Can you just bleach out the RFP with strong monochromatic light? According to Tsien's paper (http://www.tsienlab.ucsd.edu/Publications/Shaner%202005%20Nature%20Methods%20-%20Choosing%20fluorescent%20proteins.pdf), dsRED have very low photostability.
-Dave
On Friday, November 21, 2008 11:55:09 AM UTC+1, Confused Dave wrote:
> Hello,
>> I'm looking for a way to reliably inactivate fluorescent proteins
> (EGFP and dsRed) without harming epitopes for immuno.
>> A little background: I'm electroporating inducible constructs into
> the chick embryo. We use dsRed as the electroporation control, and
> EGFP as a marker for activation of our doxicyclin-inducible construct.
> We want to use immunofluorescence to on these samples. We currently
> use 4% formalin to fix, washes with PBS+tween, usually ethanol
> dehydration for storage, and sucrose-agar embedding for sectioning,
> and the fluorescence still comes up strong. Obviously, imaging with
> epifluorescence with a green and red channel, we can't add another
> fluorophore.
>> Best case would be a way to specifically kill the RFP leaving the
> green intact, although as long as we can recover the GFP with an
> antibody, this isn't a problem. Also looking for something that won't
> damage our epitopes to badly!
>> Thanks,
> Dave
