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>> I have used TO-PRO-3 as live dead discriminator with red
>> excitation. Note that the -3 is important, Molecular Probes
>> also sells TO-PRO-1! For TP3, saturating concentration is 2-4 uM.
> I'll say.....esp. in view of what follows. How much DNA stain do
> people want to whack about, on the reasonable asumption that that
> which binds to DNA may not be frightfully good for it?
To clarify, by saturating I meant "I use it to get best CVs when staining
nuclei or fixed and permabilized cells for DNA content/cell cycle"
> Out of interest, what is here the 'gold standard'? We in Aber are
> beginning to have suspicions that supposedly pure(ish) TO-PRO[3],
> or likely a metabolite / degradation product thereof, penetrates (and
> ergo stains) ""live"" [i.e. not all that knackered] cells (bugs).
> What say you, and netters to whom this is copied?
I mention in passing in Cytometry 21:265 that TO-PRO-3 enters apoptotic
thymocytes more rapidly than the normal cells in the same sample. PI and EB
are well know to behave the same. While "apoptotic" may or may not mean
something to bugs, I would hesitate to suggest problems with TO-PRO-3 unless
I knew that PI and EB do not show the same effect. You may, of course,
know this.
This does suggest my opinion of a gold standard. TO-PRO-3 is only available
from Molecular Probes, and should be compared to similar molecules like ethidium
and propidium which have long histories and multiple sources.
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