Initial dosing and dose adjustment of intravenous tobramycin in children with cystic fibrosis (CF) is challenging. The objectives of this study were to develop nonparametric population pharmacokinetic (PK) models of tobramycin in children with CF to be used for dosage design and model-guided therapeutic drug monitoring. We performed a retrospective analysis of tobramycin PK data in our children's CF center. The Pmetrics package was used for nonparametric population PK analysis and dosing simulations. Both the ratios of maximal concentration to the MIC (Cmax/MIC) and daily area under the concentration-time curve to the MIC (AUC24/MIC) were considered efficacy targets. Trough concentration (Cmin) was considered the safety target. A total of 2,884 tobramycin concentrations collected in 195 patients over 9 years were analyzed. A two-compartment model including total body weight, body surface area, and creatinine clearance as covariates best described the data. A simpler model was also derived for implementation in the BestDose software to perform Bayesian dose adjustment. Both models were externally validated. PK/pharmacodynamics (PD) simulations with the final model suggest that an initial dose of tobramycin of 15 to 17.5 mg/kg/day was necessary to achieve Cmax/MICs of ≥10 for MICs up to 2 mg/liter in most patients. The AUC24/MIC target was associated with higher dosage requirements and higher Cmin. A daily dose of 12.5 mg/kg would optimize both efficacy and safety target attainment. We recommend performing tobramycin therapeutic drug monitoring (TDM), model-based dose adjustment, and MIC determination to individualize intravenous tobramycin therapy in children with CF.