Zenyth 200rt microplate reader

THC-stimulated changes in the mitochondrial membrane potential (MMP) were assessed using the fluorescent reagent tetraethylbenzimidazolylcarbocyanine iodide (JC-1) with the JC-1-Mitochondrial Membrane Potential Assay Kit (Abcam, Cat. no. ab113850) following the manufacturer’s protocol. Cells were washed once with 1× dilution buffer and then incubated with 20 µM JC-1 dye in 1× dilution buffer for 10 min at 37 °C, protected from light. JC-1 dye was then removed, cells were washed once with 1× dilution buffer, and 100 µL of fresh 1× dilution buffer was added to each well. The red fluorescence in excitation (535 nm)/emission (590 nm) and green fluorescence excitation/emission (475 nm/530 nm) was measured using a Biochrom Anthos Zenyth 200 rt Microplate Reader (Cambridge, UK). Background fluorescence was subtracted from the fluorescence of treated cells, then the ratio of red (polarized) fluorescence divided by that of green (depolarized) fluorescence was obtained.