Three-dimensional microscopy can be used to reveal the shape, distribution, and connectivity of three-dimensional (3D) features that lie buried within an opaque material. This article discusses several experimental techniques that can be used to generate 3D images. These include serial sectioning, focused ion beam tomography, atom probe tomography, and X-ray microtomography. Nine case studies are presented that represent the work of the various research groups currently working on 3D microscopy using serial sectioning and illustrate the variants of the basic experimental techniques. The article also discusses the techniques for reconstruction and visualization of 3D microstructures with advanced computer software and hardware.