Perforin staining is not enhanced by preventing degranulation, preventing acidification of cytotoxic granules, blocking secretion, or inhibiting proteolysis.

A T-cell line from a healthy donor specific for an HLA B8-restricted EBV peptide was incubated for 2 hours at 37°C with the indicated agents before staining for CD8, the EBV tetramer, and perforin or GzmA. Chloroquine (CHQ), which blocks lysosomal acidification, enhanced GzmA staining but not perforin staining. The ratio of mean fluorescence intensity (MFI) of perforin or GzmA expression of treated cells to mock-treated cells is shown. The agents tested were protease inhibitors (PI), cathepsin inhibitors (CI), CHQ, EGTA, Brefeldin A (BFA), or cycloheximide (CHX) at concentrations given in “Patients, materials, and methods.” Med indicates medium.