JEOL Resources

Atomic resolution structures of biological macromolecules using microED on JEOL TEMs

Wed, 09 Dec 2020 13:23:39 GMT

Also see: JEOL CRYO ARM™ Achieves Top Resolution August 2023.

Micro electron diffraction, or microED, is a technique aimed at solving structures of biological macromolecules by electron diffraction. Barn-storming work by the group from Prof. Gonen showed the impressive impact and promise of this technique¹. The technique borrows from X-ray crystallography in that precession techniques are used for data collection and that much of the well-established software for solving structures by X-ray crystallography can be used for microED. However, it differs in a fundamental way in that electrons are used, which, owing to the substantially larger scattering cross-section of electrons with biological matter, means much smaller crystals can be used. Thus, crystals the size of a few microns are perfectly adequate for microED. The current crop of cameras and levels of microscope automation allow for the rapid collection of a full data set in a matter of minutes. By combining microED with the principle steps of SPA as employed in SerialEM², a single overnight run can yield hundreds of microED data sets³. Note that because diffraction data is collected, the acquisition process is immune to mechanical vibrations and drift; only a shift caused by poor eucentricity that would move the crystal out of the selected aperture field of view would impact the data collection.

MicroED can be applied on every new electron microscope in the JEOL TEM line-up. Precise control of the goniometer’s tilting speed compatible with microED is standard and can be set through either SerialEM or Recorder, the latter being part of JEOL’s TEMography package (SIF). SerialEM was first introduced on a JEOL JEM-2200FS in 2006 and is now fully compatible with all of JEOL’s electron microscopes. A record-breaking 1.34Å structure of apo-ferritin was recently obtained from JEOL’s latest generation of cryo-TEMs, the CRYO ARM™, a microscope available at 200 and 300 kV with a cold field emission gun and in-column energy filter⁴.

This application note reports on microED results obtained on a JEOL JEM-F200 outfitted with a cold FEG and a Gatan Elsa holder to keep the sample at -177ºC for the sake of reducing radiation damage⁵. Figure 1 shows a gallery of principle checks whether crystals of L-histidine will deliver high-resolution diffraction patterns for microED. The micro-crystals are deposited on ultrathin carbon films overlaid on lacey carbon.

Fig. 1: Diffraction images of L-histidine in defocused diffraction mode without (left) and with (middle) a selected area aperture inserted showing the L-histidine crystals. The focused diffraction pattern is shown on the right showing strong Bragg reflections.

Figure 2 shows several frames from one of the microED data sets acquired under continuous rotation at 0.25º/sec from -30º to +30º. The beam stopper is used to prevent the central, undiffracted beam from hitting the camera’s sensor.

Fig. 2: Selected focused diffraction patterns from -30º to +30º microED series acquired at 0.25º/sec. Total dose was 2.4 el/Å2.

After unpacking the movies, the individual frames are processed using standard X-ray crystallographic packages – SIR2019 and SHELXL. Table 1 shows the experimental results whereas Figure 3 shows the structures calculated from the data using direct methods. The refined map after running SHELXL shows clearly the expected atoms in the structure.

Table 1: experimental results
Molecular weight (Da)	155.16
Measurement temperature (K)	96
Space group	P212121
a, b, c (Å)	5.27, 7.44, 18.99
# of reflections	417
R factor (%)	19.81

Fig 3: Structure of L-histidine after initial direct method calculation using SIR2019 (left) and after refinement using SHELXL (middle) with the structure formula shown right.

Conclusion: Micro electron diffraction data can be readily acquired using a JEOL cryo-electron microscope using relatively shallow tilt series. The resulting data can immediately be processed using available pipe lines for X-ray diffraction data enabling rapid structure determination of small molecules by electron diffraction.

Contact your local JEOL representative to learn more about JEOL TEMs and microED using SIF Recorder or SerialEM.

References:

B.L. Nannenga et al., Nature Methods 11 (2014) 927.
D.N. Mastronarde, J. Struct. Biol. 152 (2005) 36.
M.J. de la Cruz et al., Ultramicroscopy 201 (2019) 77.
M. Tegunov et al. (2020) https://doi.org/10.1101/2020.06.05.136341.
Guzmán-Afonso et al., Nature Comm 10 (2019) 3537.

F2: 3D Tomography

Thu, 10 Dec 2020 12:51:19 GMT

The F2 is a new concept of 20-200kV TEM equipped with a Cold FEG. This new generation of multi-purpose electron microscope is designed specifically to meet today’s diversified needs.

Transmission electron microscopy gives access to two-dimensional information coming from the sample. The third dimension is available thanks to the tomography technique. The sample is tilted and for each step, an image (TEM, STEM, EDS) is acquired. Then the 3D volume of the sample can be reconstructed.

The F2 has a “Dual SDD system”, which is composed of two silicon drift detectors (SDD) with large sensor areas, resulting in a total solid angle of 1.7 sr. The combination of two detectors and their placement around the sample, allows the ability to record EDS information throughout the full tilt series (from -80° to +80°).

F2: EDS - Double Your Capabilities

Thu, 10 Dec 2020 12:53:16 GMT

The F2 is a new concept of 20-200kV TEM equipped with a Cold FEG. This new generation of multi-purpose electron microscope is designed specifically to meet today’s diversified needs.

Thanks to the high brightness and small probe size of the Cold FEG, the F2 is able to reach an unprecedented guaranteed resolution for STEM (0.14nm), EDS (1.7sr) and EELS (0.3eV) at the same time, creating a new class of high-end non-corrected TEM.

