Cell Viability Measurement by MTT Assay

Cell viability status was measured with the colorimetric MTT assay kit (ab211091, Abcam, Cambridge, MA, USA), according to the manufacturer’s protocol. Briefly, cells from the eighth-day chicken embryo femoral muscle were seeded into 96-well plate (Nest Scientific, Rahway, NJ, USA) at a density of 15 × 10³ in 100 μL of medium per well in eight replicates per each group treatment. Then, cells were incubated with experimental factors (L-Glu, nGO) for 48 h, in standard conditions (at 37 °C in a humidified atmosphere containing 5% CO₂). After that time, the growth medium was removed from the wells and cells were incubated with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reagent for 3 h at 37 °C. After incubation, formazan crystals were dissolved in MTT solvent and incubated for 15 min. The absorbance of samples (n = 6) was measured at 570 nm on a microplate reader, Infinite^® 200 PRO microplate reader with i-control™ software (Tecan Group Ltd., Männedorf, Germany). Calculations were performed as described by Strojny et al. [53 (link)].