Hi Vida, I would try the 100 nozzle, as a rough rule of thumb the nozzle size should be 3-4 times the diameter of the cells. For all large cells we use a 100 nozzle (not yet managed to get a bigger one from BD). The fuzzy side streams also give you an indication (especially if you are using a 1 drop sort mode). Accudrops will be fine with all size nozzles. We routinely run these for set up, then backflush the sample line and run a combination of rinse and sometimes ethanol to clear them out before the sort. Good luck! Derek On 7/5/08 0:35, "vhodara@sfbr.org" <vhodara@sfbr.org> wrote: > Dear experts in flow sorting: > > I am trying to sort HUH7 cells GFP (brigth and not so bright populations) Just > two gates in a single color FL1 vs SSC (very simple sorting should be). Those > are big cells so I tried with the 70um nozzle but with medium pressure. I > haven't had problems with settings, or with accudrop. However, I couldn't have > pure populations. When setting the streams I can see the two perfect dots on > the camera on the screen and accudrop run perfectly, but when I am sorting I > don't see but a smear of white dots to the right and to the left of the main > stream (is that because of the big cells?). I am planning to try the 100um > nozzle, but can I run the accudrop with this nozzle eventhough they are very > small? If somebody can give me some tips, I will be very grateful. > And another question, if I prepare the machine for asceptic sorting, can I run > the accudrop before or I need to prepare a sterile solution of these beads or > run them after asceptic cleaning and then run again bleach and water for a few > minutes to keep the "sterility". (We do not have the FACSAria in a BSL-3 lab > yet) > Thanks a Lot > Vida > -- *************************************************************** Derek Davies, FACS Laboratory, London Research Institute, Cancer Research UK, 44 Lincolns Inn Fields, London, UK. Tel: (44) 20 7269 3394 FAX: (44) 20 7269 3479 mobile: 07790 604112 e_mail: derek.davies@cancer.org.uk Web Page: http://science.cancerresearchuk.org/sci/facs/ In tenebris lux ***************************************************************Received on Fri May 9 14:38:00 2008
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