2.12. Cell Cycle Analysis by Flow Cytometry

The cells were grown in 24-well plate for 24 h to study the effect of colchicine on cell cycle and were then treated with colchicine of different concentrations (0, 2.5, 5, 10, 20, 40, and 60 nM). The cells were then incubated overnight at 37 °C in a humidified atmosphere of 5% CO₂. The cells treated with colchicine were harvested and fixed in ice cold 70% ethanolin ice, for 1 h. Then, the cells were incubated for 30 min at 37 °C in a PBS solution containing 1 mg/mL RNaseA and 1 µg/mL propidium iodide (PI) solution. The cell cycle analysis was performed using Amnis Image Stream flow cytometer (Millipore, United States) and the data were analyzed based on 8000 events recorded using IDEAS image Stream Analysis Software.