We have studied the intracellular bioactivity of several antimicrobial agents against two strains of Staphylococcus aureus, a pathogen sequestrated in phagolysosomes, using peripheral blood neutrophils from human volunteers. This was compared to the activity of cell-associated drugs also measured in vitro. Several discrepancies (high cellular association, low bioactivity) were observed (coumermycin, glycopeptides, erythromycin and clindamycin) which can be due to the binding of the drug to a particular cellular organelle, to intracellular metabolization or inactivation, to unfavourable conditions in the phagolysosome (pH drop), or to a toxic effect of the drug on the functions of the neutrophil. Addition of the antibiotic during ingestion was frequently associated with better neutrophil-dependent killing through several potential mechanisms: coingestion of the antibiotic with the inoculum, modification of opsonization, release of activating substances and fragilization of the microorganism to oxygen-dependent or -independent killing mechanisms.