Background: The aim of this study was to examine the influence of B-scan averaging on choroidal thickness using wide-field enhanced-depth imaging optical coherence tomography. Methods: Six high-resolution trans-foveal horizontal enhanced-depth imaging line scans (spanning a 60° field) were acquired consecutively from the right eye of 10 healthy adults (mean age 30 ± 5 years), with each line scan an average of 10, 20, 30, 40, 50 or 100 B-scans, using the automated real-time image averaging and follow-up features of a Spectralis device. The impact of B-scan averaging on regional measures of wide-field choroidal thickness (across macular and peripheral regions) and their accuracy was investigated, assuming that averaging 100 B-scans would provide the most accurate estimate of choroidal thickness. Results: Regional estimates of wide-field choroidal thickness did not vary across the different B-scan averaging conditions (all p > 0.05). The mean choroidal thickness averaged across the full wide-field area exhibited the closest agreement to measures obtained with 100 averaged B-scans, when frame averaging exceeded 30 B-scans (95 per cent limits of agreement +10 to −7, +7 to −7 and +6 to −3 μm for 30, 40 and 50 averaged B-scans, respectively), compared to 10 and 20 averaged B-scans (95 per cent limits of agreement +13 to −8 and +13 to −6 μm, respectively; p < 0.01 and p < 0.02 compared to the accuracy of 50 averaged B-scans). Conclusion: Averaging 30 B-scans for an individual enhanced-depth imaging optical coherence tomography line scan provided accurate measures of choroidal thickness across a wide-field (60°) area in young healthy eyes. This information can assist in designing the volumetric scan protocols required for detailed examination of the macular and peripheral choroid.