Annexin v 7 aad apoptosis detection kit

Eca109 and EC9706, two human ESCC cell lines, were obtained from the Chinese Academy of Sciences Cell Bank (Shanghai, China). Dulbecco modified Eagle medium (DMEM; Gibco, Carlsbad, CA, USA) supplemented with 10% fetal bovine serum (Sijichun Bioengineering Materials Inc., Zhejiang, China) was used as the cell culture medium. For some studies, SH (Sigma-Aldrich Co., St. Louis, MO, USA) was dissolved in DMEM to achieve a concentration of 10 mM. Cell cultures were housed at 37°C in a humidified incubator with 5% CO₂. The CCK-8 kits and cell cycle analysis kits were acquired from Beyotime Institute of Biotechnology (Jiangsu, China). Annexin V-7AAD apoptosis-detection kits were purchased from BD Biosciences (Franklin Lakes, NJ, USA).

According to the manufacturer’s instructions, the cells were washed with PBS and the apoptosis was performed using flow cytometric analyses with Annexin V: 7-AAD Apoptosis Detection Kits (BD Biosciences, USA). After incubation, the samples were analyzed using flow cytometry (FACSCalibur, BD Biosciences, San Jose, CA). All samples were assayed in triplicate.