Apoptosis Detection in Cancer Cells

Apoptosis assays were performed on the cancerous cell lines to test whether or not 1 induced apoptosis or necrosis. Apoptosis detection was determined by flow cytometry (Cytomics FC 500 Series Beckman Coulter) using annexin V-FITC/PI kit (Beckman Coulter, Miami, FL) in combination with PI (essentially as previously described [32 (link)]. Untreated cells and cells treated with 1μM curcumin, 1μM EF-24 (Sigma, 300 μM H₂O₂, and 1% v/v DMSO (all reagents from Sigma-Aldrich, St. Louis, MO, USA) were included as controls and incubated for a total of 24 h. FITC (excitation/emission maxima of 495/519 nm) and PI (excitation/emission maxima of 493/636 nm) emitting green and red signals, respectively, were excited with a 20 mW argon ion laser operation at 488 nm and their ensuing fluorescence signals were captured by using FL1 and FL2 detectors, respectively. Data acquisition and analysis was performed by using CXP software (Beckman Coulter).