Re: options of vital dyes for Hoechst DNA / Pyronin Y RNA on cytometers without UV laser

Stephen Hou wrote:

> I have a user who wanted to sort quiescent stem cells based on Hoechst
> DNA / Pyronin Y RNA analysis. However, our instrument does not have UV
> laser. Do we have other vital dyes available for measuring DNA/RNA
> contents? 

If you happen to have a violet laser. you could try Molecular
Probes/Invitrogen's Vybrant DyeCycle Violet in place of Hoechst 33342;
if you have a red laser, DRAQ5 might work (I have heard that it does,
but have not verified this). There is now a 488 nm-excited green
fluorescent RNA-selective dye, SYTO RNASelect, from Molecular Probes,
that might work better than pyronin Y, but I haven't heard of anyone
trying to use it in combination with Hoechst 33342 as I used pyronin Y.

With 488 excitation alone, there is a possibility that you could get DNA
and RNA content using Vybrant DyeCycle Orange and SYTO RNASelect,
measuring the former above 600 nm and the latter at 525 or so with a 30
nm bandwidth. You'd probably have to fiddle with concentrations and
possibly need some compensation, but it's worth trying, meaning I would
have done it if I were still playing such games with mammalian cells.

Vybrant DyeCycle Violet can be used to demonstrate side populaion (SP)
cells and is therefore presumably a substrate for the ABCG2 transporter.
Whether the other DyeCycle dyes or SYTO RNASelect might be, I don't know.

Season's Greetings to All,

-Howard