Apoptosis Analysis by Flow Cytometry

Flow cytometry was performed as described previously (Wang et al., 2018 (link)). Cell apoptosis was analyzed by Annexin V-FITC/PI apoptosis detection kit according to the manufacturer’s protocol (Sigma-Aldrich Co. St. Louis, MO). Briefly, cells (CNE-2, H1299, and HSF) were seeded in 6-well plates. After 24 h of culture, cells were treated with CKI, IR, or IR combined with CKI, and then incubated at 37°C for 24 h. Afterwards, cells were collected, centrifuged for 5 min at 1,500 rpm, and resuspended in 1 × binding buffer. Finally, 5 μL Annexin V-FITC and 5 μL PI were added into the cells at room temperature for 15 min. The cells were then analyzed using flow cytometer (Beckman FC 500, Beckman Coulter, Inc., CA, United States).