灵敏度高,可获得实时动态的细胞生长曲线,适合研究分选后细胞长效动态的活性变化,可为基础和临床分选后活细胞的功能研究提供检测手段。In this paper,flow cytometric analysis,cell counting kit-8(CCK-8)and real time cellular analysis(RTCA)were used to detect cell viability before and after flow cytometric sorting,respectively.The experimental operation and statistical analysis of the three methods were compared comprehensively.The result showed that the apoptosis percentages for cells determined by flow cytometric analysis were(3.85±0.008)%in the group after sorting and(14.09±0.021)%in the group before sorting.And the apoptosis percentage for cells after sorting was lower than that before sorting,with a statistical difference(P)less than 0.05.Further statistical analysis for the proportion of living cells before and after sorting showed that the proportion of living cells after sorting was higher than that before sorting,with a significant statistical difference(P)less than 0.001.The CCK-8 assay showed that the group after sorting had higher cell proliferation viability than the group before sorting(P<0.001).RTCA assay showed a highly significant statistical difference(P<0.0001)in the continuous proliferation activity of the cells after sorting.Compared to the assays of flow cytometric analysis and CCK-8,RTCA method has a low request of cell numbers,recyclable,easy operation,labeling-free,high detection sensitivity,which displays a real-time dynamic cell growth curve suitable for studying the long-term dynamic changes of cell viability after sorting.Furthermore,RTCA could be used as a method for the functional studies of cells after basic and clinical sorting.

关 键 词：实时细胞功能分析 流式细胞分析 流式细胞分选 CCK-8 细胞活性 

分 类 号：O615.4[理学—无机化学] Q256[理学—化学]