Re: Best FRET pair

Posted by Vitaly Boyko on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Best-FRET-pair-tp4804703p4805626.html

Dear Ammasi,

Is that so important? CeFP-YFP is a great pair, though the quantum yield of the CeFP could be better, and it works better than the YFP-mRFP1 pair for example. Though I did not have a chance to compare that to GFP-mOrange or GFP-mRFP1. YFP-mRFP1 pair works quite good, nearly ten-fold better than the YFP-mCherry one. Having an optimal filter set is very important!, e.g. to keep the donor/FRET bleedthrough much below 40%.

Vitaly

301-515-7833 

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From: "Periasamy, Ammasi (ap3t)" <[hidden email]>
To: [hidden email]
Sent: Fri, March 26, 2010 1:54:49 PM
Subject: Re: Best FRET pair

YFP-mOrange should work for you
We are using GFP-mOrange as FRET pair. You should have a better quantum yield for the donor compared to acceptor.
Ammasi


-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Maria Calvo
Sent: Friday, March 26, 2010 11:48 AM
To: [hidden email]
Subject: Best FRET pair

Dear all,

We are planning to do FRET experiments with a fluorescent protein (YFP, mCherry or mOrange)and a directly labelled primary antibody.

We are restricted to YFP, mCherry or mOrange for one of the proteins (the most abundant).

Does anyone know what's the best FRET pair for any of these FPs, in terms of J (overlap), brightness ?

Thanks for your help,

Maria Calvo

--
Ammasi Periasamy, Ph.D.
Director, Keck Center for Cellular Imaging (KCCI)
Professor of Biology and Biomedical Engineering
Biology, Gilmer Hall (064), 485 McCormick Rd
University of Virginia
Charlottesville, VA 22904
Voice: 434-243-7602 (Office); 982-4869 (lab)
Fax:[phone]; Email:[hidden email]
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