Protocol for investigating mitochondrial structure, function, and metabolism in human cervical cancer cells

Mitochondria regulate a variety of biological activities, including metabolism, oxidative stress, and cell death. Here, we present a protocol for the investigation of mitochondrial structure, function, and metabolism in human cervical cancer cells. We describe steps for staining and visualizing mitochondria using confocal microscopy to assess morphology, mass, membrane potential, calcium, reactive oxygen species (ROS), and lipid droplet accumulation. We then detail procedures for isolating mitochondria and performing metabolomic analysis of mitochondrial metabolites via mass spectrometry. For complete details on the use and execution of this protocol, please refer to Adiga et al.¹.