Aude, Broad %c.v.s can come from inadequate staining, or an inadequate cell suspension. But there's no reason to expect that your primary tissues would be inherently more difficult to analyze for DNA than your cell line. Perhaps you need to improve disaggregation. Look at the cells by fluorescence microscopy. I would guess you'll find lots of aggregates, so the answer most likely lies there. Beyond this, maybe what you're looking at is a mix of ploidies . . . aneuploid vs. diploid? Using selective antisera to separate subpopulations can often help. MAK. -- Mark A. KuKuruga, Managing Director University of Michigan Flow Core 7416 CCGC 0946 (734) 647-3216, fax (734) 936-7376 kukuru@umich.edu >>> Aude Barani <aude.barani@univ-st-etienne.fr> 01/24/01 02:56AM >>> Hello, I am a PhD student working in the Laboratory of Physiology, in Saint Etienne medecine university (France). I have performed some cell cycle analyses using normal cells isolated from fresh tissu and allowed to proliferate for 7 day in presence of serum. My problem is that DNA profiles obtained with this kind of cells completely differ from the one obtained with cell lines and oncogenique cells : the profile of the lastest can be discribed as 2 distinct peaks (2N, 4N) separated by a valley. Profiles obtained with primary cells can be discribed as follow : an aymetric peak with a large CV, corresponding to a large increase in the relative S-Phase population. This increase do not allows us to clearly distinguish G0/G1 peak from G2/M peak. Is anybody has ever worked on "proliferating primary normal cell" analyse of the cell cycle? Did someone has ever obtained these kind of DNA profile? If you know any references about this kind of DNA profile and its analysis, it would be a very helpful. If anyone can help me, it would be grateful Aude Barani ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Aude Barani Laboratoire de Physiologie GIP-Exercice, Groupe PPEH Faculte de Medecine Jacques Lisfranc 15 Rue Ambroise Pare 42023 Saint-Etienne cedex 02 - France Tel: (0)4 77 42 14 75 Fax: (0)4 77 42 14 70 mail to : aude.barani@univ-st-etienne.fr ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
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