To test the validity of the mutant selection window, we simulated mutant prevention concentration-targeted fluoroquinolone concentrations using an in vitro model with infected fibrin clots. Therapeutic ciprofloxacin (peak 5 microg/mL; t(1/2) 4 h), gatifloxacin (3.5 microg/mL; 8h), gemifloxacin (1.25 microg/mL; 8 h), levofloxacin (6 microg/mL; 6 h) and moxifloxacin (4.5 microg/mL; 12 h) were tested against methicillin-susceptible and -resistant Staphylococcus aureus, as were mutant prevention concentration (MPC)-targeted regimens achieving a trough of 1/4x or 2x MPC. MIC/MPC for MSSA K553 were 0.125/2, 0.03/0.125, 0.03/0.063, 0.125/1 and 0.015/0.25 microg/mL for ciprofloxacin, gatifloxacin, gemifloxacin, levofloxacin and moxifloxacin, respectively. Corresponding values for MRSA 494 were 0.125/1, 0.063/0.125, 0.03/0.063, 0.125/0.5 and 0.063/0.125 microg/mL. All regimens produced efflux mutants of MSSA K553. For MRSA 494, therapeutic and 1/4x MPC levofloxacin regimens produced resistance, whereas only 1/4x MPC regimens of gatifloxacin, gemifloxacin, and moxifloxacin produced resistance. All ciprofloxacin regimens produced resistance. Ciprofloxacin 1/4x MPC and therapeutic levofloxacin caused outgrowth of GrlA mutants (S80Y amino acid substitution); efflux mutants were isolated in all other cases. Overall, gatifloxacin, gemifloxacin, and moxifloxacin displayed a lesser propensity to select resistant isolates of S. aureus than ciprofloxacin and levofloxacin. The mutant selection window premise appeared valid for MRSA only. Additional studies are necessary to define the applicability of the MPC.