Immune cells communicate by releasing large quantities of cytokines. Although the mechanisms of cytokine secretion are increasingly understood, quantitative knowledge of the number of cytokines per vesicle is still lacking. Here, we measured with quantitative microscopy the release rate of vesicles potentially carrying interleukin-6 (IL-6) in human dendritic cells. By comparing this to the total secreted IL-6, we estimate that secretory vesicles contain about 0.5–3 IL-6 molecules, but with a large spread among cells/donors. Moreover, IL-6 did not accumulate within most cells, indicating that synthesis and not trafficking is the bottleneck for IL-6 production. IL-6 accumulated in the Golgi apparatus only in ~ 10% of the cells. Understanding how immune cells produce cytokines is important for designing new immunomodulatory drugs.