To describe the relationship between the ratio of the 24-h area under the concentration–time curve (AUC 24 ) to minimum inhibitory concentration (MIC) as well as the effect of linezolid on Staphylococcus aureus, the killing kinetics of three S. aureus strains was studied by in vitro simulation of 5-day antibiotic dosing over a wide range of AUC 24 /MIC ratios. Similarly susceptible meticillin-resistant S. aureus ATCC 43300 and S. aureus 479 and vancomycin-intermediate S. aureus ATCC 700699 (Mu50) at a starting inoculum of 10 8 colony-forming units (CFU)/mL were exposed to multiple-dose pharmacokinetics of twice-daily linezolid for 5 days. The simulated AUC 24 /MIC ratios varied from 30h to 1200h (S. aureus ATCC 43300), from 30h to 600h (S. aureus 479) and from 50h to 400h (S. aureus ATCC 700699). The cumulative antimicrobial effect was expressed by its intensity (I E ) measured from the start of treatment to the time after the last antibiotic dose when numbers of antibiotic-exposed bacteria reached ≥10 8 CFU/mL. With each organism, bacterial re-growth followed a pronounced reduction of the starting inoculum that occurred at each simulated AUC 24 /MIC ratio except for the lowest value (30h). This reduction was AUC 24 /MIC-dependent: the minimum numbers of surviving organisms decreased with increasing AUC 24 /MIC ratios. A sigmoid relationship was established between I E and the simulated AUC 24 /MIC ratio. This relationship was bacterial strain-independent; a logistic function fits the combined data with r 2 =0.95. The established AUC 24 /MIC–I E relationship is useful to predict the antistaphylococcal effects of linezolid at clinically attainable AUC 24 /MIC values.