From: ZETTLER, ERIK (EZETTLER@LIFE.JSC.NASA.GOV)
Date: Tue Jan 19 1993 - 08:48:07 EST
For calculating cells/ml in samples, we first calibrate a stock of beads gravimetrically (i.e. weigh sample to 4 decimal places, run and accumulate sample, weigh sample again and calculate volume run based spec. grav of solute) We use this technique with both EPICS and FACScan flow cytometers and it is described more thoroughly in Olson et al. 1992, "Phytoplankton analysis using described more thoroughly in Olson et al., in press, Current Methods in Aquatic Microbial Ecology, ed. P.F. Kemp et al. Since this is not out yet, if anyone is interested, let me know and I'll give more details. The problem with just running a known volume with a metered system is that if there is coincidence (and there is, even at count rates as low as 500/s) you may miss events. With the beads, the theory is that you can claibrate the stock at a very low count rate (e.g. < 100/s.) and then during a run of sample with beads, the beads are missed by coincidence at the same rate as cells and the beads counted in the final file are a measure of the volume the electronics actually "saw".
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