Apoptosis Induction in Colon Cancer Cells

HCT116 colon cancer cells were seeded in 24-well plates with a density of 1 × 10⁵ cells and incubated overnight before the treatment. The cells were treated with the IC50 concentration of dolasetron and ketoprofen and incubated for 48 h. After washing the control and drug-treated wells with PBS, the cells were stained with a mixture of acridine orange and ethidium bromide (100 µg/mL each). The stained cells were visualized and imaged immediately using an inverted fluorescent microscope, and images were captured using the Thermo Fisher scientific EVOS Floid imaging system [25 (link)]. For apoptosis, caspase-3/7activity was measured using the Apo-one Homogeneous Caspase-3/7 assay kit (Promega Madison, WI, USA; cat no. G7790), and the manufacturer’s protocol was followed. Cleavage of the non-fluorescent substrate, Z-DEVD-Rhodamine-110, by caspase 3/7 resulted in fluorescent rhodamine-110. The fluorescence of the sample was measured at 530 nm emission and 490 nm excitation in the multimode reader, BioTek [26 (link),27 (link)].