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2–1 × 106 cells were harvested per condition, fixed in 1 mL ice-cold 70% EtOH, and kept at −20°C for at least 2 h and up to 2 weeks. Fixed cells were washed two times with PBS + 2% FBS and spun down at 700 × g for 10 min after each washing. Cells were stained during 1 h at 37°C with 5 μg/mL Hoechst 33342 (BD Biosciences, San Jose, CA) in PBS + 2% FBS at a final volume of 250 μL to 1 mL, depending on the number of cells.
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