Determination of mitochondrial membrane potential

Cells were cultured as indicated and mitochondrial membrane potential was determined by cell permeable JC-1 dye. JC-1 selectively enters the mitochondria and reversibly changes from green to red as the membrane potential increases. In healthy cells with normal ΔψM, JC-1 spontaneously forms J-aggregates with intense red fluorescence. Low ΔψM in apoptotic cells maintains JC-1 in the monomeric form and only shows green fluorescence. JC-1 fluorescence images of cells exposed to Celastrol, with or without NAC pretreatment, were captured using an epifluorescence Nikon microscope (Nikon, Japan).