4.5. Cytotoxicity Assay

The MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay is a colorimetric assay to measure cellular metabolic activity as an indicator of cell viability, proliferation and cytotoxicity. J744A.1 cells were seeded in a 96-well plate with concentration 10⁴ cells/100μL/well and incubated for 24 h. After the medium was removed, 100 μL fresh medium containing peptide (ranging from 64 μg/mL to 2 μg/mL) was added to the wells. Following 24 h incubation, fresh medium with MTT (0.5 mg/mL) was replaced and incubated for 3 h. After the medium/MTT was removed, DMSO was added at 100 μL for dissolving the formazan crystal. Cell survival rate was calculated by measuring the absorbance at 540 nm using a Tecan Sunrise microplate reader (Tecan, Männedorf, Switzerland). Medium without peptide and mixed with H₂O_2(aq) represented positive and negative controls, respectively.