Visualization of mitochondrial DNA replication in individual cells by EdU signal
Authors Haines KM, Feldman EL, Lentz SI
Submitted By Eva Feldman on 8/1/2012
Status Published
Journal Journal of visualized experiments : JoVE
Year 2010
Date Published 5/1/2010
Volume : Pages Not Specified : Not Specified
PubMed Reference 21113116
Abstract Mitochondria are key regulators of cellular energy and mitochondrial biogenesis
is an essential component of regulating mitochondria numbers in healthy cells.
One approach for monitoring mitochondrial biogenesis is to measure the rate of
mitochondrial DNA (mtDNA) replication. We developed a sensitive technique to
label newly synthesized mtDNA in individual cells in order to study mtDNA
biogenesis. The technique combines the incorporation of
5-ethynyl-2'-deoxyuridine (EdU) with a tyramide signal amplification (TSA)
protocol to visualize mtDNA replication within subcellular compartments of
neurons. EdU is superior to other thymidine analogs, such as
5-bromo-2-deoxyuridine (BrdU), because the initial click reaction to label EdU
does not require the harsh acid treatments or enzyme digests that are required
for exposing the BrdU epitope. The milder labeling of EdU allows for direct
comparison of its incorporation with other cellular markers. The ability to
visualize and quantify mtDNA biogenesis provides an essential tool for
investigating the mechanisms used to regulate mitochondrial biogenesis and would
provide insight into the pathogenesis associated with drug toxicity, aging,
cancer and neurodegenerative diseases. Our technique is applicable to sensory
neurons as well as other cell types. The use of this technique to measure mtDNA
biogenesis has significant implications in furthering the understanding of both
normal cellular physiology as well as impaired disease states.

Investigators with authorship
Eva FeldmanUniversity of Michigan