Direct observation of iron-induced conformational changes of mitochondrial DNA by high-resolution field-emission in-lens scanning electron microscopy.

PubMed Central

Yaffee, M; Walter, P; Richter, C; Müller, M

1996-01-01

When respiring rat liver mitochondria are incubated in the presence of Fe(III) gluconate, their DNA (mtDNA) relaxes from the supercoiled to the open circular form dependent on the iron dose. Anaerobiosis or antioxidants fail to completely inhibit the unwinding. High-resolution field-emission in-lens scanning electron microscopy imaging, in concert with backscattered electron detection, pinpoints nanometer-range iron colloids bound to mtDNA isolated from iron-exposed mitochondria. High-resolution field-emission in-lens scanning electron microscopy with backscattered electron detection imaging permits simultaneous detailed visual analysis of DNA topology, iron dose-dependent mtDNA unwinding, and assessment of iron colloid formation on mtDNA strands. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8643576

Imaging of endodontic biofilms by combined microscopy (FISH/cLSM - SEM).

PubMed

Schaudinn, C; Carr, G; Gorur, A; Jaramillo, D; Costerton, J W; Webster, P

2009-08-01

Scanning electron microscopy is a useful imaging approach for the visualization of bacterial biofilms in their natural environments including their medical and dental habitats, because it allows for the exploration of large surfaces with excellent resolution of topographic features. Most biofilms in nature, however, are embedded in a thick layer of extracellular matrix that prevents a clear identification of individual bacteria by scanning electron microscopy. The use of confocal laser scanning microscopy on the other hand in combination with fluorescence in situ hybridization enables the visualization of matrix embedded bacteria in multi-layered biofilms. In our study, fluorescence in situ hybridization/confocal laser scanning microscopy and scanning electron microscopy were applied to visualize bacterial biofilm in endodontic root canals. The resulting fluorescence in situ hybridization /confocal laser scanning microscopy and scanning electron microscopy and pictures were subsequently combined into one single image to provide high-resolution information on the location of hidden bacteria. The combined use of scanning electron microscopy and fluorescence in situ hybridization / confocal laser scanning microscopy has the potential to overcome the limits of each single technique.

Chemistry Viewed through the Eyes of High-Resolution Microscopy.

ERIC Educational Resources Information Center

Beer, Michael; And Others

1981-01-01

This special report, prepared by several chemists working in the field of electron microscopy, provides information regarding the most recent developments in transmission and scanning electron microscopy that have chemical significance. (CS)

Ultrafast Science Opportunities with Electron Microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Durr, Hermann

X-rays and electrons are two of the most fundamental probes of matter. When the Linac Coherent Light Source (LCLS), the world’s first x-ray free electron laser, began operation in 2009, it transformed ultrafast science with the ability to generate laser-like x-ray pulses from the manipulation of relativistic electron beams. This document describes a similar future transformation. In Transmission Electron Microscopy, ultrafast relativistic (MeV energy) electron pulses can achieve unsurpassed spatial and temporal resolution. Ultrafast temporal resolution will be the next frontier in electron microscopy and can ideally complement ultrafast x-ray science done with free electron lasers. This document describes themore » Grand Challenge science opportunities in chemistry, material science, physics and biology that arise from an MeV ultrafast electron diffraction & microscopy facility, especially when coupled with linac-based intense THz and X-ray pump capabilities.« less

Contributed review: Review of integrated correlative light and electron microscopy.

PubMed

Timmermans, F J; Otto, C

2015-01-01

New developments in the field of microscopy enable to acquire increasing amounts of information from large sample areas and at an increased resolution. Depending on the nature of the technique, the information may reveal morphological, structural, chemical, and still other sample characteristics. In research fields, such as cell biology and materials science, there is an increasing demand to correlate these individual levels of information and in this way to obtain a better understanding of sample preparation and specific sample properties. To address this need, integrated systems were developed that combine nanometer resolution electron microscopes with optical microscopes, which produce chemically or label specific information through spectroscopy. The complementary information from electron microscopy and light microscopy presents an opportunity to investigate a broad range of sample properties in a correlated fashion. An important part of correlating the differences in information lies in bridging the different resolution and image contrast features. The trend to analyse samples using multiple correlated microscopes has resulted in a new research field. Current research is focused, for instance, on (a) the investigation of samples with nanometer scale distribution of inorganic and organic materials, (b) live cell analysis combined with electron microscopy, and (c) in situ spectroscopic and electron microscopy analysis of catalytic materials, but more areas will benefit from integrated correlative microscopy.

