Determination of Cell Viability Using MTT Assay

To determine the effect of phenethyl isothiocyanate on normal human keratinocyte, HaCaT cells and cervical cancer cells (CaSki and HeLa) were analyzed using MTT assay. Briefly, the cells were seeded into a 96-well plate at a density of 5 × 10³ cells per well in triplicate and incubated at 37°C for 24 h. The cells were treated with different concentrations of PEITC (0, 2, 5, 10, 15, 20, and 25 µM) for 24 and 48 h. Control group cells were treated with 0.01% DMSO in media. Subsequently, cell survival was determined by adding 10 µL of 5 mg/ml MTT reagent in each well, followed by incubation for 4 h at 37°C, and then 100 µL of dimethyl sulfoxide was added to dissolve formazan by gentle shaking. The absorbance of each well was measured at 570 nm, and cell survival was calculated as percentage over the control. Additionally, cisplatin was also used to determine and compare cytotoxicity on the cell lines, either alone or in combination with the tested compound. The purpose of including HaCaT cells (normal keratinocyte cell line) is to compare the cytotoxic effect of phenethyl isothiocyanate.