-----Original Message----- From: Mike Evans [mailto:xyinc3@frii.com] Sent: Friday, April 21, 2000 7:20 PM To: cyto-inbox Subject: RE: macrophage apoptosis and flow Normally Pmt's are not very responsive at the voltages you spoke of and will not be linear as well. What is the filter you are using for the PI. Is it a long pass. If it is you are going to get loads of light. Maybe you can put in a Band Pass filter with a fairly tight bandwidth. I would recommend a 620/20 filter for the PI. This should drop the overall intensity enough that it should allow you to run under linear conditions as well as get everything that should be on scale on scale. Mike Evans -----Original Message----- From: Jane S. Miller [mailto:miller@medicine.tamu.edu] Sent: Thursday, April 20, 2000 7:15 AM To: cyto-inbox Subject: Re: macrophage apoptosis and flow I'm wondering why you are using FL3 instead of FL2 for propidium iodine? Would that effect the brightness? >>> Maciej Simm <simmmmer@yahoo.com> 04/18/00 05:14PM >>> Dear all, One of the people I'm working with right now is studying apoptosis in periteneal macrophages using a kit with annexin V monoclonal FITC ab and propidium iodine. The cells are autofluorescent. The "unstained" tube has signal in up to 2nd log. The stained tubes are VERY HIGHLY BRIGH on FL3 (PI) and I tried to make things "fit" on the dotplot by lowering fl3 voltage by 200 or so, but I'm still not getting everything (and also some of the dimmer cells are squooshed on the axis). The FITC signal is of normal brighness. Needless to say we're not getting the same results as the kit suggests we should given the incubation/conditions. He's gonna talk to the company. I am posting this message with hopes that some of you may have seen this method or one similar to it and are willing to comment. Any suggestions are welcome thanks a bunch, Maciej ===== `---------------------------------------------` | Maciej S. Simm | 525 E 68th Street | | Research Technician | Room N-805 | | Cornell Medical Center | Tel. 212.746.3428 | `---------------------------------------------` | www.cd4cd8.com | `---------------------------------------------` __________________________________________________ Do You Yahoo!? Send online invitations with Yahoo! Invites. http://invites.yahoo.com
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