Cell Cycle Analysis of PC-3 Cells

According to the procedure previously outlined by Alqahtani et al. [48 (link)] and Nasr et al. [49 (link)], the cell cycle distribution was evaluated. PC-3 cells were cultured for 24 h after being treated with B. aegyptiaca extract at the previously estimated IC₅₀ (92 µg/mL). The cells were then removed, thrice rinsed in cold PBS, fixed in cold ethanol (70%), and kept at 4 °C for four hours. PBS was used to rehydrate the fixed cells followed by the addition of RNase A (100 g/mL) and propidium iodide (100 g/mL, Abcam, Boston, MA, USA) for DNA staining. Using BD FACSCalibur flow cytometer with CellQuest software (BD Biosciences, San Diego, CA, USA), the DNA content was calculated after 30 min of incubation. propidium iodide fluorescence intensity was collected on FL₂ of a flow cytometer and 488 nm laser excitation.