The mitochondrion is different from other organelles in the eukaryotic cell because it possesses its own genome that exists in the form of circular DNA. Due to this unusual characteristic, mitochondria reproduce independently of the cells in which they are found and are widely considered to have evolved from free-living aerobic eubacteria that developed an endosymbiotic relationship with primitive anaerobic eukaryotic cells. From this association, the eubacteria would have gained shelter and nutrients from the larger organism, which in return received power from the smaller cell in the form of ATP molecules. The ancient partnership, which apparently led to the modern relationship between the eukaryotic cell and the mitochondrion, is estimated to have begun about 1.5 billion years ago when oxygen became a significant part of the Earth's atmosphere. Interestingly, unlike nuclear DNA, mitochondrial DNA is not shuffled every generation, so it changes at a slower rate, a fact that has resulted in its use for a wide range of applications, especially the study of human evolution.

The popular dye combination of MitoTracker Red CMXRos, Alexa Fluor 488 conjugated to phalloidin, and Hoechst 33342 were utilized to stain an adherent culture of Madin-Darby ovine kidney epithelial cells, which is illustrated above. This triple probe regimen labels the tubular mitochondria, filamentous actin network, and DNA in the nucleus. Images were recorded in grayscale with a QImaging Retiga Fast-EXi camera system coupled to an Olympus BX-51 microscope equipped with bandpass emission fluorescence filter optical blocks provided by Omega Optical. During the processing stage, individual image channels were pseudocolored with RGB values corresponding to each of the fluorophore emission spectral profiles.