Electron Microscopy Localization and Characterization of Functionalized Composite Organic-Inorganic SERS Nanoparticles on Leukemia Cells

PubMed Central

Koh, Ai Leen; Shachaf, Catherine M.; Elchuri, Sailaja; Nolan, Garry P.; Sinclair, Robert

2008-01-01

We demonstrate the use of electron microscopy as a powerful characterization tool to identify and locate antibody-conjugated composite organic-inorganic (COINs) surface enhanced Raman scattering (SERS) nanoparticles on cells. U937 leukemia cells labeled with antibody CD54-conjugated COINs were characterized in their native, hydrated state using wet Scanning Electron Microscopy (SEM) and in their dehydrated state using high-resolution SEM. In both cases, the backscattered electron detector (BSE) was used to detect and identify the silver constituents in COINs due to its high sensitivity to atomic number variations within a specimen. The imaging and analytical capabilities in the SEM were further complemented by higher resolution Transmission Electron Microscope (TEM) images and Scanning Auger Electron Spectroscopy (AES) data to give reliable and high-resolution information about nanoparticles and their binding to cell surface antigens. PMID:18995965

New and unconventional approaches for advancing resolution in biological transmission electron microscopy by improving macromolecular specimen preparation and preservation.

PubMed

Massover, William H

2011-02-01

Resolution in transmission electron microscopy (TEM) now is limited by the properties of specimens, rather than by those of instrumentation. The long-standing difficulties in obtaining truly high-resolution structure from biological macromolecules with TEM demand the development, testing, and application of new ideas and unconventional approaches. This review concisely describes some new concepts and innovative methodologies for TEM that deal with unsolved problems in the preparation and preservation of macromolecular specimens. The selected topics include use of better support films, a more protective multi-component matrix surrounding specimens for cryo-TEM and negative staining, and, several quite different changes in microscopy and micrography that should decrease the effects of electron radiation damage; all these practical approaches are non-traditional, but have promise to advance resolution for specimens of biological macromolecules beyond its present level of 3-10 Å (0.3-1.0 nm). The result of achieving truly high resolution will be a fulfillment of the still unrealized potential of transmission electron microscopy for directly revealing the structure of biological macromolecules down to the atomic level. Published by Elsevier Ltd.

Neuroanatomy from Mesoscopic to Nanoscopic Scales: An Improved Method for the Observation of Semithin Sections by High-Resolution Scanning Electron Microscopy

PubMed Central

Rodríguez, José-Rodrigo; Turégano-López, Marta; DeFelipe, Javier; Merchán-Pérez, Angel

2018-01-01

Semithin sections are commonly used to examine large areas of tissue with an optical microscope, in order to locate and trim the regions that will later be studied with the electron microscope. Ideally, the observation of semithin sections would be from mesoscopic to nanoscopic scales directly, instead of using light microscopy and then electron microscopy (EM). Here we propose a method that makes it possible to obtain high-resolution scanning EM images of large areas of the brain in the millimeter to nanometer range. Since our method is compatible with light microscopy, it is also feasible to generate hybrid light and electron microscopic maps. Additionally, the same tissue blocks that have been used to obtain semithin sections can later be used, if necessary, for transmission EM, or for focused ion beam milling and scanning electron microscopy (FIB-SEM). PMID:29568263

Neuroanatomy from Mesoscopic to Nanoscopic Scales: An Improved Method for the Observation of Semithin Sections by High-Resolution Scanning Electron Microscopy.

