Dear experts in flow cytometry, I would like to perform INF g and TNF b intracellular staining of a small (0.1%) tetramer + PBMC. I have some doubts and I would be grateful if you could give me any suggestion. My lymphocytes are frozen and I wonder if there are any particular conditions for its stimulation with specific peptide. The other thing that bothers me is the precision and specificity of facs analysis. My population of Tetramere + cells is already small and its subpopulation of INF g + will be even more difficult to distinguish from the background staining. I wonder if it would be possible (and correct) to culture the lymphocytes after peptide stimulation during 48h to increase the number of double positive cells without changing considerably the proportion of tetramer+/ INF g + cells? Thanks for any suggestions! Michal -- Michal Abel, MD, PhD Laboratory of Clinical Immunology INSERM U25 Necker Hospital, 161 Rue de Sèvres. Paris tel: 33 1 40 19 28 87 fax: 33 1 44 49 53 74
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