Before sample collection, the cells were visually observed and photographed using an Axio Vert A1 Zeiss microscope with AxioCam MRm camera and Zen 2011 program (Carl Zeiss Microscopy GmbH, Jena, Germany). Thereafter, half (500 μL) of the cell culture medium was collected and centrifuged, as described above. LDH release was measured from medium samples immediately after the collection using the CytoTox96® Non-Radioactive Cytotoxicity Assay (Promega, Madison, WI, USA) according to the manufacturer’s instructions. Absorbance values were measured at a wavelength of 490 nm with a spectrophotometer BioTek, ELx808 and the Gen-5 2.04 program (Instruments Inc., Winooski, VT, USA).
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide salt (MTT; 0.5 mg/mL, Sigma-Aldrich, St. Louis, MO, USA) was added to cells remained on 12-well culture plates in the rest of medium (500 μL) and incubated for 3 h in the dark at +37 °C and 5% CO2, as described previously [15 (link)]. Absorbance values were measured at the wavelength of 560 nm using the spectrophotometer BioTek, ELx808.
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide salt (MTT; 0.5 mg/mL, Sigma-Aldrich, St. Louis, MO, USA) was added to cells remained on 12-well culture plates in the rest of medium (500 μL) and incubated for 3 h in the dark at +37 °C and 5% CO2, as described previously [15 (link)]. Absorbance values were measured at the wavelength of 560 nm using the spectrophotometer BioTek, ELx808.
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