Fluorescent probes are currently used to evaluate the mitochondrial transmembrane potential in situ, However, in parallel experiments using the probes JC-1 and TMRM in different cell types (human astrocytes, HEp-2, Vero, KB, and HeLa tells), we found that the distribution of JC-1 and TMRM is highly variable not only in different tell types but also in different cells of the same cell type, a condition that has never been documented until our work, This phenomenon depends on a hidden, widespread multidrug resistance (MDR) phenotype that can be recognized only by comparative assays with MDR inhibitors (progesterone, verapamil, and cyclosporin A) and represents a serious risk of error in the evaluation of the mitochondrial potential.