James, Assuming you're using PI after ethanol . . . We've seen this kind of (probable) artifact when staining other types of cells. You're most likely seeing a difference due to a) differential permeabilization, b) DNA conformation differences, like that you see in activated monocytes/macrophages, or c) non-specific staining of the cytoplasm (mitochondrial DNA?), something we've seen in squamous cells. I'd try different permeabilization, or perhaps a hypotonic citrate / detergent system to strip most of the cytoplasm. Also, if you have access to a UV source, try DAPI . . . DAPI's been shown to be least sensitive to these problems. MAK. -- Mark A. KuKuruga, Managing Director University of Michigan Flow Core 7416 CCGC 0946 (734) 647-3216, fax (734) 936-7376 kukuru@umich.edu >>> "James S. Clark" <jclark@clinmed.gla.ac.uk> 07/16/01 07:01AM >>> Hi All, I have an unusual situation. I have been looking at a range of congenic rats and have found that the vascular smooth muscle cells seem to be aneuploid. The profile is normal at the 1N and there is a leak about 1.2N and a large spread at G2M (maybe 2 populations). I have ran a large number of tumor samples in the past and this profile shows great similarity to a number of them. The rats are otherwise normal apart from this unusual DNA profile . Has anyone any idea what could be reason for this unusual DNA profile? Best wishes James
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