PubMed

Rodríguez, José-Rodrigo; Turégano-López, Marta; DeFelipe, Javier; Merchán-Pérez, Angel

2018-01-01

Semithin sections are commonly used to examine large areas of tissue with an optical microscope, in order to locate and trim the regions that will later be studied with the electron microscope. Ideally, the observation of semithin sections would be from mesoscopic to nanoscopic scales directly, instead of using light microscopy and then electron microscopy (EM). Here we propose a method that makes it possible to obtain high-resolution scanning EM images of large areas of the brain in the millimeter to nanometer range. Since our method is compatible with light microscopy, it is also feasible to generate hybrid light and electron microscopic maps. Additionally, the same tissue blocks that have been used to obtain semithin sections can later be used, if necessary, for transmission EM, or for focused ion beam milling and scanning electron microscopy (FIB-SEM).

High-resolution low-dose scanning transmission electron microscopy.

PubMed

Buban, James P; Ramasse, Quentin; Gipson, Bryant; Browning, Nigel D; Stahlberg, Henning

2010-01-01

During the past two decades instrumentation in scanning transmission electron microscopy (STEM) has pushed toward higher intensity electron probes to increase the signal-to-noise ratio of recorded images. While this is suitable for robust specimens, biological specimens require a much reduced electron dose for high-resolution imaging. We describe here protocols for low-dose STEM image recording with a conventional field-emission gun STEM, while maintaining the high-resolution capability of the instrument. Our findings show that a combination of reduced pixel dwell time and reduced gun current can achieve radiation doses comparable to low-dose TEM.

In-situ Isotopic Analysis at Nanoscale using Parallel Ion Electron Spectrometry: A Powerful New Paradigm for Correlative Microscopy

NASA Astrophysics Data System (ADS)

Yedra, Lluís; Eswara, Santhana; Dowsett, David; Wirtz, Tom

2016-06-01

Isotopic analysis is of paramount importance across the entire gamut of scientific research. To advance the frontiers of knowledge, a technique for nanoscale isotopic analysis is indispensable. Secondary Ion Mass Spectrometry (SIMS) is a well-established technique for analyzing isotopes, but its spatial-resolution is fundamentally limited. Transmission Electron Microscopy (TEM) is a well-known method for high-resolution imaging down to the atomic scale. However, isotopic analysis in TEM is not possible. Here, we introduce a powerful new paradigm for in-situ correlative microscopy called the Parallel Ion Electron Spectrometry by synergizing SIMS with TEM. We demonstrate this technique by distinguishing lithium carbonate nanoparticles according to the isotopic label of lithium, viz. 6Li and 7Li and imaging them at high-resolution by TEM, adding a new dimension to correlative microscopy.

Live CLEM imaging to analyze nuclear structures at high resolution.

PubMed

Haraguchi, Tokuko; Osakada, Hiroko; Koujin, Takako

2015-01-01

Fluorescence microscopy (FM) and electron microscopy (EM) are powerful tools for observing molecular components in cells. FM can provide temporal information about cellular proteins and structures in living cells. EM provides nanometer resolution images of cellular structures in fixed cells. We have combined FM and EM to develop a new method of correlative light and electron microscopy (CLEM), called "Live CLEM." In this method, the dynamic behavior of specific molecules of interest is first observed in living cells using fluorescence microscopy (FM) and then cellular structures in the same cell are observed using electron microscopy (EM). Following image acquisition, FM and EM images are compared to enable the fluorescent images to be correlated with the high-resolution images of cellular structures obtained using EM. As this method enables analysis of dynamic events involving specific molecules of interest in the context of specific cellular structures at high resolution, it is useful for the study of nuclear structures including nuclear bodies. Here we describe Live CLEM that can be applied to the study of nuclear structures in mammalian cells.

