2.12. RNA Level Detection by Pyronin Y Staining

Cells were resuspended in Hank's solution, washed two times, and fixed in cold methanol : acetone (4 : 1) for 30 min at 4°C. The samples were then washed twice and incubated with 0.5 μg/ml pyronin Y for 30 min at 37°C. Before analysis by the BD FACSCalibur flow cytometer, cells were transferred into ice for at least 10 min. Pyronin Y is excited at 488 nm, emits at 575 nm, and analyzed in a linear scale.