摘要
流式分选术是对细胞或生物颗粒进行无菌筛分的技术,因具有检测速度快、通量高、灵敏度高等优势,故在医学及生命科学领域有广阔的应用前景。离心是流式分选后富集细胞的常用技术,直接影响流式分选后样本的细胞活性和得率。该文以KPC细胞系(即小鼠胰腺组织特异性kras缺失p53突变形成胰腺癌分离的癌细胞,简称KPC)和小鼠原代脾脏细胞为例,利用FACS SORP Aria II分选仪进行分选。对离心力、离心时间等参数进行优化,结果表明KPC细胞分选后,300×g、4°C离心25 min,能获得高得率高活性的细胞用于下游实验,而小鼠原代脾脏细胞则300×g、4°C离心15 min最为合适。该方法解决了流式分选后细胞离心难、离心损失大的问题,可提高分选实验实际得率,为流式分选术的应用提供技术参考。
Flow cytometry is a kind of aseptic screening technology for cells or biological particles.Due to its advantages such as high speed,high flux,high sensitivity,etc,which has broad applications in medicine and life sciences.Centrifugation is a common technique for cell enrichment after flow cell sorting,which directly affects the cell viability and yield of samples sorted by flow cytometry.In this paper,based on FACS SORP Aria II cell sorter,KPC cell line(i.e.mouse pancreatic tissue-specific kras deletion p53 mutant cancer cells isolated from pancreatic cancer,KPC)and mouse primary spleen cells are sorted.The experimental conditions such as centrifugal time and centrifugal force are optimized.The results show that KPC cells with high yield and viability can be obtained by centrifugation at 300×g,for 25 min at 4°C,while for mouse primary spleen cells,centrifugation at 300×g,for 15 min at 4°C is the most appropriate.This method can solve the problems of difficult centrifugation and large centrifugation loss,then improve the actual yield of cell sorting experiments and provide technical reference for the application of flow cytometry.
作者
黄莹莹
邢月婷
孟晓瑜
桑文华
李艳伟
宋兴辉
郭春
王佳佳
HUANG Yingying;XING Yueting;MENG Xiaoyu;SANG Wenhua;LI Yanwei;SONG Xinghui;GUO Chun;WANG Jiajia(Core Facilities,School of Medicine,Zhejiang University,Hangzhou 310058,China;School of Basic Medical Sciences,Zhejiang University,Hangzhou 310058,China)
出处
《中国细胞生物学学报》
CAS
CSCD
2021年第10期2001-2008,共8页
Chinese Journal of Cell Biology
基金
浙江省教育厅一般科研项目(批准号:Y202044997)
中央高校基本科研业务费专项资金(青年科研创新专项)(批准号:2021QNA7018)资助的课题。
关键词
流式分选
离心时间
离心力
得率
细胞活性
flow cell sorting
centrifugal time
centrifugal force
yield
cell viability
