GFP sorting question
From: Andrew Oberyszyn (oberyszyn.2@osu.edu)
Date: Wed Oct 16 2002 - 15:45:14 EST
Hi FLow-er in Flow land,
I've come across an interesting (and confusing) problem which sorting GFP
cells. I've noticed several times that I get alot of fanning while
sorting. I don't believe it is the Frequency, drive or phase since
setting up the sort gives me beautiful side streams and great
separation. I first notice this on my Elite and now I have a
FACSVantage and 2 of the last 3 GFP sorts showed this problem.
After todays sort, which had terrible fanning, I put on a sample of
unstained fixed PBL's to see if it was instrument related. Not
surprisingly, that sample gave great side streams. Obviously the
problem is with the sample. I've never (never say never!) seen a
fanning problem with surface stained cells and I don't understand what
could be causing this to happen with GFP cells? Could the
transfection method somehow "charge" the cells which interferes
with the charge applied for sorting? As far as I can tell the media
(which is MEM with Pen, strep, L-Glut, FCS, bicarb.) shouldn't do
anything?
Any help or suggestions would be greatly appreciated!
Thanks in advance.
Andy
^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^
Andy Oberyszyn,
M.S.
Core lab
manager
The Ohio State
University
University Cell Analysis & Sorting
Core
424 Davis Heart & Lung
Institute
473 West 12th
Avenue
Columbus, Ohio
43210
Tel:
614/292-FLOW(3569)
Fax:
614/292-7335
E-Mail: cytometry@osu.edu
Website:
http://heartlung.osu.edu/hlri/corelabs/flowcore.jsp
^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^
"What if the Hokey Pokey is
really what it's all about?!?"
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