2.12. Sulforhodamine B (SRB) Assay

HEK293A, MCF10A, MDA-MB-231, and BT549 cells were seeded into each well of a 96-well plate. After incubation for 24 h, the cells were treated with DMSO and 100 μg/mL of PLE for up to 5 days; the cell medium with serially diluted PLE was refreshed every 2 days. The plates were harvested every 2 days at the same time and fixed with 10% trichloroacetic acid (TCA) solution (Sigma-Aldrich) at 4°C overnight. The plates were washed with distilled water more than three times. After that, the plates were stained with 0.4% SRB (Sigma-Aldrich) in 1% acetic acid solution for 30 min at 25°C. The cells were then washed with 0.1% acetic acid solution and dried at 25°C for 1 h. Finally, 10 mM Trizma base (Sigma-Aldrich) was added, and the plates were incubated on a rocker for 1 h to resolve the stained cells. The SRB levels were measured using a microplate spectrophotometer (BioTek Instruments) at an absorbance of 540 nm.