Uptake of granzyme B is mediated through specific interactions at the cell surface. Jurkat cells were incubated in granyzme B at 4°C or 37°C for 60 minutes, washed in citrate (pH 3.0) or medium pH 7.4. Cell death was assayed by TUNEL and annexin V-FITC and analyzed by flow cytometry. Percentage of specific killing by granzyme B and Ad2 at 4°C was calculated relative to that observed at 37°C. Sufficient granzyme B remained bound to targets at 4°C after a neutral wash to induce apoptosis after addition of Ad2 at 37°C, whereas acid wash of granzyme B-treated cells at 4°C significantly reduced the level of cell death.