293 cells are very sensitive and easyly dye in suspension. we routinely detach them by gentle pipetting and keep/wash them in 20% serum until analysis/sort, that we do as soon as possible. filtering and DNase can also help. Saverio Alberti Head, Lab. of Experimental Oncology Department of Cell Biology and Oncology Consorzio Mario Negri Sud 66030 Santa Maria Imbaro (Chieti), Italy Phone: (39) 872-570.293 FAX: (39) 872-570.412 E-mail: alberti@cmns.mnegri.it On Wed, 25 Aug 1999, Barsky, Lora wrote: > > > Can anyone offer insight as to why my users have been flow problems when > using 293A and 293T cell lines on BD FACScan and BD FACSCalibur? > > The cell lines are adherent, so they trypsonize to release from the plates, > they bring the cells in DPBS to FACS. They're all looking for GFP > expression. > > I've suggested they pre-filter with a nylon mesh - 55um. But this doesn't > help much. > > No other problems with flow on other sample types. > > Any info would be appreciated! > > Thanks, > Lora W. Barsky > CHLA > Research Immunology/BMT >
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