Hello everyone
I want to investigate mitochondrial membrane potential changes over a 48-hr period in cancer cell lines
. All the dyes employed (tmrm, mitotracker cmxros etc) have been reported to have some toxic effects on mitochondria if used for over some hours. Does anyone have protocol for confocal monitoring with mitotracker cmxros ?
Yes, I've seen the toxic effects of MitoTrackers. In egregious cases (such as using too high to concentration), you'll even see the mitochondria swell into "donut" shapes then burst.
TMRM is better, probably because it lacks the chloromethyl group and thus doesn't mind to proteins and is reversible. We've done some timelapses out to several hours, but I think stretching it to days would be too much. You'd have to test it.
I don't know of any mt potential dyes that can be visualized for as long as 48 hours.
I strongly agree with Jason. You find details about use of TMRM in Ward et al 2000 in Journal of Neuroscience and Dussmann et all 2003 in Journal of Cell Science. TMRM has the lowest partition coefficient in membranes compared to rh123 and TMRE but still acts like a photosensitizer. Keep the concentrations around 30nM for Cancer cell lines and 10nM if you work with neurons, keep the excitation light intensity and exposure time low, keep the imaging frequency low as well. We used TMRM successfully for more than 72h (Laussmann et al 2011, CDD). If you need more advice you might want to give details about the imaging setup and the cells you intend to use.
Best
Heiko
Vad Pérez Not that I've seen. That was just my personal observation.
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