Hi folks, A few weeks (months?!) ago I asked about sorting single sperm for PCR, and recieved quite a few encouraging responses. Th interested party has obtained the dye (we're using Ho348) and tested their PCR after staining, and they're quite satisfied and ready to sort. Now the ball is in my court.... We have a 753 with Cicero. I dug out our auto-clone unit, got the 753 sorting beautifully with the normal sort drawer, and then installed the auto-clone. Now I'm stuck! The next step is calibration and testing, to adjust the instrument so that the sorted test stream (actually the "unsorted" stream) goes through a hole in a plate in the sort drawer. The multiwell plate is underneath this hole, and any droplet that is going to land in the well must go through this hole. Problem is, my stream won't go through the hole! The waste (what I would call the center stream and the left stream) is deflected to the left and lands in the waste catcher, but the stream I want (the right stream, which should be falling straight down) is too far to the right. The manual indicates that I should adjust the right-left and front-back knobs on the autoclone until the stream goes through the hole, but the right-left knob has no noticeable effect. Does anyone know what it does? It looks like a potentiometer, and I'm speculating that it modulates some bias in the plate charges to shift all streams right or left. I thought about yanking it and substituting one w/ a larger range of resistances, but there should be better way. I tried decreasing deflection gain, and that does move it a bit to the left, but not enough, and I'm limited by the fact that at too low a setting the waste streams start splattering off the edge of the waste catcher. Does anyone have any ideas? I won't be able to work w/ the instrument again for another week, but we really need to get this working. Suspecting that deflection in one direction may be higher than the other, I'm going to try switching the plate polarity and cable setups so that right is left. Does anyone have any experience w/ this beast, or know of any other tricks? ------------------------------------------------------------- Steve G. Hilliard Join the fight--help Cell Analysis Facility stamp out smartass University of Georgia sig files! -------------------------------------------------------------
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