OBJECTIVE: To quantify the test-retest repeatability of mean diffusivity (MD) and fractional anisotropy (FA) derived from diffusion tensor imaging (DTI) tractography in a cohort of paediatric patients with localization-related epilepsy. METHODS: 30 patients underwent 2 DTI acquisitions [repetition time/echo time (ms), 7000/90; flip, 90°; b-value, 1000 s mm(-2); voxel (mm), 2 × 2 × 2]. Two observers used Diffusion Toolkit and TrackVis ( www.trackvis.org ) to segment and analyse the following tracts: corpus callosum, corticospinal tracts, arcuate fasciculi, inferior longitudinal fasciculi and inferior fronto-occipital fasciculi. Mean MD and mean FA were calculated for each tract. Each observer independently analysed one of the DTI data sets for every patient. RESULTS: Segmentation identified all tracts in all subjects, except the arcuate fasciculus. There was a highly consistent relationship between repeated observations of MD (r = 0.993; p < 0.0001) and FA (r = 0.990; p < 0.0001). For each tract, coefficients of variation ranged from 0.9% to 2.1% for MD and from 1.5% to 2.8% for FA. The 95% confidence limits (CLs) for change ranged from 2.8% to 6% for MD and from 4.3% to 8.6% for FA. For the arcuate fasciculus, Cohen's κ for agreement between the observers (identifiable vs not identifiable) was 1.0. CONCLUSION: We quantified the repeatability of two commonly utilized scalar metrics derived from DTI tractography. For an individual patient, changes greater than the repeatability coefficient or 95% CLs for change are unlikely to be related to variability in their measurement. ADVANCES IN KNOWLEDGE: Reproducibility of these metrics will aid in the design of future studies and might one day be used to guide management in patients with epilepsy.