The vertebral endplate and cortical shell play an important structural role and contribute to the overall strength of the vertebral body, are at highest risk of initial failure, and are involved in degenerative disease of the spine. The ability to accurately measure the thickness of these structures is therefore important, even if difficult due to relatively low resolution clinical imaging. We posit that digital tomosynthesis (DTS) may be a suitable imaging modality for measurement of endplate and cortical shell thickness owing to the ability to reconstruct multiplanar images with good spatial resolution at low radiation dose. In this study, for 25 cadaveric L1 vertebrae, average and standard deviation of endplate and cortical shell thickness were measured using images from DTS and microcomputed tomography (μCT). For endplate thickness measurements, significant correlations between DTS and μCT were found for all variables when comparing thicknesses measured in both the overall endplate volume (R² = 0.25–0.54) and when measurements were limited to a central range of coronal or sagittal slices (R² = 0.24–0.62). When compared to reference values from the overall shell volume, DTS thickness measurements were generally nonsignificant. However, when measurement of cortical shell thickness was limited to a range of central slices, DTS outcomes were significantly correlated with reference values for both sagittal and coronal central regions (R² = 0.21–0.49). DTS may therefore offer a means for measurement of endplate thickness and, within a limited sagittal or coronal measurement volume, for measurement of cortical shell thickness.