Cell proliferation evaluation by the CCK-8 assay

Cells in the logarithmic growth phase were collected. Digestion of the cells was performed using 0.25% trypsin, followed by cell suspension preparation. We adjusted the cell density to 1 × 10⁵ cells/ml. Thereafter, the cells were plated in 96-well plates (200 μl/well). The experiments were performed in 5 replicate wells. The cells were grown for 24 h in an incubator programmed at 37 °C, 5% CO2, and saturated humidity. After the cells were allowed to grow and adhere for 24 h, the drug solution was added to the cells at various concentrations. CCK-8 reagent (10 μl) was added to each well, the samples were thoroughly mixed, and the plates were placed in an incubator for 2 h. Absorbance values (OD values) were determined at 450 nm using an enzyme standard instrument and used in the following formula:cell growth inhibition rate = (test OD value − blank OD value)/(control OD value − blank OD value) × 100%. Bar graphs of the final value were generated, and the results were analyzed. The experiment was replicated three times, and the results were averaged.