scholarly journals Coma-corrected rapid single-particle cryo-EM data collection on the CRYO ARM 300

2021 ◽  
Vol 77 (5) ◽  
pp. 555-564
Author(s):  
Rouslan G. Efremov ◽  
Annelore Stroobants
Keyword(s):  
High Resolution ◽  
Data Collection ◽  
Single Particle ◽  
Protein Complexes ◽  
Large Area ◽  
Beam Image ◽  
Resolution Electron ◽  
High Resolution Images ◽  
Electron Microscopes ◽  
Data Collection Procedure

Single-particle cryogenic electron microscopy has recently become a major method for determining the structures of proteins and protein complexes. This has markedly increased the demand for throughput of high-resolution electron microscopes, which are required to produce high-resolution images at high rates. An increase in data-collection throughput can be achieved by using large beam-image shifts combined with off-axis coma correction, enabling the acquisition of multiple images from a large area of the EM grid without moving the microscope stage. Here, the optical properties of the JEOL CRYO ARM 300 electron microscope equipped with a K3 camera were characterized under off-axis illumination conditions. It is shown that efficient coma correction can be achieved for beam-image shifts with an amplitude of at least 10 µm, enabling a routine throughput for data collection of between 6000 and 9000 images per day. Use of the benchmark for the rapid data-collection procedure (with beam-image shifts of up to 7 µm) on apoferritin resulted in a reconstruction at a resolution of 1.7 Å. This demonstrates that the rapid automated acquisition of high-resolution micrographs is possible using a CRYO ARM 300.

scholarly journals High-resolution cryo-EM using beam-image shift at 200 keV

2020 ◽  
Author(s):  
Jennifer N. Cash ◽  
Sarah Kearns ◽  
Yilai Li ◽  
Michael A. Cianfrocco
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Data Collection ◽  
Data Quality ◽  
Single Particle ◽  
Aberration Correction ◽  
Cryo Electron Microscopy ◽  
Beam Image ◽  
Recent Advances ◽  
Shift Data

ABSTRACTRecent advances in single-particle cryo-electron microscopy (cryo-EM) data collection utilizes beam-image shift to improve throughput. Despite implementation on 300 keV cryo-EM instruments, it remains unknown how well beam-image shift data collection affects data quality on 200 keV instruments and how much aberrations can be computationally corrected. To test this, we collected and analyzed a cryo-EM dataset of aldolase at 200 keV using beam-image shift. This analysis shows that beam tilt on the instrument initially limited the resolution of aldolase to 4.9Å. After iterative rounds of aberration correction and particle polishing in RELION, we were able to obtain a 2.8Å structure. This analysis demonstrates that software correction of microscope aberrations can provide a significant improvement in resolution at 200 keV.

On Dark-Field High-Resolution Images Of Two-Dimensional Antiphase Structures

1983 ◽  
Vol 21 ◽  
Author(s):  
O. Terasaki ◽  
G.J. Wood ◽  
D. Watanabe
Keyword(s):  
High Resolution ◽  
Dark Field Image ◽  
Antiphase Domain ◽  
Dark Field ◽  
Two Dimensional ◽  
Domain Structures ◽  
Resolution Electron ◽  
True Representation ◽  
High Resolution Images ◽  
Electron Microscopes

ABSTRACTDark-field images of two-dimensional antiphase domain structures with stepped boundaries have been simulated for imaging conditions corresponding to some current high resolution electron microscopes. The simulations reveal that the positions of the dots in the dark-field images do not give a true representation of atomic structure; thus, in contrast to the brightfield superstructure images, it is very misleading to say that bright dots in the dark-field image correspond exactly to the positions of the minority atoms.

scholarly journals Assessing the JEOL CRYO ARM 300 for high-throughput automated single-particle cryo-EM in a multiuser environment

IUCrJ ◽  
2020 ◽  
Vol 7 (4) ◽  
pp. 707-718
Author(s):  
Marcus Fislage ◽  
Alexander V. Shkumatov ◽  
Annelore Stroobants ◽  
Rouslan G. Efremov
Keyword(s):  
High Resolution ◽  
Data Collection ◽  
Structural Biology ◽  
High Throughput ◽  
Single Particle ◽  
Illumination System ◽  
Resolution Electron ◽  
Access Mode ◽  
User Access ◽  
And Performance

Single-particle cryo-EM has become an indispensable structural biology method. It requires regular access to high-resolution electron cryogenic microscopes. To fully utilize the capacity of the expensive high-resolution instruments, the time used for data acquisition and the rate of data collection have to be maximized. This in turn requires high stability and high uptime of the instrument. One of the first 300 kV JEOL CRYO ARM 300 microscopes has been installed at the cryo-EM facility BECM at VIB-VUB, Brussels, where the microscope is used for continuous data collection on multiple projects. Here, the suitability and performance of the microscope is assessed for high-throughput single-particle data collection. In particular, the properties of the illumination system, the stage stability and ice contamination rates are reported. The microscope was benchmarked using mouse heavy-chain apoferritin which was reconstructed to a resolution of 1.9 Å. Finally, uptime and throughput statistics of the instrument accumulated during the first six months of the facility operation in user access mode are reported.

