4.2. Fluorescence Labeling of Mitochondria

To label the mitochondria, fibroblast cells were incubated with MEM NEAA basal medium (Thermo Fisher Scientific) containing 150 nM Mitotracker Red CM-H2Xros (Invitrogen, Waltham, MA, USA) for 30 min. In the FCCP treatment group, fibroblast cells were incubated with 0.7 µM FCCP for 30 min before the addition of the Mitotracker Red CM-H2Xros. At the end of the incubation period, the cells were washed 3 times with prewarmed Dulbecco’s phosphate-buffered saline (dPBS). The nucleus was stained by further incubating cells with basal medium containing Nucblue Hoechst (Invitrogen) for 15 min. Following this incubation, cells were washed several times with prewarmed dPBS to remove excess dye. At the end of the wash, MEM NEAA basal medium was added to each dish prior to image acquisition.