We have encountered similar problems trying to assay ion flux, especially if it is a transient change. We set up a kinetics experiment where the sample is run to get a baseline. We add an agent to induce ion flux, mix, and then get the sample running again as quickly as possible. A lot of the time we would miss the major shift in fluorescence, which is linked to the ion flux obviously, and just observe the fluorescence returning to baseline. Upon further observation we discovered that mixing is a major problem. 1) It takes too long to mix the sample in the sample tube. 2)After mixing the sample with your ion flux inducer(or inhibitor), when the sample starts running again, it is going to mix with the untreated sample that is in the sample tubing. We are trying to overcome this problem using some hardware and software from Alitea USA, Medina, WA. 800-963-1101. It is computer driven peristaltic pumping system that can deliver small volumes precisely and at controlled rates. However, it isn't cheap ($3-4,000?). We haven't invested a whole lot of time in it so far (busy,busy). Gary Durack,the facility director, would know more but he's out until the 17th. I'm sure that Alitea would also be helpful. David McFarland University of Illinois Biotechnology Center Flow Cytometry Facility ---------- Forwarded message ---------- Date: Thu, 6 Mar 1997 12:41:33 -0600 (CST) From: Raymond B. Hester <rhester@jaguar1.usouthal.edu> To: cyto-inbox Subject: Transit time Dear Group, An investigator wants to look (using flow) at Ca++ responses that occur within seconds (as in 1 or 2 seconds) after the addition of a particular agent to dissociated endothelial/epithelial cells. Can anyone tell us the fastest transit time from sample uptake to laser intercept with either a Coulter Elite or FACSVantage? I also seem to remember a commercially available device that could be attached to (any?) flow cytometer and that increased the speed of sample injection. Does anyone know the particulars about this device - manufacturer, cost, and how well it works? Can it can be attached to both the Coulter Elite and the FACS Vantage? Thanks for any info. Ray ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ Raymond B. Hester, Ph.D. Flow Cytometry Lab CSAB 357, Biotechnical Services The University of South Alabama Mobile, AL 36688-0002 To err is human - Email: rhester@jaguar1.usouthal.edu but it feels divine. Voice: (334) 460-6029 Mae West FAX: (334) 460-6073 ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++
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