4.12. Apoptosis Assay

EA was evaluated for its apoptosis-inducing potential on selected cancer cell lines using Annexin-V staining [61]. In brief, cells were plated in a six-well culture plate, and once they reached ~70%–80% confluency, incubated with IC₅₀ concentration of EA or vehicle control for 24 h at 37 °C. Following the incubation with EA (24 h), cells were collected, washed two times with PBS, and incubated with Annexin-V/PI stain and analyzed by Flow Cytometry as per manufacturer’s protocol (Annexin-V kit, BD-Biosciences, San Jose, USA).