Both NK cells and T cells exhibit significant production of IFN-γ in response to Herceptin and IL-12 costimulation. The MDA-468 and SKBR3 cell lines were cultured with PBMCs within our in vitro coculture assay. Standard controls were also included. Brefeldin A (10 μg/ml) was included in all culture wells in order to halt protein trafficking from the Golgi. After 6 hours of culture, cells were harvested and labeled for surface expression of either CD56, CD3, CD20, or CD14 with PE-labeled mAb’s. Cells were then permeabilized, fixed, and stained for intracellular IFN-γ protein with an FITC-labeled mAb. Shown are representative dot plots of the PBMC population (vs. the SKBR3 cell line) stained for IFN-γ production within (a) CD56⁺ NK cells and (b) CD3⁺ T cells. The percentage of cells secreting IFN-γ along with the mean fluorescent intensity (MFI) is indicated within each plot. The fluorescent intensity of cells stained with isotype control Ab’s routinely fell within the first log (not shown).