Fluorescence-based Mitochondrial and ROS Assays

Fluorescence-based measurements of mitochondrial membrane potential and intracellular ROS generation were performed as previously described 50 (link). In brief, following treatment, hepatocytes were incubated with the potentiometric JC-1 sensor (Mitochondrial Membrane Potential Detection Kit, #30001, Biotium). For ROS measurements, fresh liver sections or primary hepatocytes were incubated with the cellular ROS indicator DHE (#PD-MY 003, MedChemExpress), cytoplasmic ROS indicator H2DCFDA (#HY-D0940, MedChemExpress), and mitochondrial ROS indicator MitoSOX RED (HY-D1055, MedChemExpress) based on methods provided by manufacturers 51 (link). Fluorescence images were acquired with same laser output and gain for control and treated samples on a Zeiss 880 confocal microscope using a 100X oil objective 52 (link).