TUNEL assay
The measurement of cell viability was also correlated with DNA damage using TUNEL assay as per the manufacturer's protocol (GenScript, Piscataway, NJ, USA). Briefly, 2 × 105 cells per well were seeded in 12-well plates and the cells were exposed to the treatments with appropriate compounds. Upon treatments, the cells were air-dried for 5 min and fixed (4% paraformaldehyde in PBS) for 30 min at room temperature. After fixation, they were incubated overnight at 4 °C and permeabilized using 1% Triton X-100. The cells were then blocked in 3% H2O2 in methanol and labeled with TUNEL reaction mixture (biotin-11-dUTP and TdT) and streptavidin-FITC. The smear of cells were washed with PBS to remove excess of labeling reagents and incubated with POD-conjugated anti-FITC substrate. Finally, 3,3′-diaminobenzidine (DAB) working reagent was added and the cells were observed under the light microscope. The images were taken using Leica DMI 300B (Leica Microsystems Inc., Buffalo Grove, IL, USA).