The electron microscopy laboratories at the Frederick National Laboratory explore emerging platforms and develop methodologies to make cryo-EM and volume EM more widely accessible techniques.
Cryo-electron microscopy (cryo-EM) allows high-resolution visualization of macromolecules in solution and near physiological conditions. The success of the approach depends on the quality of specimen preparation, data collection, and image processing. Cryo-EM workflow standardization is needed to enable researchers to obtain high-quality results that are essential to understand structure-function relationships and guide drug design.
As the method is increasingly used by non-specialists, it is important to develop basic standard protocols which will ensure that reproducibility and rigor are maintained. Such protocols will serve as important resources for novice cryo-EM researchers from a broad range of backgrounds.
Our researchers in the National Cryo-EM Facility and the Advanced Cryo-EM Technology Program have created this standardized workflow and protocols for the process.
Standard Operating Procedures
Volume electron microscopy
Recent hardware and software developments as well as workflow advances have resulted in the scanning electron microscope (SEM) being paired with various sectioning approaches to enable imaging of large volumes of resin-embedded cells and tissue samples at high speed and with significant automation. These approaches that allow efficient ultrastructural imaging of µm to mm volumes in 3-D and at nm level resolutions, we refer to collectively as volume EM (vEM) techniques.
The heterogeneity inherent to vEM sample preparation, imaging, image processing and analysis steps both underscores the need for, and also complicates establishment of standards in this emerging field.
Guidelines for vEM may help scientists fully exploit new technologies and resources, increase experimental efficiencies and generate and share reliable insights in their research.