One of the major mechanisms of cell-mediated cytotoxicity involves exocytosis of cytoplasmic granules from the effector toward the target cell. The granules contain a number of proteins, including the pore-forming protein perforin and a family of serine proteases called granzymes, including Granzyme B. Granzyme B is present mainly in the granules of CD8+ CTL and natural killer (NK) cells and mediates the lethal hit that kills virus infected and tumorigenic cells. Measurement of release of Granzyme B in response to the appropriate target is useful for evaluating cell-mediated cytotoxicity. The Granzyme B ELISPOT assay is a valuable non-radioactive alternative to the 51Cr-release killing cell assays for measuring antigen-specific CTL cytotoxicity. This Granzyme B ELISPOT Ready-SET-Go! reagent set contains the necessary reagents for performing enzyme linked immunosorbent spot (ELISPOT) assays for high resolution frequency analysis of Granzyme B-secreting cells. This ELISPOT reagent set is pre-titrated for optimal spot development.