Mitochondria and Cell Cycle

Howard Shapiro 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
>suggestions?

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.

-Howard





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