RE: GFP
Mayumi Naramura (MNARAMURA@atlas.niaid.nih.gov)
Tue, 25 Feb 1997 16:39:49 -0500
The excitation wavelength of the original GFP from jellyfish (wtGFP)
was not really suitable for 488 nm laser. Also, jellyfish codon usage
was not efficiently translated by mammalian cells. I remember there
was also some discussion on temperature sensitivity. But now there are
a lot of commercially available GFP mutants which were actually
selected for detection by flow cytometry. I have used pGreenLantern-1
from GIBCO BRL and pEGFP-1 from CLONTECH, and both worked very well in
flow. I compared pGreenLantern-1 with wtGFP in the same cell line, and
the difference in the mean fluorescence intensity was 2 to 3 log's.
Mayumi Naramura, M.D.
NIH/NIAID
Lab of Immunology
Twinbrook II, Rm. 125
12441 Parklawn Drive
Rockville, MD 20852
Phone: 301-402-4595
FAX: 301-594-2522
e-mail:mnaramura@atlas.niaid.nih.gov
----------
From: David L. Haviland, Ph.D.
Sent: Tuesday, February 25, 1997 11:29
To: cyto-inbox
Subject: GFP
Hi:
I was curious if people had been successfully analysed GFP levels
within
transfected cells using flow?
I once had an investigator try something like this (though I do *not*
know
if it was GFP, per se) and the experiment was a complete bust! There
was
something about the excitation/emmision that was not near
instantaneous and
there was enough delay that no fluoresence could be detected except
when
using a flourescent microscope.
David