Apoptosis detection by flow cytometry

SW620 and HT29 cells were incubated in 6-well plates for 48 hours with indicated concentrations of AE. After that, the cells were harvested, washed with PBS and fixed in 70% ethanol at 4°C for 30 min. Then the cells were exposed to 20 μg/mL RNase I and 50 μg/mL PI (Cell Signaling Technology, Danvers, MA, USA) for 30 min at 37°C. Flow cytometry was performed according to the manufacturer’s protocol. Apoptotic cells were assessed by flow cytometry on a FACScan (Beckman Instruments, Fullerton, CA, USA).