Possibilities and limitations of advanced transmission electron microscopy for carbon-based nanomaterials

PubMed Central

Bittencourt, Carla; Van Tendeloo, Gustaaf

2015-01-01

Summary A major revolution for electron microscopy in the past decade is the introduction of aberration correction, which enables one to increase both the spatial resolution and the energy resolution to the optical limit. Aberration correction has contributed significantly to the imaging at low operating voltages. This is crucial for carbon-based nanomaterials which are sensitive to electron irradiation. The research of carbon nanomaterials and nanohybrids, in particular the fundamental understanding of defects and interfaces, can now be carried out in unprecedented detail by aberration-corrected transmission electron microscopy (AC-TEM). This review discusses new possibilities and limits of AC-TEM at low voltage, including the structural imaging at atomic resolution, in three dimensions and spectroscopic investigation of chemistry and bonding. In situ TEM of carbon-based nanomaterials is discussed and illustrated through recent reports with particular emphasis on the underlying physics of interactions between electrons and carbon atoms. PMID:26425406

Atomic-resolution transmission electron microscopy of electron beam–sensitive crystalline materials

NASA Astrophysics Data System (ADS)

Zhang, Daliang; Zhu, Yihan; Liu, Lingmei; Ying, Xiangrong; Hsiung, Chia-En; Sougrat, Rachid; Li, Kun; Han, Yu

2018-02-01

High-resolution imaging of electron beam–sensitive materials is one of the most difficult applications of transmission electron microscopy (TEM). The challenges are manifold, including the acquisition of images with extremely low beam doses, the time-constrained search for crystal zone axes, the precise image alignment, and the accurate determination of the defocus value. We develop a suite of methods to fulfill these requirements and acquire atomic-resolution TEM images of several metal organic frameworks that are generally recognized as highly sensitive to electron beams. The high image resolution allows us to identify individual metal atomic columns, various types of surface termination, and benzene rings in the organic linkers. We also apply our methods to other electron beam–sensitive materials, including the organic-inorganic hybrid perovskite CH3NH3PbBr3.

On the Progress of Scanning Transmission Electron Microscopy (STEM) Imaging in a Scanning Electron Microscope.

PubMed

Sun, Cheng; Müller, Erich; Meffert, Matthias; Gerthsen, Dagmar

2018-04-01

Transmission electron microscopy (TEM) with low-energy electrons has been recognized as an important addition to the family of electron microscopies as it may avoid knock-on damage and increase the contrast of weakly scattering objects. Scanning electron microscopes (SEMs) are well suited for low-energy electron microscopy with maximum electron energies of 30 keV, but they are mainly used for topography imaging of bulk samples. Implementation of a scanning transmission electron microscopy (STEM) detector and a charge-coupled-device camera for the acquisition of on-axis transmission electron diffraction (TED) patterns, in combination with recent resolution improvements, make SEMs highly interesting for structure analysis of some electron-transparent specimens which are traditionally investigated by TEM. A new aspect is correlative SEM, STEM, and TED imaging from the same specimen region in a SEM which leads to a wealth of information. Simultaneous image acquisition gives information on surface topography, inner structure including crystal defects and qualitative material contrast. Lattice-fringe resolution is obtained in bright-field STEM imaging. The benefits of correlative SEM/STEM/TED imaging in a SEM are exemplified by structure analyses from representative sample classes such as nanoparticulates and bulk materials.

Breaking resolution limits in ultrafast electron diffraction and microscopy.

PubMed

Baum, Peter; Zewail, Ahmed H

2006-10-31

Ultrafast electron microscopy and diffraction are powerful techniques for the study of the time-resolved structures of molecules, materials, and biological systems. Central to these approaches is the use of ultrafast coherent electron packets. The electron pulses typically have an energy of 30 keV for diffraction and 100-200 keV for microscopy, corresponding to speeds of 33-70% of the speed of light. Although the spatial resolution can reach the atomic scale, the temporal resolution is limited by the pulse width and by the difference in group velocities of electrons and the light used to initiate the dynamical change. In this contribution, we introduce the concept of tilted optical pulses into diffraction and imaging techniques and demonstrate the methodology experimentally. These advances allow us to reach limits of time resolution down to regimes of a few femtoseconds and, possibly, attoseconds. With tilted pulses, every part of the sample is excited at precisely the same time as when the electrons arrive at the specimen. Here, this approach is demonstrated for the most unfavorable case of ultrafast crystallography. We also present a method for measuring the duration of electron packets by autocorrelating electron pulses in free space and without streaking, and we discuss the potential of tilting the electron pulses themselves for applications in domains involving nuclear and electron motions.

