Introduction: Combining currently available antibiotics to optimize their use is a promising strategy to reduce treatment failures. Nevertheless, most of the combination assays are usually performed in vitro on bacteria exposed to constant concentrations of antibiotics over less than 24 h which seems poorly predictive for clinical situations. The aim of this study was to compare static and dynamic in vitro assays to check if the use of dynamic models could improve the predictability of efficacy for combination treatments. Materials and Methods: The potential efficacy of vancomycin and/or amikacin on a Methicillin‐Sensitive Staphylococcus aureus (MSSA, Hg001 derived from NCTC 8325) strain and a Methicillin‐Resistant S. aureus (MRSA, ATCC 33591) strain was tested by Checkerboard assays (CA), Time‐Kill Studies (TKS) and a Hollow‐Fiber Model (HFM). TKS were performed over 24 h by exposing two different initial inoculum sizes, standard (5.7 log10 CFU ml−1) and large (9.7 log10 CFU ml−1) at the maximal free plasma concentrations that would be attained in patients. In the HFM, the inocula were exposed to free concentration profiles similar to those that would be observed in human patients receiving daily administrations of antibiotics over 5 days. Results and Conclusions: The 2 strains were considered as susceptible or intermediate to vancomycin and amikacin based on EUCAST breakpoints. The CA showed no synergism nor antagonism between the two drugs. On standard inocula, amikacin and vancomycin alone or in combination at constant concentrations decreased the bacterial burden below the limit of detection in less than 24 h except amikacin on the MRSA strain in TKS. In TKS on large inocula, bacterial reductions were only observed after exposure of the MRSA strain to vancomycin alone or in combination and after exposure of the MSSA strain to amikacin alone or in combination. It means that the combination had similar activity as the best drug alone and never improved the efficacy compared to the monotherapy. In the HFM, after 5 days of exposure of the large inoculum to fluctuating concentrations of amikacin or vancomycin in monotherapy, no reduction of bacterial load was observed whereas we identified a synergistic effect of their combination with a mean reduction of 5.1 log10 CFU ml−1 of the bacterial population. In conclusion, this study suggests that in vitro static assays by exposing bacteria to constant concentration over a short duration, could be insufficient to evaluate the potential benefit of a combination therapy on large inocula