Hi Jose, from my experience I would say, 1. never trust the result upon apoptosis definition based on only one criterion. Always include two or three assays, for example the TUNEL assay and the appearance of sub-G1 cells. On the other hands, I would recommend to stain the DNA of the cells with a stain like Hoechst 33342 and check for the appearance of condensed chromatin or apoptotic bodies by a microsope. 2. Using PI and annexin, Pi positiv and annexin negative should be necrotic cells, annexin positive and PI negative are clearly apoptotic cells and both positive could be both, late apoptotic or necrotic cells. Andreas From: "Jose Benito" <jbenito1@hotmail.com> To: cyto-inbox Subject: Apoptosis and anexin-V Date sent: Wed, 10 May 2000 11:15:25 CEST > > Hi everyone. > I want to study apoptosis in PBMCs using the staining with annexin as a > marker of apoptotic cells. I have some questions regarding this issue: > 1.- How good the annexin staining correlates with apoptosis. Can a cell be > defined as apoptoic simply based in the annexin criterion? > 2.- Is there any test rally specific for the fenomenon of apoptosis? > 3.- What is the rationale of using propidium iodide in combination with > annexin? Is it to differentiate apoptotic from necrotic cells? If this is > the case, does it mean that cells positive for annexin and negative for PI > should be classified as apoptotic and double positive cells as necrotic? > Could you find cells positive for PI and negative for annexin? > Sorry for so many questions and Thanks > > José Miguel benito > Infectious Diseases Division > Hospital Carlos III > Madrid > ________________________________________________________________________ > Get Your Private, Free E-mail from MSN Hotmail at http://www.hotmail.com > > My address: PD Dr. Andreas Simm Institut fuer Klinische Biochemie und Pathobiochemie Versbacher Str. 5 D-97078 Wuerzburg Germany Tel.: +49 931-201-3144 FAX.: +49 931-201-3153 E-mail: simm@klin-biochem.uni-wuerzburg.de
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