Tony, Simplest answer . . . sounds like air in the flow cell . . . have you done a drain and fill? If that's not it, then look for the sipper to be plugged. With the dial in the run position, rotate the sample tube rest into the on position WITHOUT a tube . . you should see a drip. Alternatively, you could remove the outer tube of the sipper (vacuum sheath), and see the drip-back in any position. If you don't see any drip, try placing a piece of tubing - - one end fitting the end of the inner sipper tube, the other connected to a syringe. Draw back on the syringe (negative pressure) to clear blockage in the sipper. Be careful not to bend or torque this inner tube . . . it goes all the way into the flow cell. Finally, there could be crud in the flow cell, either blocking the laser, or causing turbulence. After a month of sitting, this could very well be the problem . . . have you done a system bleaching recently? MAK. --- Mark A. KuKuruga, Managing Director BRCF Flow Cytometry Core The University of Michigan Medical Center 7416 CCGC 0946 1500 East Medical Center Drive Ann Arbor, MI 48109 734-647-3216 (voice) 734-936-7376 (fax) kukuru@umich.edu >>> "Tony Schountz" <tschount@mesastate.edu> 11/14/01 18:39 PM >>> I've run into a problem with our FACScan (HP system) that seems rather bizarre. I started to set compensation manually using the FACScan Research Software, but when I put cells on there appeared to be no flow through the instrument (on the computer screen or the instrument panel). So I checked all the clear lines for occlusions and none were obvious. I marked the fluid level on the sheath tank and after a few minutes the level had dropped below the mark, indicating that fluid was going throgh the system. So I quit FRS and launched Autocomp and after pressing the "PMT" button I had events occurring in the lower left quadrant of the FL1/FL2 window and events on the FSC axis - with only water in my tube (i.e. no beads). The flow rate of these "events" was between 3500 and 5000. Of course, after a couple of minutes the software informs me of an error. Could someone shed a little light on what might be occurring here? What are these "events" that Autocomp is detecting? The instrument was used about a month ago without any troubles, and I take great care in trying to keep the lines free of occlusions since it frequently sets idle for this amount of time. This instrument was donated to our college a couple of years ago, and I don't have any formal training for maintaining it. I'd sure like to take such a course, but it is fiscally prohibitive for me. So, I'm relying on the collective expertise (and generosity) of the group to help me out. Thanks, Tony -- Tony Schountz, Ph.D. Dept. of Biology Mesa State College
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