2.6. Determination of Cytotoxicity Using MTT Assay

Human colorectal adenocarcinoma (HT-29), human colon carcinoma (HCT-116), human cervical cancer (HeLa), human gastric adenocarcinoma (HGT-1), human hepatocellular carcinoma (HepG2), and mouse fibroblast (BALB/c 3T3) cells were seeded in 96-well plates (1 × 10⁵ cells/well) and incubated overnight. Phy-CS-MNP nanocomposite (1.56–00 μg/mL) was added into the 96 wells of HT-29, HCT-116, HeLa, HGT-1, HepG2, and BALB/c 3T3 cells. Untreated BALB/c 3T3 and cancer cell lines were included. Following 24 h, 48 h, and 72 h of incubation, 20 µL (5 mg/mL) of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was added into each well followed by 2–4 h incubation. Lastly, an aliquot of 100 µL of dimethyl sulfoxide was added into each well. The absorbance was read at 570 nm using ELISA microplate reader (Tecan, Männedorf, Switzerland). The wavelength of 630 nm was used as a reference.