cryoelectron

Cryoelectron (cryo-EM) refers to cryogenic electron microscopy methods used in structural biology to image biological specimens at cryogenic temperatures, typically by vitrifying samples in water and imaging with a transmission electron microscope. Cryoelectronic imaging aims to preserve native conformation without crystallization, enabling visualization of macromolecules and complexes at near-atomic resolution.

The general workflow involves preparing a thin sample on a grid, rapidly freezing it by plunge or high-pressure methods to form vitreous ice, and transferring the grid to a cold microscope. Images are collected at cryogenic temperatures with direct electron detectors and phase-contrast mechanisms; computational processing aligns thousands to millions of particles to reconstruct a 3D model. Cryo-electron tomography extends this to in situ imaging in cells by tilting the specimen and collecting series of images.

History and impact: Cryo-EM emerged in the late 20th century and gained dramatic improvements in resolution in the 2010s due to direct electron detectors and better software. It has become a standard tool for structural biology, contributing to structures of complex proteins, ribosomes, viral capsids, and membrane assemblies. Nobel Prize in Chemistry 2017 recognized developments in cryo-EM techniques.

Advantages and limitations: Cryo-EM can study large, flexible, or heterogeneous complexes without crystallization and can approach near-atomic resolution. Limitations include difficulty with small proteins, sample heterogeneity, radiation sensitivity, need for expensive instrumentation, and complex data processing requiring substantial computing resources and expertise.

Future directions: continued improvements in detectors, automation, sample preparation, and in situ and time-resolved cryo-EM aim to capture dynamic biological processes and to visualize smaller macromolecules.