Mitochondrial Membrane Potential Assay in SH-SY5Y Cells

The mitochondrial membrane potential assay was carried out using a JC-10 kit (Abcam, CA, USA) [31 (link)]. SH-SY5Y cells were seeded in 96-well plates and pretreated with various concentrations of GH (dissolved in PBS). After incubation for 1 h at 37°C, scopolamine was cotreated with GH for 24 h at 37°C. After treatment, the cell mitochondria were stained using 50 μL of JC-10 solution for 30 min at 37°C and kept away from light. After incubation, 50 μL of buffer B solution was added into the JC-10 loading plate before reading the fluorescence intensity, and this was analyzed using a microplate reader (BioTek Instruments Inc., Winooski, VT, USA). The fluorescence intensities were measured at Ex/Em = 490/525 nm and 490/590 nm for ratio analysis.