Hi Sania, You are not really comparing like with like here. TUNEL will pick up strand breaks in DNA which will be probably a late apoptotic and also a necrotic event. Annexin will pick up cells which have some membrane asymmetry and the PI in that assay will show which cells have passed through that stage. The critical difference is that for TUNEL staining, cells are fixed so we see all cells that have had damaged DNA whereas the annexin is a live assay and we just get a picture of the cells as they are when we measure them, many of them will move through or will have moved through - apoptosis to death. With both assays it would be good to look at different time points after bacterial invasion and, in the annexin assay, to look at the cumulative annexin+/PI- and annexin+/PI+ populations. In this way you may see that your annexin results, once you include all cells that show a death commitment, may be more similar to the Tunel. Good luck! Derek On 27/6/08 13:52, "Sānia Alves dos Santos" <saniasan@usp.br> wrote: > Hi Flowers > > I'm working with hepatocyte culture, studying the frequency of apoptotic death > in cells invaded by bacteria. Using the Tunel staining I've obtaining higher > values in comparison to negative control cells (30%). Using Annexin staining > the values are very low, about 1-5% of apoptotic cells. Otherwise the PI > staining indicate necrosis in about 20% of cells. How can I interprete these > results? > > Thanks in advance, > > Sania Alves dos Santos > Laboratory of Bacteriology > Faculty of Medicine > University of Sao Paulo > tel: (5511) 3061-7029 > -- *************************************************************** Derek Davies, FACS Laboratory, London Research Institute, Cancer Research UK, 44 Lincolns Inn Fields, London, UK. Tel: (44) 20 7269 3394 FAX: (44) 20 7269 3479 mobile: 07790 604112 e_mail: derek.davies@cancer.org.uk Web Page: http://science.cancerresearchuk.org/sci/facs/ In tenebris lux ***************************************************************Received on Mon Jun 30 13:38:00 2008
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