Green Fluo. Prot.
nick at lmb1.rug.ac.be
Thu Jan 12 11:04:18 EST 1995
> Hello All,
> I'm looking for advice and experience in using the Green Fluorescent
> Protein (GFP) marker in combination with FACS analysis. I'd like to use
> it in mammalian (mouse thymocyte) cells. The idea is to drive GFP
> expression by a promoter of interest and isolate the subpopulation of
> GFP-positive cells for further study. Anyway, are people finding the GFP
> marker useful for FACS analysis in mammalian or other tissue culture
> systems? Positive and negative experiences are greatly appreciated.
> Thanks, Dave Askew
> askewd at gas.uug.arizona.edu
I tried to use GFP in L929 murine fibrosarcoma cells. However, no
fluorescence could be detected by FACS nor by fluorescence-microscopic analysis.
GFP was under control of the - strong - CMV promotor. Not all cells seem to
be able to apply the cyclization of the GFP peptide, necessary to obtain
fluorescence. Also, try to use excitement-wavelength 395 nm; this gives
stronger fluorescence than 488 nm.
Someone else on the lab had success with myeloma cells. Detection
was performed by microscopy.
If anyone else has had any success with other cells, please
mention it here. It would be very usefull. Thanx-a-lot...
Nick Vercammen (nick at lmb1.rug.ac.be)
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