Conventional Electron Microscopy, Cryogenic Electron Microscopy, and Cryogenic Electron Tomography of Viruses

Electron microscopy (EM) techniques have been crucial for understanding the structure of biological specimens such as cells, tissues and macromolecular assemblies. Viruses and related viral assemblies are ideal targets for structural studies that help to define essential biological functions. Whereas conventional EM methods use chemical fixation, dehydration, and staining of the specimens, cryogenic electron microscopy (cryo-EM) preserves the native hydrated state. Combined with image processing and three-dimensional reconstruction techniques, cryo-EM provides three-dimensional maps of these macromolecular complexes from projection images, at atomic or near-atomic resolutions. Cryo-EM is also a major technique in structural biology for dynamic studies of functional complexes, which are often unstable, flexible, scarce, or transient in their native environments. State-of-the-art techniques in structural virology now extend beyond purified symmetric capsids and focus on the asymmetric elements such as the packaged genome and minor structural proteins that were previously missed. As a tool, cryo-EM also complements high-resolution techniques such as X-ray diffraction and NMR spectroscopy; these synergistic hybrid approaches provide important new information. Three-dimensional cryogenic electron tomography (cryo-ET), a variation of cryo-EM, goes further, and allows the study of pleomorphic and complex viruses not only in their physiological state but also in their natural environment in the cell, thereby bridging structural studies at the molecular and cellular levels. Cryo-EM and cryo-ET have been applied successfully in basic research, shedding light on fundamental aspects of virus biology and providing insights into threatening viruses, including SARS-CoV-2, responsible for the COVID-19 pandemic.