Maher, In my experience with whole blood I/C cytokine production there is usually complete loss of CD4 following activation of lymphocytes with PMA+Ion. I usually use FL3 gating for lymphocyte subset identification, and even the bright stains eg, PC5 (PECY5) will not identify these cells following activation with these compounds. I have subsequently used the CD3 pos / CD8 neg identification of these cells, which one has to run anyway to identify the true CD8 pos T cells (assuming the CD3 pos / CD8 neg / CD4 neg and the CD3 pos /CD8 pos / CD4 pos populations are negligible). Alternatively, if you are using purified cells you could sort the CD4 pos cells prior to stimulation. Greg Hodge Flow Lab / Haematology Department, Women's and Children's Hospital, 72 King William St, North Adelaide, South Australia 5006 ph:08 82046522 fax: 08 82046043 Email: hodgeg@mail.wch.sa.gov.au
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