Hello world! I have a user who is transfecting cells with a construct for a cytosolic GFP. We are then assessing the cell cycle status of the GFP+ve cells using Hoechst 33342 as the DNA stain. Thats fine. NO problems. However, due to time constraints, accessibility to the UV laser and the large number of potential samples, what we also want to be able to do is perform the same assay on fixed cells. Ethanol fixation abolishes or greatly reduces GFP staining, and paraformaldehyde fixation leads to insufficiently peaky DNA histograms. Now before we re-invent the wheel, is there anyone who can point us to the best way of doing this. Would PF then post-fixing in ethanol be better? Or detergent? A different GFP? Any suggestion gratefully received! TIA, Derek Derek Davies FACS Lab Imperial Cancer Research Fund, London, UK http://www.icnet.uk/axp/facs/davies/index.html
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