Hi Andy, I've noticed the same thing (ie. fanning of the streams with some samples), but my vague impression was that it happened with large cell types such as fibroblasts, not necessarily with GFP. Does this ring any bells with anyone? The purity of the sorts doesn't seem to be compromised, but I suppose I must be losing some cells that don't make it into the collection tube. Cheryl on 10/16/02 4:45 PM, Andrew Oberyszyn at oberyszyn.2@osu.edu wrote: 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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