To all of those who have contributed (or are planning to contribute)
to this thread:
In view of the fact you seem to get different results concerning the
brightness of DsRed compared to other FPs, would you mind describing
your imaging setup (excitation source, filters, detector/camera
model)? This would help a lot to get a better picture of the
situation.
TIA,
Beat
>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,
>respectively.
>> > 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.
>>Frank
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