Re: best FRET pair _2012 update

[offterdi1]

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Dear all
Thanks a lot for your input, which is very much appreciated, indeed.
What surprises me a bit, though, is the focus on the CFP-YFP system.
One big problem with that pair has not yet been discussed at all: The generation
of a CFP-like photoproduct in acceptor bleaching experiments. I have seen this
effect myself and it was very strong! So are there any further comments on this
or improved YFP version that don't photoconvert to CFP-like emitting species...?

Thanks again

[Arne Seitz]

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Hi Martin,

in the following paper we also quantified the photoconversion for different YFP constructs. Citrine and Venus also showed photoconversion but reduced by a factor of around six compared to the eYFP construct.

Quantifying the influence of yellow fluorescent protein photoconversion on acceptor photobleaching-based fluorescence resonance energy transfer measurements
J. Biomed. Opt. 17, 011010 (Feb 13, 2012);

http://spiedigitallibrary.org/jbo/resource/1/jbopfo/v17/i1/p011010_s1


Cheers Arne

P.S. Sorry for the shameless self-promotion.


---------------------------------------------------------------
Arne Seitz
Head of Bioimaging and Optics Platform (PT-BIOP)
Ecole Polytechnique Fédérale de Lausanne (EPFL)
Faculty of Life Sciences
Station 15, AI 0241
CH-1015 Lausanne

Phone: [phone]
Fax:      [phone]
http://biop.epfl.ch/
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> -----Original Message-----
> From: Confocal Microscopy List
> [mailto:[hidden email]] On Behalf Of Martin
> Offterdinger
> Sent: lundi 6 août 2012 09:32
> To: [hidden email]
> Subject: Re: best FRET pair _2012 update
>
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Dear all
> Thanks a lot for your input, which is very much appreciated, indeed.
> What surprises me a bit, though, is the focus on the CFP-YFP system.
> One big problem with that pair has not yet been discussed at all: The
> generation of a CFP-like photoproduct in acceptor bleaching experiments. I
> have seen this effect myself and it was very strong! So are there any further
> comments on this or improved YFP version that don't photoconvert to CFP-
> like emitting species...?
>
> Thanks again

[Martin Spitaler]

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Dear Martin,

 Re: non-CFP-like FRET pairs: in our (and many other people's) hands ,
GFP-mCherry works very well: It's based on the most optimised FPs of all
(GFP) - monomeric, single-exponential decay, bright, photostable, ..... - ,
efficiency is good yet the spectra nicely separated for stimulated emission.

Best,

Martin
###############################
Martin Spitaler, PhD
FILM - Facility for Imaging by Light Microscopy
- Facility Manager -
Sir Alexander Fleming Building, desk 401
Imperial College London / South Kensington
Exhibition Road
London SW7 2AZ
UK

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Website: http://imperial.ac.uk/imagingfacility 

On Mon, 6 Aug 2012 02:31:43 -0500, Martin Offterdinger
<[hidden email]> wrote:
>
>Dear all
>Thanks a lot for your input, which is very much appreciated, indeed.
>What surprises me a bit, though, is the focus on the CFP-YFP system.
>One big problem with that pair has not yet been discussed at all: The
generation
>of a CFP-like photoproduct in acceptor bleaching experiments. I have seen this
>effect myself and it was very strong! So are there any further comments on
this
>or improved YFP version that don't photoconvert to CFP-like emitting
species...?
>
>Thanks again