Although I rarely say "never", I will say it here: NEVER try to phenotype dead cells. Too many artefacts. There is no such thing as "how many dead cells are PE positive." Depending on the "affinity" of your antibody clone for dead cells, this can be anywhere from what you want to 100%. mr At 10:54 AM -0500 12/18/01, Mark Kukuruga wrote: >. . . unless, of course, if instead of dead cell elimination one is >interested in >live/dead discrimination and enumeration. Example: label cells >with a PE conjugated >antibody . . . now, of the dead cells, how many are PE positive, and >how many are >just dead? >mak. > >-- >Mark A. KuKuruga, Managing Director >University of Michigan Flow Core >7416 CCGC 0946 >(734) 647-3216, fax (734) 936-7376 >kukuru@umich.edu > > >>>> "DAVID M CODER" <d_coder@MSN.com> 12/17/01 09:33AM >>> >Either PI or 7-AAD will work. Moreover, with either DNA-binding dye, >compensation in >only direction is needed: You must make sure that PE-positive cells >do not appear to be >7-AAD or PI-positive. Compensation 7-AAD or PI into PE positive >space is unnecessary >as those cells are eliminated from further analysis. The attached >images illustrate >compensated vs. uncompensated PE. > >For details see web page that I developed when I was directing the >lab at the University >of Washington: >http://nucleus.immunol.washington.edu/Research_facilities/Apps/7aad.html > >Dave >---------------- >David M. Coder, Ph.D. >Consulting in all aspects of cytometry. > >email: d_coder@msn.com >tel./messages: 206-499-3446 > ----- Original Message ----- > From: Jáksó Pál > To: Cytometry Mailing List > Sent: Monday, November 26, 2001 11:21 PM > Subject: RE: 7AAD PROTOCOLS > > > Dear Rachel, > > 7AAD is may be better in some application because of the lower >crosstalking to the > FL2 channel. Accordingly you can use it together with PE stains. >PI is very hard to > compensate from the FL2. But if you use only FITC as second stain >(and nothing else) > you can use both 7AAD and PI for dead cell labeling. > > Pal Jakso > > University of Pecs > Faculty of Medicine > Dept. of Pathology > 7643, Hungary > 12. Szigeti str. > > > While this subject has come up: it seems like both 7AAD and propidium iodide > can be (and are) used for detecting dead or apoptotic cells - are they > interchangeable ? If people have distinct preferences, what is the > difference ? > > thanks, > Rachel
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