The JC10 dye measures mitochondrial membrane potential (MMP) which can be a measure of the viability of cells. However, I want to roughly assess OxPhos with JC10 (given that the MMP can be a measure of OxPhos activity), even though the literature (Abcam and Sigma JC10) only talks about detecting apoptosis. I want to use the BD Acuri C6 flow cytometer to carry out the assay. Am I mistaken? Maybe I have misunderstood the whole thing.
Thanks Mohit, however the paper doesnt answer my question unfortunately
Hi Phil,
MMP can be useful to determine mitochondrial health in cells, provided you have some control compounds you can add. If you have access to them, the best way to OxPhos activity is by measuring respiration rates in the presence and absence of oligomycin (Complex V inhibitor). When you add the oligomycin, you will be inhibiting ATP synthesis, and slowing respiration - the remaining respiration rate is caused by "leak" across your inner mitochondrial membrane. This will provide you with a crude estimation of your respiratory control ratio - or integrity of your inner membrane. If you don't have access to this equipment, you could try performing a titration with the protonophore FCCP - a little goes a long way, so you may want to start out with a top concentration of ~10 uM. comparing the sensitivity each of your experimental groups to FCCP could indicate changes in mitochondrial health between them.
Also, when using JC-10 or JC-1 be sure to use the probe ratiometrically, looking at the mean fluorescence intensity (J-aggregates:J-monomers) of each channel. Alternatively, you might want to try a non-ratiometric dye such as TMRE, or TMRM as these catatonic probes increase fluorescence as they concentrate into the mitochondrial matrix.
we actually offer a kit that may help you out...
https://www.caymanchem.com/product/701560
I hope that this helps
Thanks David. This definitely helps! I have FCCP to use as a control with JC-10- hope it works fine.
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