3.5. Cellular Viability Assay—MTT Test

The cancer cells were seeded in 24-well plates, with 1 mL of cell suspension per well, in the following densities: A2780—10⁴ cells/mL; A2780cis—1.5 × 10⁴ cells/mL; and Toledo and Toledo-cis—5 × 10⁴ cells/mL. Lymphocytes were seeded in 48-well plates, 0.5 mL of cell suspension per well, at −2 × 10⁵ cells/mL. The cultures were left overnight to adapt. The next day, the tested compounds were added, pre-dissolved in DMF, and then in a complete culture medium, at concentrations two-fold higher than the final ones. The resulting solutions were added to cell cultures to obtain the demanded concentration. Stock solutions of the compounds were freshly made before every experiment. The final concentration of the solvents in culture medium was 0.2% v/v. After 72 h, MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) solution was added to PBS (final concentration 0.25 mg/mL in culture) and incubated at 37 °C for 2–3 h. Then the culture medium was removed, and the purple formazan crystals were dissolved in DMSO. The absorbance was measured at 540 nm on a Spectrostar Nano spectrophotometer with a microplate reader (BMG Labtech, Ortenberg, Germany). The experiments were performed at least in triplicate. The results were transformed to relative values, as the percentage of the solvent control (assumed as 100%) to subtract any toxic effect of DMF.