three cube FRET on a spectral confocal

[Cromey, Douglas W - (dcromey)]

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We have a person here interested in doing three cube FRET on our newest spectral confocal. I am aware of this technique, which as I understand it is usually done in widefield fluorescence with (wait for it) three filter cubes. Can (should) it be done on a confocal?

References, tips/pointers, or cautions would be appreciated. I've done a little acceptor photobleaching type FRET with users (sometimes even successfully!), but not a lot. Our concern is that the user has not used a confocal or ever done FRET before...

Doug

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Douglas W. Cromey, M.S. - Associate Scientific Investigator
Dept. of Cellular & Molecular Medicine, University of Arizona
1501 N. Campbell Ave, Tucson, AZ  85724-5044 USA

office:  LSN 463              email: [hidden email]<mailto:[hidden email]>
voice:  520-626-2824       fax:  [phone]

http://swehsc.pharmacy.arizona.edu/micro
Home of: "Microscopy and Imaging Resources on the WWW"

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[Periasamy, Ammasi (ap3t)]

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Here is few papers on confocal Spectral FRET.
Hope this helps
Ammasi


Chen, Y. J.P. Mauldin, R.N. Day and Periasamy, A. (2007) Characterization of spectral FRET imaging microscopy for monitoring the nuclear protein interactions. J Microscopy, 228:139-152.

Sun, Y. Booker, C.F., Kumari, S., Day, R.N., Davdison, M. and Periasamy, A. (2009) Characterization of an Orange Acceptor Fluorescent Protein for Sensitized Spectral FRET Microscopy using a White Light Laser. J. Biomed. Opt. 14(5), 054009.

Sun, Y., Wallrabe, H. Booker, C., Day, R.N. and Periasamy, A. (2010) Three-color spectral FRET microscopy localizes three interacting proteins in living cells. Biophysical J. Vol. 99, 1274-1283.

Dr. Ammasi Periasamy
Center Director, W.M. Keck Center for Cellular Imaging
Professor of Biology and Biomedical Engg.
Univeristy of Virginia, Charlottesville
http://www.kcci.virginia.edu/people/profile/ap3t
Phone: [phone] or 982-4869
Fax: [phone]
E-mail: [hidden email]
Office and Center Location
(90 Geldard Dr., At the intersection of Geldard dr. and White Head Rd.),
Physical and Life Sciences Building (PLSB)
FRET/FLIM Workshop-March 6-10, 2017: http://www.kcci.virginia.edu/workshop 

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Cromey, Douglas W - (dcromey)
Sent: Friday, August 05, 2016 11:38 AM
To: [hidden email]
Subject: three cube FRET on a spectral confocal

*****
To join, leave or search the confocal microscopy listserv, go to:
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*****

We have a person here interested in doing three cube FRET on our newest spectral confocal. I am aware of this technique, which as I understand it is usually done in widefield fluorescence with (wait for it) three filter cubes. Can (should) it be done on a confocal?

References, tips/pointers, or cautions would be appreciated. I've done a little acceptor photobleaching type FRET with users (sometimes even successfully!), but not a lot. Our concern is that the user has not used a confocal or ever done FRET before...

Doug

------------------------------------------------------------------------------------------
Douglas W. Cromey, M.S. - Associate Scientific Investigator Dept. of Cellular & Molecular Medicine, University of Arizona
1501 N. Campbell Ave, Tucson, AZ  85724-5044 USA

office:  LSN 463              email: [hidden email]<mailto:[hidden email]>
voice:  520-626-2824       fax:  [phone]

http://swehsc.pharmacy.arizona.edu/micro
Home of: "Microscopy and Imaging Resources on the WWW"

UA Microscopy Alliance -  http://microscopy.arizona.edu<http://microscopy.arizona.edu/>