Publisher Summary This chapter focuses on the experimental procedures designed to measure apoptosis-associated mitochondrial membrane permeabilization, either in intact cells or in isolated mitochondria. Following a variety of death signals, mitochondria show early alterations in their function that at least in some instances may be explained by the opening of the permeability transition (PT) pore. The PT pore—also called mitochondrial megachannel or multiple conductance channels—is a dynamic multiprotein complex located at the contact site between the inner and the outer mitochondrial membranes—one of the critical sites of metabolic coordination among the cytosol, the mitochondrial intermembrane space, and the matrix. Mitochondrial inner membrane depolarization is one of the major alterations observed in apoptotic cells. Accurate quantitation of the mitochondrial inner transmembrane potential (Δψm), using appropriate potentiometric fluorochromes, reveals a Δψm decrease in most models of apoptosis. The chapter proposes several protocols for quantitation of Δψm and other mitochondrial alterations in purified mitochondria, as well as in intact cells. Several different cationic fluorochromes can be employed to measure mitochondrial transmembrane potentials. Mitochondrial alterations in intact cells and cytometric analysis.