Cyan adp cell analyzer

Flow cytometry analysis was performed as described previously [61 (link)]. Briefly, H2B-GFP-labelled cells were washed and resuspended in PBS buffer for flow cytometry analysis. Cells used for PI staining were first fixed in ice-cold 70% ethanol overnight at 4°C. The fixed cells were washed, resuspended in 0.5 ml of NS buffer (10 mM Tris-HCl [pH=7.2], 0.25 M sucrose, 1 mM EDTA, 1 mM MgCl₂, 0.1 mM ZnCl₂, 0.4 mM phenylmethylsulfonyl fluoride, 7 mM β-mercaptoethanol). RNase A (0.5 mg/ml) and propidium iodide (10 μg/ml) were added into the suspension and incubated for 2 h at 37°C or overnight at 4°C in the dark. The cells were sonicated for 10 s before analysis with a CyAn ADP cell analyzer (Beckman Coulter, Hialeah, FL) or BD Acuri™ C6 flow cytometer (BD Biosciences). We noted that different cell analyzers may yield FACS patterns with subtle differences. Profiles obtained from the same cell analyzer should be used for direct comparison. Data were analyzed with FlowJo software (Treestar, Inc., Ashland, OR, USA).