Imaging Cellular Architecture with 3D SEM

A number of methods for ultrastructural imaging by scanning electron microscopy (SEM) in three dimensions (3D) have been developed over the last decade. These techniques, which we collectively term 3D SEM, offer 2D imaging of quality approaching that attainable by traditional transmission electron microscopy, but with fine resolution in the third dimension as well. In this article, we cover a number of 3D SEM methodologies, including serial block face SEM (SBF-SEM) and focused ion beam SEM (FIB-SEM), as well as other complementary techniques. In addition, we describe the process from preparing a sample to generating a volume, using data from a murine duodenal epithelial cell junction to exemplify the cellular structural detail that may be studied with 3D SEM.