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细胞周期分析PI染色法

发布者:biolead 发布时间:2019-10-19 分享到:

细胞周期流式检测(PI染色法)

1. 收集细胞:若悬浮生长细胞离心弃培养基,用PBS洗涤一次(建议用PBS + 1% FBS或PBS + 0.1% BSA),若是贴壁生长细胞,加入合适体积的0.25%胰酶消化一分钟左右(可显微镜观察待细胞脱壁),吹起细胞后加入5ml 含10%血清的培养基,终止作用,转移细胞至15ml离心管中,300g 离心5分钟,用PBS洗涤细胞一次;

2. 用PBS洗涤细胞一次,300g离心5分钟后弃上清;

3. 用500ul PBS 重悬细胞,振荡混匀后加入5ml 70%冷乙醇溶液(放置在-20度预冷的70%乙醇溶液),混匀后放置4度过夜(可放置一周);

4. 500g离心5分钟后弃乙醇,用PBS悬浮细胞洗涤细胞一次;

5. 用PBS调节细胞浓度为5 X 106个/ml,取500ul细胞悬液加入100ul  500ug/ml 的PI染色(用PBS配制500ug/ml的PI染色液),4°C避光放置30分钟;

6. 加入100ul 煮沸过的RNase A (10mg/ml) 37度孵育30分钟;

7. 混匀细胞后待上流式仪获取分析。

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