2.7. Cell Cycle Analysis

Cell cycles were determined by using a Muse™ cell cycle assay kit according to the manufacturer’s protocols. Cells were seeded at a concentration of 5 × 10⁴ cells/well and cultured for 24 h. After culturing, the cells were treated with sanggenol L (10, 20, and 30 µM) for 48 h. The cells were fixed in ice-cold 70% ethanol at 4 °C for 24 h. After resuspension, the cells were added 200 μL of Muse™ cell cycle reagent and incubated for 30 min at room temperature. Cells were analyzed using a Muse™ cell analyzer (Merck KGaA, Darmstadt, Germany).