Bone strength depends on both mineral content and bone structure. The aim of this in vitro study was to develop a method of quantitatively assessing trabecular bone structure by applying three-dimensional image processing to data acquired with multi-slice and cone-beam computed tomography using micro-computed tomography as a reference. Fifteen bone samples from the radius were examined. After segmentation, quantitative measures of bone volume, trabecular thickness, trabecular separation, trabecular number, trabecular nodes, and trabecular termini were obtained. The clinical machines overestimated bone volume and trabecular thickness and underestimated trabecular nodes and number, but cone-beam CT to a lesser extent. Parameters obtained from cone beam CT were strongly correlated with μCT, with correlation coefficients between 0.93 and 0.98 for all parameters except trabecular termini. The high correlation between cone-beam CT and micro-CT suggest the possibility of quantifying and monitoring changes of trabecular bone microarchitecture in vivo using cone beam CT.