Frank O. Fackelmayer
Frank.Fackelmayer at uni-konstanz.de
Thu Feb 17 04:41:16 EST 2000
Jered Floyd wrote:
> MMILLING at MEDAU.JNJ.com ("MILLINGTON, Michelle [JJRAU]") writes:
> > We have tried EBFP but are unable to detect by flow due to low fluorescence
> > and photobleaching. Is dsRFP brighter and more resistant to photobleaching
> > than EBFP?
> With an epifluorescence microscope, DsRed is significantly brighter
> and much more resistant to photobleaching than GFP/EGFP;
We´ve also used RFP and EGFP, but with reverse result: EGFP is MUCH brighter the
RFP, at least in COS cells transfected with vectors dsRed-N1 and pEGFP-N1,
> I imagine as
> such it is much better than EBFP as well. I have not yet tried it
> with flow, but will soon.
> One word of warning with DsRed: At least in E. coli, there is a very
> large delay (36-48 hours) before it is visible to the eye. This is
> unfortunate, as the other FPs express visibly much faster.
Not only in E.coli. When looking at transfected cells, RFP is not detectable 24h
after transfection (when EGFP is already VERY bright), but shows a good signal
only after at least 48h. Seems the "maturation" of the protein - which makes it
fluorescent - takes much longer for RFP than for GFP. This is an important thing
to remember when doing cotransfections with RFP and GFP.
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