Mitochondria Structure

Mitochondria are found in most eukaryotic cells, where they make up as much as 10% of the cell volume. The mitochondria structure is a double-membrane that consists of an outer membrane and an inner membrane called a cristae that extends into the inner matrix [1]. Mitochondria are pleomorphic organelles, with structural variations depending on cell type, cell cycle stage, and intracellular metabolic state. The key function of mitochondria is energy production through oxidative phosphorylation and lipid oxidation. However, mitochondria are involved in various other metabolic functions including urea production, steroid biogenesis, and calcium homeostasis.

Learn more about additional mitochondria stains

MitoTracker probes are cell-permeant mitochondria stains that contain a mildly thiol-reactive chloromethyl moiety. Following incubation, MitoTracker dyes passively diffuse across the plasma membrane and accumulate in the mitochondria of live cells. These dyes are offered in a range of wavelengths that can all be used for mitochondrial localization in multicolor experiments (Figures 1–3).

Signal retention of the MitoTracker probes is variable after cell fixation. MitoTracker Green and MitoTracker Red lose signal, while MitoTracker Orange CMTMRos, MitoTracker Red CMXRos, and MitoTracker Deep Red are well retained. Cells stained with MitoTracker Deep Red can be subsequently permeabilized and still retain its staining pattern.

MitoTracker Orange CM-H₂TMRos and MitoTracker Red CM-H₂XRos are additional mitochondrial stains. These probes do not fluoresce until they enter live cells, where they are oxidized to become fluorescent and then are sequestered in the mitochondria.

Learn more about mitochondrial functional assays

CellLight fluorescent fusion proteins can be used to stain mitochondria and follow the dynamics of mitochondria behavior in live cells independent of membrane potential. Thus, they can be used in combination with MitoTracker dyes to investigate relationships between mitochondria morphology and membrane potential. CellLight Mitochondria-GFP (Figure 4) and CellLight Mitochondria-RFP (Figure 5) are ready-to-use constructs that use the leader sequence of E1 alpha-pyruvate dehydrogenase (3.1 kDa) to target mitochondria.

Introducing CellLight fluorescent fusion proteins involves a simple transfection step, using the BacMam technology, and they work like cell stains with minimal toxicity or chemical disruption. These mitochondrial fusion proteins are compatible with other fluorescent probes for multiplex analysis in live cells, or after formaldehyde fixation for colocalization studies.

Learn more about these and other CellLight fluorescent reagents

1. Cooper GM (2000). The Cell: A Molecular Approach. 2nd edition. Sunderland (MA): Sinauer Associates; 2000. Mitochondria.