Elmlund, D. & Elmlund, H. Cryogenic electron microscopy and single-particle analysis. Annu Rev Biochem, 2015, 84, 499-517
About 20 years ago, the first three-dimensional (3D) reconstructions at subnanometer (10-Å) resolution of an icosahedral virus assembly were obtained by cryogenic electron microscopy (cryo-EM) and single-particle analysis. Since then, thousands of structures have been determined to resolutions ranging from 30 Å to near atomic (4 Å). Almost overnight, the recent development of direct electron detectors and the attendant improvement in analysis software have advanced the technology considerably. Near-atomic-resolution reconstructions can now be obtained, not only for megadalton macromolecular complexes or highly symmetrical assemblies but also for proteins of only a few hundred kilodaltons. We discuss the developments that led to this breakthrough in high-resolution structure determination by cryo-EM and point to challenges that lie ahead.