MTT Cell Viability Assay

The MTT assay was carried out in accordance with the supplier’s guidelines (Sigma-Aldrich, cat#M5655, St. Louis, MO, USA). Briefly, the culture medium was cautiously removed from each plated well following each treatment. Each well was then filled with 50 µL of serum-free medium and 50 µL of MTT^® solution, and the mixture was incubated for three hours at 37 °C. Each well received 150 µL of MTT solvent after incubation. For fifteen minutes, the foil-wrapped plate was shaken on an orbital shaker. Sometimes, the liquid needed to be pipetted in order to completely dissolve the MTT formazan. Within an hour, plate absorbance at OD = 590 nm was measured. In three separate experiments, the evaluation was conducted again. The percentage of cell viability was calculated using the following equation: % Viability = Mean OD(_{untreated sample}) − Mean OD_{(treated sample)}/Mean OD(_{untreated sample}) × 100. Samples were reported in comparison to the negative control (untreated wild-type ChLN cells) considered to have 100% viability.