SRRF: Universal live-cell super-resolution microscopy.

PubMed

Culley, Siân; Tosheva, Kalina L; Matos Pereira, Pedro; Henriques, Ricardo

2018-08-01

Super-resolution microscopy techniques break the diffraction limit of conventional optical microscopy to achieve resolutions approaching tens of nanometres. The major advantage of such techniques is that they provide resolutions close to those obtainable with electron microscopy while maintaining the benefits of light microscopy such as a wide palette of high specificity molecular labels, straightforward sample preparation and live-cell compatibility. Despite this, the application of super-resolution microscopy to dynamic, living samples has thus far been limited and often requires specialised, complex hardware. Here we demonstrate how a novel analytical approach, Super-Resolution Radial Fluctuations (SRRF), is able to make live-cell super-resolution microscopy accessible to a wider range of researchers. We show its applicability to live samples expressing GFP using commercial confocal as well as laser- and LED-based widefield microscopes, with the latter achieving long-term timelapse imaging with minimal photobleaching. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

Electron microscopy of electromagnetic waveforms.

PubMed

Ryabov, A; Baum, P

2016-07-22

Rapidly changing electromagnetic fields are the basis of almost any photonic or electronic device operation. We report how electron microscopy can measure collective carrier motion and fields with subcycle and subwavelength resolution. A collimated beam of femtosecond electron pulses passes through a metamaterial resonator that is previously excited with a single-cycle electromagnetic pulse. If the probing electrons are shorter in duration than half a field cycle, then time-frozen Lorentz forces distort the images quasi-classically and with subcycle time resolution. A pump-probe sequence reveals in a movie the sample's oscillating electromagnetic field vectors with time, phase, amplitude, and polarization information. This waveform electron microscopy can be used to visualize electrodynamic phenomena in devices as small and fast as available. Copyright © 2016, American Association for the Advancement of Science.

4D electron microscopy: principles and applications.

PubMed

Flannigan, David J; Zewail, Ahmed H

2012-10-16

The transmission electron microscope (TEM) is a powerful tool enabling the visualization of atoms with length scales smaller than the Bohr radius at a factor of only 20 larger than the relativistic electron wavelength of 2.5 pm at 200 keV. The ability to visualize matter at these scales in a TEM is largely due to the efforts made in correcting for the imperfections in the lens systems which introduce aberrations and ultimately limit the achievable spatial resolution. In addition to the progress made in increasing the spatial resolution, the TEM has become an all-in-one characterization tool. Indeed, most of the properties of a material can be directly mapped in the TEM, including the composition, structure, bonding, morphology, and defects. The scope of applications spans essentially all of the physical sciences and includes biology. Until recently, however, high resolution visualization of structural changes occurring on sub-millisecond time scales was not possible. In order to reach the ultrashort temporal domain within which fundamental atomic motions take place, while simultaneously retaining high spatial resolution, an entirely new approach from that of millisecond-limited TEM cameras had to be conceived. As shown below, the approach is also different from that of nanosecond-limited TEM, whose resolution cannot offer the ultrafast regimes of dynamics. For this reason "ultrafast electron microscopy" is reserved for the field which is concerned with femtosecond to picosecond resolution capability of structural dynamics. In conventional TEMs, electrons are produced by heating a source or by applying a strong extraction field. Both methods result in the stochastic emission of electrons, with no control over temporal spacing or relative arrival time at the specimen. The timing issue can be overcome by exploiting the photoelectric effect and using pulsed lasers to generate precisely timed electron packets of ultrashort duration. The spatial and temporal resolutions achievable with short intense pulses containing a large number of electrons, however, are limited to tens of nanometers and nanoseconds, respectively. This is because Coulomb repulsion is significant in such a pulse, and the electrons spread in space and time, thus limiting the beam coherence. It is therefore not possible to image the ultrafast elementary dynamics of complex transformations. The challenge was to retain the high spatial resolution of a conventional TEM while simultaneously enabling the temporal resolution required to visualize atomic-scale motions. In this Account, we discuss the development of four-dimensional ultrafast electron microscopy (4D UEM) and summarize techniques and applications that illustrate the power of the approach. In UEM, images are obtained either stroboscopically with coherent single-electron packets or with a single electron bunch. Coulomb repulsion is absent under the single-electron condition, thus permitting imaging, diffraction, and spectroscopy, all with high spatiotemporal resolution, the atomic scale (sub-nanometer and femtosecond). The time resolution is limited only by the laser pulse duration and energy carried by the electron packets; the CCD camera has no bearing on the temporal resolution. In the regime of single pulses of electrons, the temporal resolution of picoseconds can be attained when hundreds of electrons are in the bunch. The applications given here are selected to highlight phenomena of different length and time scales, from atomic motions during structural dynamics to phase transitions and nanomechanical oscillations. We conclude with a brief discussion of emerging methods, which include scanning ultrafast electron microscopy (S-UEM), scanning transmission ultrafast electron microscopy (ST-UEM) with convergent beams, and time-resolved imaging of biological structures at ambient conditions with environmental cells.