scholarly journals High-resolution cryo-EM using beam-image shift at 200 keV

IUCrJ ◽  
2020 ◽  
Vol 7 (6) ◽  
pp. 1179-1187 ◽  
Author(s):  
Jennifer N. Cash ◽  
Sarah Kearns ◽  
Yilai Li ◽  
Michael A. Cianfrocco
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Data Collection ◽  
Single Particle ◽  
Particle Motion ◽  
Data Set ◽  
Cryo Electron Microscopy ◽  
Beam Image ◽  
Shift Data ◽  
Resolution Structure

Recent advances in single-particle cryo-electron microscopy (cryo-EM) data collection utilize beam-image shift to improve throughput. Despite implementation on 300 keV cryo-EM instruments, it remains unknown how well beam-image-shift data collection affects data quality on 200 keV instruments and the extent to which aberrations can be computationally corrected. To test this, a cryo-EM data set for aldolase was collected at 200 keV using beam-image shift and analyzed. This analysis shows that the instrument beam tilt and particle motion initially limited the resolution to 4.9 Å. After particle polishing and iterative rounds of aberration correction in RELION, a 2.8 Å resolution structure could be obtained. This analysis demonstrates that software correction of microscope aberrations can provide a significant improvement in resolution at 200 keV.

High resolution electron microscopy of molecular crystals I. Quaterrylene, C 40 H 20

1982 ◽  
Vol 381 (1780) ◽  
pp. 225-240 ◽  
Keyword(s):  
High Resolution ◽  
Film Growth ◽  
High Resolution Electron Microscopy ◽  
Boundary Structure ◽  
Grain Boundary Structure ◽  
Resolution Electron ◽  
High Resolution Images ◽  
Electron Microscopes ◽  
Dose Measurements ◽  
Image Simulations

The polynuclear aromatic hydrocarbon quaterrylene, C 40 H 20 , has been studied by using high resolution electron microscopes operating at 100 and 500 kV. Despite the susceptibility of quaterrylene to damage by the incident electron beam, the use of low-dose techniques has allowed a detailed investigation of thin film growth and grain boundary structure in quaterrylene. Comparisons with image simulations establish that high resolution images recorded in the (110)-projection at 500 kV extend to about 0.35 nm, with this figure being limited by radiation damage rather than microscope performance. Moreover, critical dose measurements by electron diffraction establish a good correlation between dose and spot resolution. Extensive crystal distortion has been observed to accommodate mismatch at crystal junctions and boundaries; it is proposed that such distortion arises from individual molecular distortion.

Electron crystallographic determination of the 3-D structure of staurolite at atomic resolution

1991 ◽  
Vol 49 ◽  
pp. 540-541
Author(s):  
Kenneth H. Downing ◽  
Hu Meisheng ◽  
Hans-Rudolf Went ◽  
Michael A. O'Keefe
Keyword(s):  
High Resolution ◽  
Three Dimensional ◽  
High Resolution Electron Microscopy ◽  
Atomic Resolution ◽  
Dimensional Structure ◽  
Structure Information ◽  
Resolution Electron ◽  
Scattering Effects ◽  
High Resolution Images ◽  
Effects Of Radiation

With current advances in electron microscope design, high resolution electron microscopy has become routine, and point resolutions of better than 2Å have been obtained in images of many inorganic crystals. Although this resolution is sufficient to resolve interatomic spacings, interpretation generally requires comparison of experimental images with calculations. Since the images are two-dimensional representations of projections of the full three-dimensional structure, information is invariably lost in the overlapping images of atoms at various heights. The technique of electron crystallography, in which information from several views of a crystal is combined, has been developed to obtain three-dimensional information on proteins. The resolution in images of proteins is severely limited by effects of radiation damage. In principle, atomic-resolution, 3D reconstructions should be obtainable from specimens that are resistant to damage. The most serious problem would appear to be in obtaining high-resolution images from areas that are thin enough that dynamical scattering effects can be ignored.

