I'm trying to isolate a stable cell line expressing very high levels of particular G protein coupled receptor genes by using a bicistronic vector co-expressing a Renilla GFP. I have sorted 3 times using the top 10%, and have not yet obtained a homogeneous population of GFP positive cells (>200 on FL1 channel). I'd like to improve this and would like to purchase an instrument to sort as many cells as possible (I need 20-100 million cells output to analyze for receptor expression). Does anyone have experience with using the Cytomation MoFlo, BD FACS Vantage, BeckmanCoulter Epics/Altra, or others machines? I'd also like to clone single cells at the end to obtain a stable line, but this may not be necessary if the pool after multiple sorts is homogeneous, stable, and high level (>3 pmol/mg binding sites, equivalent to ~200,000 sites/cell). Thank you for your help. Please reply directly to my email address alan.finegold@perkinelmer.com. Alan A. Finegold, Ph.D. Scientist, Receptor Biology R&D PerkinElmer Life Sciences 10,000 Virginia Manor Rd, Suite 360 Beltsville, MD 20705 alan.finegold@perkinelmer.com (301) 210-4700 x14 (301) 210-6266 fax
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