Mitochondria are often depicted as static, cylinder-shaped organelles, but time-lapse studies reveal that the important energy generators may change shape and move about the cytoplasm on an almost continuous basis. These activities often appear to involve the cytoskeletal microtubules, which influence the characteristic direction and dissemination of the mitochondria in various kinds of cells. The number of mitochondria present in a cell is related to the metabolic needs of that cell, and may range from a single large mitochondrion to thousands of the organelles. Their size generally ranges from 1 to 10 micrometers, making mitochondria large enough to be observed with a light microscope. The organelles were originally identified during the 1800s, and were commonly believed to transmit hereditary information until after the beginning of the twentieth century. Most of the modern understanding of the functional role of mitochondria did not develop until after a method for isolating the intact organelles was developed in 1948.

The popular triple fluorophore combination of MitoTracker Red CMXRos, BODIPY FL conjugated to phalloidin, and Hoechst 33258 was used to label the adherent log phase culture of HJ1.Ov cells presented above for mitochondria, the filamentous actin network, and nuclear DNA. The cells were first treated with MitoTracker Red CMXRos in growth medium for one hour, washed and fixed with paraformaldehyde (prepared in growth medium), permeabilized, and blocked with bovine serum albumen. The cells were subsequently labeled with the conjugated phalloidin and counterstained with the bisbenzimide reagent. 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.