Atomic structure imaging of the Si/SiO2 interface with high-resolution electron microscopy

1986 ◽  
Vol 44 ◽  
pp. 390-391
Author(s):  
J.L. Batstone ◽  
J.M. Gibson ◽  
Alice.E. White ◽  
K.T. Short
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Atomic Structure ◽  
High Resolution Electron Microscopy ◽  
Medium Voltage ◽  
Voltage Electron ◽  
Resolution Electron ◽  
Structural Image ◽  
Electron Microscopes ◽  
Point To Point

High resolution electron microscopy (HREM) is a powerful tool for the determination of interface atomic structure. With the previous generation of HREM's of point-to-point resolution (rpp) >2.5Å, imaging of semiconductors in only <110> directions was possible. Useful imaging of other important zone axes became available with the advent of high voltage, high resolution microscopes with rpp <1.8Å, leading to a study of the NiSi2 interface. More recently, it was shown that images in <100>, <111> and <112> directions are easily obtainable from Si in the new medium voltage electron microscopes. We report here the examination of the important Si/Si02 interface with the use of a JEOL 4000EX HREM with rpp <1.8Å, in a <100> orientation. This represents a true structural image of this interface.

The Contribution of Intermediate-Voltage, High-Resolution Electron Microscopy In Materials Science: An Appraisal

1990 ◽  
Vol 48 (1) ◽  
pp. 478-479
Author(s):  
John L. Hutchison
Keyword(s):  
High Resolution ◽  
Materials Science ◽  
High Resolution Electron Microscopy ◽  
Optical Diffraction ◽  
The Past ◽  
Resolution Electron ◽  
Extreme Sensitivity ◽  
Electron Microscopes ◽  
New Generation ◽  
Small Metal Particles

Over the past five years or so the development of a new generation of high resolution electron microscopes operating routinely in the 300-400 kilovolt range has produced a dramatic increase in resolution, to around 1.6 Å for “structure resolution” and approaching 1.2 Å for information limits. With a large number of such instruments now in operation it is timely to assess their impact in the various areas of materials science where they are now being used. Are they falling short of the early expectations? Generally, the manufacturers’ claims regarding resolution are being met, but one unexpected factor which has emerged is the extreme sensitivity of these instruments to both floor-borne and acoustic vibrations. Successful measures to counteract these disturbances may require the use of special anti-vibration blocks, or even simple oil-filled dampers together with springs, with heavy curtaining around the microscope room to reduce noise levels. In assessing performance levels, optical diffraction analysis is becoming the accepted method, with rotational averaging useful for obtaining a good measure of information limits. It is worth noting here that microscope alignment becomes very critical for the highest resolution.In attempting an appraisal of the contributions of intermediate voltage HREMs to materials science we will outline a few of the areas where they are most widely used. These include semiconductors, oxides, and small metal particles, in addition to metals and minerals.

The Contribution of Phonon-Scattered Electrons to High-Resolution Electron Micrographs of Crystals

1990 ◽  
Vol 48 (1) ◽  
pp. 62-63
Author(s):  
H. Kohl
Keyword(s):  
High Resolution ◽  
Spherical Aberration ◽  
High Resolution Electron Microscopy ◽  
Static Potential ◽  
Qualitative Comparison ◽  
The Past ◽  
Resolution Electron ◽  
High Resolution Images ◽  
High Resolution Electron Micrographs ◽  
Extract Information

High-Resolution Electron Microscopy is able to determine structures of crystals and interfaces with a spatial resolution of somewhat less than 2 Å. As the image is strongly dependent on instrumental parameters, notably the defocus and the spherical aberration, the interpretation of micrographs necessitates a comparison with calculated images. Whereas one has often been content with a qualitative comparison of theory with experiment in the past, one is currently striving for quantitative procedures to extract information from the images [1,2]. For the calculations one starts by assuming a static potential, thus neglecting inelastic scattering processes.We shall confine the discussion to periodic specimens. All electrons, which have only been elastically scattered, are confined to very few directions, the Bragg spots. In-elastically scattered electrons, however, can be found in any direction. Therefore the influence of inelastic processes on the elastically (= Bragg) scattered electrons can be described as an attenuation [3]. For the calculation of high-resolution images this procedure would be correct only if we had an imaging energy filter capable of removing all phonon-scattered electrons. This is not realizable in practice. We are therefore forced to include the contribution of the phonon-scattered electrons.

Combined Experimentaland Theoretical Determination of the Atomic Structure of the (310) Twin in NB

1991 ◽  
Vol 238 ◽  
Author(s):  
Geoffrey H. Campbells ◽  
Wayne E. King ◽  
Stephen M. Foiles ◽  
Peter Gumbsch ◽  
Manfred Rühle
Keyword(s):  
High Resolution ◽  
Theoretical Models ◽  
High Resolution Electron Microscopy ◽  
Interatomic Potentials ◽  
Image Simulation ◽  
Theoretical Determination ◽  
Atomic Structures ◽  
Resolution Electron ◽  
High Resolution Images

ABSTRACTA (310) twin boundary in Nb has been fabricated by diffusion bonding oriented single crystals and characterized using high resolution electron microscopy. Atomic structures for the boundary have been predicted using different interatomic potentials. Comparison of the theoretical models to the high resolution images has been performed through image simulation. On the basis of this comparison, one of the low energy structures predicted by theory can be ruled out.

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