3.4. MTT Cell Viability Assay

Cell viability was assessed using an MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide)-based cytotoxicity assay. HEK293 cells were seeded in 10⁴ cells/well in 96-well plates and allowed to adhere for 24 h prior to compound treatment. Cells were treated at various concentrations in 96-well plates and incubated for 24 h. The final concentration of DMSO in the culture medium was maintained at 0.05% to avoid solvent toxicity. Subsequently, 20 μL of the 2 mg/mL MTT solution was added to each well of the plate and incubated for 2 h. Then, the absorbance was measured at 570 nm using a Versamax microplate reader (Molecular Devices Corporation, Sunnyvale, CA, USA). The percentage of cell viability is expressed as inversely proportional to the toxicities of the compounds, meaning that the higher the toxicity, the lower the cell viability. Cell viability is defined as the absorbance in the experimental well compared to that in the DMSO control wells.