Hello Jan, I think the problem is not with the permeabilisation. Cells are producing but also secreting the cytokines. If you want to detect it, you should block the exocytosis 12-24 h before the labeling. Try to use brefeldin-A or monesin. Gogy Peter Gogolak, Ph.D. Institute of Immunology University of Debrecen, Hungary Phone/Fax: +36 52 417159 Dear All, I have a user who wishes to detect cytokine production in a transfected cell line, see below . Use of at least one commercial permeabilisation kit has failed. Literature and email archive searches have yielded virtually no information, so any insights are appreciated! Jan Grawé >Hi, > >I am trying to develop an intracellular staining protocol to detect >intracellular protein in a tumor cell line >by flow cytometry. I have checked the literature and the technique >seems to be used almost entirely for >lymphoid cells. Does this mean that other types of cells are >difficult to permeabilize? > >Does anyone have any experience detecting intracellular protein >within tumor cells using flow cytometry? > >I am transducing the cell line MB49 (murine bladder carcinoma) with >recombinant adenovirus expressing IL-15, >and I would like to detect IL-15 by intracellular staining. > >I would really appreciate any insight and/or comments. > >Thanks, >Christina Ninalga -- -------------------------------- Jan Grawé Cellanalyslab Rudbecklaboratoriet/C5 Uppsala Universitet Dag Hammarskjölds väg 20 75185 Uppsala tel: +(0)18-4714656 mobil:+(0)70-2577874 fax: +(0)18-552739 epost: jan.grawe@rudbeck.uu.se http://www.rudbeck.uu.se/cellanalys
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