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Human peripheral blood mononuclear cells (PBMCs) were either left unstimulated (left images) or stimulated with SEB for six hours. After two hours, brefeldin A was added. The cells were fixed, permeabilized, and intracellularly stained with Anti-TNF-α antibodies as well as with CD4 antibodies and CD154 antibodies. Cells were then analyzed by flow cytometry using the MACSQuant®
Analyzer. The Tandem Signal Enhancer has been used to increase binding specificity of tandem-dye–conjugated antibodies. CD4+
lymphocets were pre-gated for the analysis. Cell debris were excluded from the analysis based on scatter signals.
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