The F2 has a Dual SDD system, which is composed of two silicon drift detectors (SDD) with large sensor areas, resulting in a total solid angle of 1.7 sr. The combination of dual SDD system, the improved Cold FEG and the advanced scan system, gives the F2 TEM the ability to perform various techniques such as highly sensitive microanalysis and 3D-EDS, atomic resolution EDS maps, high speed mapping, light element mapping (from Be to U), study of beam sensitive samples, and more.

High-resolution analytical systems, such as Transmission Electron Microscopes (TEM) and Scanning Transmission Electron Microscopes (STEM) coupled with high solid-angle EDS system, are attracting increasing attention. Higher resolution and higher efficiency are required for modern systems, along with upgraded and intuitive ease of operation. As the Schottky was not able to meet all these needs, the Cold FEG F2 has been developed as a next-generation analytical electron microscope.

F2: Improved EELS

Thu, 10 Dec 2020 12:55:27 GMT

The F2 is a new concept of 20-200kV TEM equipped with a Cold FEG. This new generation of multi-purpose electron microscope is designed specifically to meet today’s diversified needs.

Thanks to the high brightness and small probe size of the Cold FEG, the F2 is able to reach an unprecedented guaranteed resolution for STEM (0.14nm), EDS (1.7sr) and EELS (0.3eV) at the same time; creating a new class of high-end non-corrected TEM.

Electron Energy Loss Spectroscopy (EELS) is a very powerful technique to characterize materials and especially light elements. The improvement of EELS study is greatly dependent on the TEM performance.

The F2 with Cold FEG dramatically improves the EELS energy resolution. In particular ELNES (Energy-Loss Near-Edge Structure) exhibits a characteristic shape depending on the chemical bonding states in a substance. The small probe size obtainable with Cold FEG improves the F2 resolution, allowing the acquisition of atomic EELS maps.

Coupled with the Cold FEG and the F2’s new DeSCAN system, wide area HAADF images can be analyzed with high energy resolution EELS, such as for surface plasmon resonance.

F2: Multi-purpose Electron Microscope

Thu, 10 Dec 2020 12:57:01 GMT

The F2 is a new concept of 20-200kV TEM. This new generation of multi-purpose electron microscope is designed specifically to meet today's diversified needs.

Thanks to the high brightness and small probe size, the F2 is able to reach an unprecedented guaranteed resolution for STEM (0.14nm), EDS (1.7sr) and EELS (0.3eV) at the same time, creating a new class of high-end non-corrected TEM.

Over the last decades, STEM has emerged as a powerful tool for studying nanomaterials. Many techniques have been developed, Bright Field, Dark Field, High Angle Annular Dark Field, and Annular Bright Field, to characterize materials. By using a STEM detector with a large inner radius, diffraction contrast can be avoided, leading to HAADF image intensity being linked to Z-contrast (atomic number). This allows for even complicated crystal systems, such as Quasicrystals, to be visualized. Coupled with the F2’s new DeSCAN system, wide area HAADF images can be analyzed with high energy resolution EELS.

High-resolution analytical systems, such as transmission electron microscope (TEM) and scanning transmission electron microscope (STEM), are attracting increasing attention. Higher resolution and higher efficiency are required for modern systems, along with upgraded and intuitive ease of operation.

F2: Smart Design - Ergonomics

Thu, 10 Dec 2020 12:58:43 GMT

The F2 is a new concept of 20-200kV TEM equipped with a Cold FEG. This new generation of multi-purpose electron microscope is designed specifically to meet today’s diversified needs.

Thanks to the high brightness and small probe size of the Cold FEG, the F2 is able to reach an unprecedented guaranteed resolution for STEM (0.14nm), EDS (1.7sr) and EELS (0.3eV) at the same time; creating a new class of high-end non-corrected TEM.

The F2 has been designed to fulfil the user’s requirements while maintaining an incredible ease of use.

The F2 incorporates a new, intuitive user interface specifically designed for analytical electron microscopy. This new interface integrates many improvements for the TEM control, such as; automatic functions (auto brightness/contrast, auto focus, auto Z, auto stigmator), coma free auto alignment and off-line data processing (Analysis Center). The software can be tuned at your convenience to provide you with the best working environment.

To facilitate the use of the F2, a new automated sample holder transfer system, the SpecPorter, has been developed to load samples. Once loaded, the PicoStage conducts precise, high speed sample movements of 0.2 nm steps, allowing the operator to move the field of view smoothly over a wide spatial scale range from millimeters to picometers.

The F2 has been developed as a next-generation electron microscope with high performance and ease of use.

Lithium Ion Battery Note

Mon, 04 Mar 2024 10:42:25 GMT

The applications for lithium ion batteries (LIB) cover a wide range, from power sources for personal computers and mobile devices to automobiles, and there is always a demand for even better performance and safety. In order to ensure the performance and quality of LIB, analysis and evaluation using high-performance assessment systems is necessary. JEOL offers a full line-up of equipment to support the development of new LIB technologies and to improve product quality, including instruments for morphology observation and surface analysis, chemical analysis systems to perform structural analysis on a molecular level, as well as fabrication systems to create high-performance coatings and powders. This LIB note offers solutions for researchers and engineers who are looking for the best equipment for their application.

OBF System - Live Low Dose, Light Element Imaging

Fri, 15 Jul 2022 09:21:59 GMT

This brochure is available here.