Hi Andy, We've been experiencing similar problems since we had our Vantage upgraded to a Vantage SE : voltages for APC (P6/FL5) and APC-Cy7 (P5/FL4) detection have to be set abnormally high (>900) (we use optical bench configuration "6" as drawn in the manual). With our old "regular" settings (in the past we considered 600V amp to be quite high already), both stained as well as unstained cell clusters disappear down to oblivion. I'm sending some dot-plots of this situation to BD soon, at this time they don't know any answer to the problem. FL3 was down shortly after the SE upgrade. However after giving a little nudge to the FL1-FL2 vs. FL3 splitter (610 SP) the signal balance was restored in favour of FL3 without affecting FL1-FL2 detection. I assume you have checked your optical bench configuration in detail - it's easy to misplace a splitter, invert a filter etc... Refer strictly to the configurations drawn in the manual. If this doesn't help it's definitely up to BD to fix the problem. I will keep you posted if anything new pops up on that front. "MORSCHAUSER, ANDREW" wrote: > This message is directed towards FACSVantage users that have upgraded their > machine from a FACSVantage to a FACSVantageSE w/Turbosort. I am > experiencing a > decreased sensitivity in FL-3. When comparing alignment particles (cRBC, > Alignflow beads, Calibrite beads) between a FACSVantage, FACSVantageSE w/TSO > and > a FACSCalibur, results have shown the FACSVantage SE signal to be > significantly > lower than the Vantage or Calibur. Furthermore, when looking at splenocytes > stained with CD4 PE-Cy7 (787/40 filter), there is no discrimination between > negative/unstained cells vs. stained cells. The signals on the FACSVantage > and > Calibur are nearing the third decade while the FACSVantageSE signal was > negative > with log amps and voltages greater than 900V. I tried increasing the log > offset > as far as it will go and found little if any improvement in the signal. > > Boards have been replaced and new PMTs in FL3 have done nothing to remedy > the > problem. BD has been very responsive to my concerns, but the problem is > still > present and I was wondering if anyone else has experienced similar problems > with > the extra mirror and different optical path for a 5-color FACSVantage SE. > Any > comments or suggestions would be greatly appreciated. > > Andy Morschauser > Schering-Plough Research Institute > Department of Immunology > 2015 Galloping Hill Road > K15-3-3945 > Kenilworth, NJ 07033 > 908-740-3089
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