The introduction of direct electron detectors significantly improved the signal-to-noise ratio of electron micrographs, resulting in a marked enhancement in the quality of cryo-electron microscope (EM) reconstructions (now referred to as the “resolution revolution”).
This approach, along with advances in image processing and data collection, has pushed the resolution of single-particle cryo-EM reconstructions into the range where the electron source could become a limiting factor. This in turn has led to the reconsideration in cryo-EM of alternatives to the standard Schottky field emission gun (FEG), such as the cold FEG, owing to their superior coherence.
Despite the known benefits of the cold FEG over thermally assisted FEG, practical limitations prevented its use in cryo-EM until JEOL’s CryoARM microscope series was developed. Cold FEG was initially used on 300 kV cryomicroscopes to push the frontier of attainable resolution, ultimately resulting in cryo-EM reconstructions at “atomic resolution” where individual atoms are visible as separated spheres of density.
Following this success, attempts have been made at using the cold FEG source on 200 kV cryomicroscopes, which are generally smaller and less expensive, to see how far the tighter energy spread would allow the resolution to be pushed on this more accessible instrument.
In this webinar, the speaker will present results from a cold FEG-equipped CryoARM200 microscope that demonstrates how 200 kV data can be used to produce sub-2Å structures for a range of samples. The resolution of the reconstructions is comparable to that obtained from the more expensive state-of-the-art 300 kV microscopes.
All of the results described in the webinar are from data collected in an automated manner using the open-source software SerialEM. The familiarity that many users have with SerialEM, as well as its overall openness and ease-of-use, make obtaining high-quality data on this platform possible even for users who do not have extensive experience using a CryoARM microscope.
Alan Merk, Electron Microscopist, Leidos Biomedical Research
Alan spent 8 years as microscopist in the lab of Sriram Subramaniam at the Center for Cancer Research, NCI, developing methods for high-resolution data collection on a 300 kV Titan Krios. Alan then joined Jana Ognjenovic’s group at the Frederick National Laboratory, where he works on pushing resolution limits on 200 kV microscopes, including a CryoARM200.
