Is it important to dilute Dihydroethidium, DHE; fluorescent probe for ROS measurement, in a oxygen free atmosphere, as argon or nitrogen? Are there any other considerations?
I guess that it could improve the shelf-life of DHE if you dissolve it under oxygen-free conditions. Although in general I would not recommend to store them for a long time. I typically prepare them fresh before doing my experiment.
You should also consider using high-quality anhydrous DMSO, to ensure a very low content of moisture that could otherwise react with your probe.
Do you prepare DHE with out oxygen-free condition?
Do you warm DHE everytime you need to use it?
I store DHE in aliquots of 25 uL at 10mM , thus I defrost only once to make a working solution at 25 uM for cell loading. To be safe I try not to use them after a ~ 3 month storage. My DMSO isn't too high-quality and for me, after measuring PROBE integrity it is suffice to say my experiments won't be effected by DHE irregularity. I am measuring ROS, so I do need them to be consistent. However, Jeroen is right about all that. However, the ones i obtain from Sigma are somewhat robust granted you are careful.
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