4.6. Cell Viability Determination Using MTT Assay

The cytotoxic activity of novel synthesized compounds (PtMet2 and PtMet2–PAMAM) against human breast cancer cell lines (MCF-7 and MDA-MB-231) and normal human breast epithelial cells (MCF-10A) was analyzed by a spectrocolorimetric assay using thiazolyl blue tetrazolium bromide (MTT) in accordance with the procedure in the literature [9]. Briefly, MCF-7, MDA-MB-231, and MCF-10A cells were seeded in six-well plates “Nunc” at a density of 5 × 10⁵ cells/well and incubated for 24 h in optimal growth conditions (37 °C, 5% CO₂). Subsequently, PtMet2, PtMet2–PAMAM, and the reference drug (cisplatin) at concentrations 0.1, 0.25, 0.5, 1.0, 1.5, 2.5, and 5.0 µM were added in duplicate, and the plates were incubated for another 24 h. Next, the plates were washed with PBS three times. Then, 1 mL PBS and 50 μL of 5 mg/cm³ MTT solution were added, and the incubation was continued for four hours. The MTT assay protocol is based on the enzymatic reduction of yellow soluble tetrazolium bromide to blue formazan, which occurs only in living cells. Then, the product is dissolved in DMSO, and the absorbance is measured at 570 nm wavelength. The absorbance result obtained in the control was taken as 100%, and the viability of the cells incubated with the tested compounds was shown as a percentage of the control cells. The analysis was performed using the Evolution 201 UV-visable spectrophotometer using the Thermo INSIGHT software (Thermo Fisher, Waltham, MA, USA, both).