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INFORMATION ........
from the Flow Cytometry Core Facility
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Us9-GFP
This GFP-fusion protein provides enhanced accuracy for quantitation
of cell cycle analysis in transfected cells by flow cytometry. The integral
membrane GFP-fusion protein Us9-GFP
(Brideau et al, J.Virol
72:4560-70,1998), is quantitatively retained in cells
following ethanol permeabilization and provides for increased sensitivity
in detection and resolution of GFP-expressing, transfected cells. It's
fluorescence is not affected by permeabilization with ethanol or other
commonly used fix/perm reagents so is superior to the other GFP-fusion
proteins used in this type of assay (example
data).
See:
An Integral Membrane Green Fluorescent Protein
Marker, Us9-GFP, Is Quantitatively Retained in Cells During Propidium Iodide-Based
Cell Cycle Analysis by Flow Cytometry. Robert F. Kalejta, Amy
D. Brideau, Bruce W. Banfield, Andrew J. Beavis. Experimental Cell Research,
248 (1), April 10, 1999, p322-328.
The original publication outlining the use of a membrane-associated
GFP-fusion protein, GFP-Spectrin, for simultaneous analysis of cell cycle
with propidium iodide in GFP-expressing, transfected cells was published
in 1997:
Use of a Membrane-Localized Green Fluorescent
Protein Allows Simultaneous Analysis of Transfected Cells and Cell Cycle
Analysis by Flow Cytometry. Kalejta RF, Shenk T, Beavis AJ.Cytometry,
29:286-291 (1997).
EGFP, ECFP
and EYFP Three-Colour Flow Cytometric Analysis
We have developed a protocol for the simultaneous analysis of the EGFP,
ECFP and EYFP using flow cytometry and single-laser excitation at 458 nm.
This was performed using a standard flow cytometer, a multiline argon-ion
laser tuned to 458 nm and a custom optical filter set. Boolean gate logic
and multicolour gating allows for identification and quantification of
the eight possible subpopulations (example
data).
See:
Simultaneous analysis of the cyan, yellow and green fluorescent
proteins by flow cytometry using single-laser excitation at 458 nm.
Beavis AJ and Kalejta RF. Cytometry 37(1), pp 68-73. 1999.
Christina DeCoste,
Manager, Flow Cytometry Core Facility,
Dept. Molecular Biology,
Princeton University, Princeton NJ 08544
Tel:(609) 258 1695
email:cdecoste@princeton.edu