Jc 10 fluorometric assay kit

Changes in ΔΨm were analyzed using a JC-10 fluorometric assay kit (MAK159 Sigma-Aldrich, St. Louis, MO, USA) according to the manufacturer's protocol. In brief, JC-10 dye loading solution (final concentration 2 μg/mL) was added to HPMEC that were incubated in the dark for 45 min, at 37°C. Fluorescence signal was detected using the iBright Imaging System. The ratio of fluorescence intensities at 590 nm/525 nm was used to determine mitochondrial membrane depolarization.

MMP was determined using JC-10 fluorometric assay kit (Sigma-Aldrich Ltd., St. Louis, MO, USA) according to the manufacturer’s instructions. MCF-7 cells were seeded in a 96-well plate (1.5 × 10 ⁴/well) in triplicates following treatment with Que, Nar or CoQN at 44.31 µg/mL, respectively, for 24 h. After that, 50 µL of JC-10 loading dye was added to each well and incubated for 30 min in the dark. Red and green fluorescence were then measured using a microplate reader at an excitation/emission wavelength of 540/590 and 490/525, respectively. The ratio of red to green fluorescence was calculated to determine the MMP. Results were expressed as a percentage of control of the red/green fluorescence ratio.