A collaborator is staining different fly parts (wings, imaginal disks, eyes, etc) for cell cycle. Flies have been modified with a bicistronic vector so that they can express different proteins in tandem w/ GFP. The current protocol she uses is to stain cells for 3 hours with 0.5ug/mL Hoechst while the cells are being digested w/ trypsin during this same time point. Staining looks "okay", but the CVs are pretty wide - unacceptably wide, actually. We can't increase Hoechst concentration (toxic), and decreasing incubation time doesn't seem to work either. The last hope is to stain after tissue trypsinization/dissociation. So...does anyone know of ways to improve this? Thanks, do ----------------------------------------------------------- Douglas P. Olson Experimental Hematology/AIDS Research Center Massachusetts General Hospital Harvard Medical School 149 13th Street Boston, MA 02129 (617)724-2668 - Phone (617)726-4691 - Fax Dolson4@partners.org - Email http://www.mgh.harvard.edu/aids
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