Mitochondria and Cell Cycle
hms at shapirolab.com
Thu Nov 17 21:58:15 EST 2005
Dan Rosson wrote:
>I have a user that has been told by his grant reviewer to measure
>mitochondria as a function of cell cycle position. Someone suggested using
>Mito Red and YOYO-1. That didn't work out to well for two reasons: the cell
>cycle profile looked like garbage and while YOYO has an emission maximum
>around 510nM, it is so broad and intense it shows up in all channels
>including 675nM where its still impossible to compensate. The best
>suggestion I could come up with was to use formaldehyde-fixed cells with PI
>for cell cycle and to find an antibody to a mitochondrial protein (e.g.
>cytochrome c) and use FITC to detect that. Does anybody have any better
Is the idea to measure mitochondrial mass vs. cell cycle position or
mitochondrial membrane potential vs. cell cycle position - or both?
If the idea is to measure mitochondrial mass (not "number," because
you'd need some high resolution imaging to even hope to come up with
that), antibody to a mitochondrial protein is a good idea; I always
suspect "mitochondrial mass" measurements made with nonyl acridine
orange, MitoTracker dyes, etc., because their uptake is at least
partially potential dependent.
If the idea is to measure potential, you can't fix or permeabilize
the cells, and you'd have to use Hoechst 33342 or one of the newer
vital cell cycle dyes from Molecular Probes as the DNA stain. Low (1
nM) concentrations of DiOC6(3) (or DiIC1(5), if you have a red laser)
may be a better bet than JC-1.
More information about the Cytometry