IC50 and Cell Viability Assays

IC₅₀ and cell viability assays were measured using a 3-(4,5-dimethylthiazole-yl)-2,5-diphenyltetrazolium bromide (MTT, Sigma-Aldrich) assay. IC₅₀ assay: Cells in the logarithmic growth stage were trypsinized, neutralized, and then plated at a density of 5×10³ cells per well in 96-well plates in 200 µL diluted media.

Quercetin was dissolved in DMSO at a concentration of 100 mM and then diluted to the appropriate dose (0, 6.25, 12.5, 25, 50, 100, or 200 µM; DMSO was used as a control) in the corresponding complete DMEM. Following the standard protocol, the 96-well plates were assessed at 24, 48, 72, and 96 hours by measuring the absorbance at 490 nm. Based on the data, we calculated the inhibition rate, performed curve fitting, and obtained the IC₅₀ value.

Cell viability assay: Cells were plated at a density of 1–2×10³ cells per well in 96-well plates in 200 µL diluted quercetin media. The plates were tested on days 1, 3, 5, and 7 by measuring the absorbance at 490 nm. Cell growth curves were generated to measure cell proliferation.