To estimate the spatial resolution of microtomographs, a test object on the submicrometer scale was prepared by focused ion beam milling and subjected to microtomographic analysis. Since human tissues are composed of cells and extracellular matrices with micrometer and submicrometer structures, it is important to investigate the three-dimensional spatial resolution of microtomographs used to visualize microstructures of human tissues. The resolutions along the direction within the tomographic slice plane (in-plane resolution) and perpendicular to it (through-plane resolution) were determined from the modulation transfer function of square-wave patterns. The in-plane resolution was estimated to be 1.2 microm from the modulation transfer function of the non-zoom image. In contrast, the zoom image gave the in-plane resolution of 0.8 microm. This in-plane resolution is comparable to the through-plane resolution, which was estimated to be 0.8 microm. Although the two-dimensional radiographs were taken with the pixel width of half the X-ray optics resolution, these three-dimensional resolution analyses indicated that the zoom reconstruction should be performed to achieve the in-plane resolution comparable to the X-ray optics resolution. The submicrometer three-dimensional analysis was applied in the structural study of human cerebral tissue stained with high-Z elements and the obtained tomograms revealed that the microtomographic analysis allows visualization of the subcellular structures of the cerebral tissue.