This review summarizes the recent advances in three-dimensional (3D) imaging techniques and their application to polymer nanostructures, for example, microphase-separated structures of block copolymers. We place particular emphasis on the method of transmission electron microtomography (electron tomography for short; hereafter abbreviated as ET). As a result of recent developments in ET, truly quantitative 3D images of polymer nanostructures can now be obtained with subnanometer resolution. The introduction of scanning optics in ET has made it possible to obtain large amounts of 3D data from micrometer-thick polymer specimens by using conventional electron microscopes at a relatively low accelerating voltage, 200 kV. Thus, ET covers structures over a wide range of thicknesses, from a few nanometers to several hundred nanometers, which corresponds to quite an important spatial range for hierarchical polymer nanostructures. ET provides clear 3D images and a wide range of new structural information that cannot be obtained using other methods. Information traditionally derived from conventional microscopy or scattering methods can be directly acquired from 3D volume data. ET is a versatile technique that is not restricted to only polymer applications; it can also be used as a powerful characterization tool in energy applications such as fuel cells.