The chemokine RANTES is secreted rapidly after activation of human CD8+ T cells, with a cycloheximide-resistant burst during the first hour. This pattern was observed in purified memory and effector phenotype CD8+ cells from blood as well as in blasts. In contrast, secretion of other chemokines and interferon-gamma by these cells was sensitive to cycloheximide and detectable only after a lag. Immunofluorescence microscopy of CD8+ memory and effector cells and blasts showed RANTES present in intracellular vesicles that do not significantly colocalize with cytotoxic granule markers or other markers of defined cytoplasmic compartments. Immunoelectron microscopy confirmed that RANTES is stored in small vesicles distinct from the lysosomal secretory granules. RANTES+ vesicles polarize rapidly in response to TcR engagement and are more rapidly depleted from the cytoplasm. These results show that CD8+ T cells have two distinct TcR-regulated secretory compartments characterized by different mobilization kinetics, effector molecules, and biological function.