Due to the high prevalence of multi-drug resistant bacteria, combination therapy is an efficient choice for treatment of infections caused by highly resistant strains. In this study, the efficacy of ceftiofur plus danofloxacin combination was investigated against resistant Escherichia coli. The interaction between the two drugs was determined by checkerboard tests and time-kill assays. The combination was defined as bactericidal or bacteriostatic based on the minimum bactericidal concentration test results. Mutant prevention concentration test was used to evaluate the resistance tendency suppression potential of the combination. The combination had a synergistic effect against 83.00% of the isolates as verified by the checkerboard and time-kill assays. The combination was defined as bactericidal against all E. coli strains, since minimum bactericidal concentration: minimum inhibitory concentration ratios were below four thresholds and also markedly reduced mutant prevention concentration values of ceftiofur up to 4000-fold compared to its single use. Ceftiofur plus danofloxacin combination inhibited growth of E. coli strains which were resistant to ceftiofur or newer generation of fluoroquinolones. Our results suggest that ceftiofur plus danofloxacin combination has a bactericidal characteristic and can be an important alternative for the treatment of infections caused by resistant E. coli.