Dear Flowers I have recently been quite flumoxed by a variation in optimum FS/SS settings for analysing a platelet prep for a colleague doing some work on a single colour CD61 labelling and inhibition experiment. I have Coulter XL and had no previous experience with platelets. When first approached by my colleague, he supplied me with settings that had previously been used for platelets on another XL. These proved to be not optimum on my instrument, I found I had to reduce the FS gain from 100 to 20; doing this changed the events from two populations with only 20% CD61+ to one population with 100% CD61+. SS Log and the FL1 settings were fine. Satisfied that I had established the optimum settings for my instrument, he then set up his inhibition experiments. I subsequently have found that the FS gain was optimum at 10, otherwise we got two populations again (with FS gain 20), and only 20% positive. My concern is.. what are the events that are not labelled when the FS gain is too high and where do they go when I reduce the gain?? I've always believed that voltage settings are "cast in stone" and once established for any observation must not be altered..I've ceratianly never seen this sort of thing with white cells. Can anyone of the gurus out there help me? Jane Hughes Chief, Institute of Child Health Lab, Red Cross Children's Hospital Rondebosch, Cape Town 7700. South Africa Tel 021 658 5313 Fax 021 689 1287
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