gfp spectral shifts?

Kevin Holmes KHOLMES at niaid.nih.gov
Tue Aug 13 15:45:48 EST 2002


Marty,
Interesting question.  When we have performed FRET experiments using YFP and
CFP, we've noticed that when performing compensation between CFP and the
detector used to collect FRET emission from YFP, it can be difficult to
correctly compensate both the bright and dull CFP+ cells.  That is, if the
bright +'s  were correctly compensated, the dull+'s were overcompensated; if
the dull+'s were correctly compensated the bright+'s were undercompensated.
This effect was seen in CFP+ only cells.  My thought has been that there is
a change in the emission spectrum of CFP dependent upon concentration and/or
some kind of spectral quenching effect occurring.
Unfortunately, I don't have any firm data to confirm the above hypothesis.
Kevin

Kevin L. Holmes, Ph.D.
Chief, Flow Cytometry Section
Research Technologies Branch
Bldg. 4, Room B1-38
NIAID, NIH

Phone: 301-496-9071
FAX:  301-402-4532
Email: kholmes at niaid.nih.gov

-----Original Message-----
From: Marty Bigos [mailto:mbigos at gladstone.ucsf.edu]
Sent: Monday, August 12, 2002 10:47 AM
To: cyto-inbox
Subject: gfp spectral shifts?


Hi -

Does anyone know of shifts in the emission spectra of the various gfp
constructs due to intracellular pH, linkage to other proteins, etc.?
Thanks.


Marty
--
Marty Bigos
Director, Flow Core
Gladstone Institute of Virology and Immunology
Building 3 SFGH Rm 509
415-695-3832
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