Synthesis, Characterization, and Application of Gold Nanoparticles in Green Nanochemistry Dye-Sensitized Solar Cells

DTIC Science & Technology

2012-06-01

resolution tunneling electron microscopy (HR-TEM). 2.4 DSSC Assembly Annealed TiO2 nanoparticle photoanodes were placed into 10 mL each of the blackberry ...resolution tunneling electron microscopy, and ultraviolet-visible spectroscopy. After characterization, the NPs were found to vary in shape but had... Blackberry Anthocyanin Extraction Procedure ...............................................................3 2.3 Au Nanoparticle Synthesis

Chemical mapping and quantification at the atomic scale by scanning transmission electron microscopy.

PubMed

Chu, Ming-Wen; Chen, Cheng Hsuan

2013-06-25

With innovative modern material-growth methods, a broad spectrum of fascinating materials with reduced dimensions-ranging from single-atom catalysts, nanoplasmonic and nanophotonic materials to two-dimensional heterostructural interfaces-is continually emerging and extending the new frontiers of materials research. A persistent central challenge in this grand scientific context has been the detailed characterization of the individual objects in these materials with the highest spatial resolution, a problem prompting the need for experimental techniques that integrate both microscopic and spectroscopic capabilities. To date, several representative microscopy-spectroscopy combinations have become available, such as scanning tunneling microscopy, tip-enhanced scanning optical microscopy, atom probe tomography, scanning transmission X-ray microscopy, and scanning transmission electron microscopy (STEM). Among these tools, STEM boasts unique chemical and electronic sensitivity at unparalleled resolution. In this Perspective, we elucidate the advances in STEM and chemical mapping applications at the atomic scale by energy-dispersive X-ray spectroscopy and electron energy loss spectroscopy with a focus on the ultimate challenge of chemical quantification with atomic accuracy.

Serial block face scanning electron microscopy--the future of cell ultrastructure imaging.

PubMed

Hughes, Louise; Hawes, Chris; Monteith, Sandy; Vaughan, Sue

2014-03-01

One of the major drawbacks in transmission electron microscopy has been the production of three-dimensional views of cells and tissues. Currently, there is no one suitable 3D microscopy technique that answers all questions and serial block face scanning electron microscopy (SEM) fills the gap between 3D imaging using high-end fluorescence microscopy and the high resolution offered by electron tomography. In this review, we discuss the potential of the serial block face SEM technique for studying the three-dimensional organisation of animal, plant and microbial cells.

Liquid scanning transmission electron microscopy: imaging protein complexes in their native environment in whole eukaryotic cells.

PubMed

Peckys, Diana B; de Jonge, Niels

2014-04-01

Scanning transmission electron microscopy (STEM) of specimens in liquid, so-called Liquid STEM, is capable of imaging the individual subunits of macromolecular complexes in whole eukaryotic cells in liquid. This paper discusses this new microscopy modality within the context of state-of-the-art microscopy of cells. The principle of operation and equations for the resolution are described. The obtained images are different from those acquired with standard transmission electron microscopy showing the cellular ultrastructure. Instead, contrast is obtained on specific labels. Images can be recorded in two ways, either via STEM at 200 keV electron beam energy using a microfluidic chamber enclosing the cells, or via environmental scanning electron microscopy at 30 keV of cells in a wet environment. The first series of experiments involved the epidermal growth factor receptor labeled with gold nanoparticles. The labels were imaged in whole fixed cells with nanometer resolution. Since the cells can be kept alive in the microfluidic chamber, it is also feasible to detect the labels in unfixed, live cells. The rapid sample preparation and imaging allows studies of multiple whole cells.

