Present and Emerging Methodologies in Cryo-EM Single-Particle Analysis.

Over the past decade, technical and methodological improvements in cryogenic electron microscopy (cryo-EM) single-particle analysis have enabled routine high-resolution structural analyses of biological macromolecules, resulting in a flood of new molecular insights into protracted biological questions. However, despite the tremendous progress and success of the field in recent years, opportunities for improvement remain in various aspects of the cryo-EM single-particle analysis workflow (e.g., sample preparation, image acquisition and processing, and structure validation). Here, we review recent advances that have contributed to the principal methods in cryo-EM and identify persisting challenges and bottlenecks that will require further methodological and hardware development.