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T cells were isolated from mouse spleen cells using the CD4+
T Cell Isolation Kit, mouse. Cells were stimulated with CD3ε pure – functional grade and CD28 pure – functional grade antibodies in the presence of recombinant mouse IL-2 and mouse IL-4, as well as Anti-IFN-γ pure – functional grade antibodies for three days. Then cells were harvested, washed, and incubated with or without (left images) PMA/ionomycin and brefeldin A. Cells were harvested, fixed, permeabilized, and intracellularly stained with Anti-IL-3, CD4, and CD154 antibodies. Cells were pre-gated for CD4+
and analyzed by flow cytometry using the MACSQuant®
Analyzer. Cell debris were excluded.
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