Andy, We've been sorting transfected GFP osteoblasts for several years now and had exactly the same problem you describe. Initially we had less fanning on our FACStar Plus than on our FACSVantage SE so we assumed the problem was due to the increased "sensitivity" of the longer deflection plates since we were also using identical pressure and drop drive on both machines but our service engineer noticed a lot of downstream signaling in the forward scatter indicating that the cells were a lot larger than we thought and a lot more varied in size. He suggested actually going to a higher pressure and ddf than we were used to employing with our 100um nozzle, specifically we settled on 12 psi with a ddf of 20.7 and an amplitude of 5.2 volts on our DIVA which usually accudrops to 16 - 17, similar settings in analog. We also filter the cells three times through the Falcon blue-capped strainer tubes, 35um mesh and this seems to have helped. I should mention we still get some fanning but it is considerably less. I think the combination of going to a larger nozzle, playing with your pressure and ddf settings and triple filtering the cells is the answer. Gene Pizzo Manager FACS Facility, UCONN Health
This archive was generated by hypermail 2b29 : Sun Jan 05 2003 - 19:26:27 EST