Fabrication of bright and thin Zn₂SiO₄ luminescent film for electron beam excitation-assisted optical microscope.

PubMed

Furukawa, Taichi; Kanamori, Satoshi; Fukuta, Masahiro; Nawa, Yasunori; Kominami, Hiroko; Nakanishi, Yoichiro; Sugita, Atsushi; Inami, Wataru; Kawata, Yoshimasa

2015-07-13

We fabricated a bright and thin Zn₂SiO₄ luminescent film to serve as a nanometric light source for high-spatial-resolution optical microscopy based on electron beam excitation. The Zn₂SiO₄ luminescent thin film was fabricated by annealing a ZnO film on a Si₃N₄ substrate at 1000 °C in N₂. The annealed film emitted bright cathodoluminescence compared with the as-deposited film. The film is promising for nano-imaging with electron beam excitation-assisted optical microscopy. We evaluated the spatial resolution of a microscope developed using this Zn₂SiO₄ luminescent thin film. This is the first report of the investigation and application of ZnO/Si₃N₄ annealed at a high temperature (1000 °C). The fabricated Zn₂SiO₄ film is expected to enable high-frame-rate dynamic observation with ultra-high resolution using our electron beam excitation-assisted optical microscopy.

Imaging single atoms using secondary electrons with an aberration-corrected electron microscope.

PubMed

Zhu, Y; Inada, H; Nakamura, K; Wall, J

2009-10-01

Aberration correction has embarked on a new frontier in electron microscopy by overcoming the limitations of conventional round lenses, providing sub-angstrom-sized probes. However, improvement of spatial resolution using aberration correction so far has been limited to the use of transmitted electrons both in scanning and stationary mode, with an improvement of 20-40% (refs 3-8). In contrast, advances in the spatial resolution of scanning electron microscopes (SEMs), which are by far the most widely used instrument for surface imaging at the micrometre-nanometre scale, have been stagnant, despite several recent efforts. Here, we report a new SEM, with aberration correction, able to image single atoms by detecting electrons emerging from its surface as a result of interaction with the small probe. The spatial resolution achieved represents a fourfold improvement over the best-reported resolution in any SEM (refs 10-12). Furthermore, we can simultaneously probe the sample through its entire thickness with transmitted electrons. This ability is significant because it permits the selective visualization of bulk atoms and surface ones, beyond a traditional two-dimensional projection in transmission electron microscopy. It has the potential to revolutionize the field of microscopy and imaging, thereby opening the door to a wide range of applications, especially when combined with simultaneous nanoprobe spectroscopy.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Timmermans, F. J.; Otto, C.

New developments in the field of microscopy enable to acquire increasing amounts of information from large sample areas and at an increased resolution. Depending on the nature of the technique, the information may reveal morphological, structural, chemical, and still other sample characteristics. In research fields, such as cell biology and materials science, there is an increasing demand to correlate these individual levels of information and in this way to obtain a better understanding of sample preparation and specific sample properties. To address this need, integrated systems were developed that combine nanometer resolution electron microscopes with optical microscopes, which produce chemicallymore » or label specific information through spectroscopy. The complementary information from electron microscopy and light microscopy presents an opportunity to investigate a broad range of sample properties in a correlated fashion. An important part of correlating the differences in information lies in bridging the different resolution and image contrast features. The trend to analyse samples using multiple correlated microscopes has resulted in a new research field. Current research is focused, for instance, on (a) the investigation of samples with nanometer scale distribution of inorganic and organic materials, (b) live cell analysis combined with electron microscopy, and (c) in situ spectroscopic and electron microscopy analysis of catalytic materials, but more areas will benefit from integrated correlative microscopy.« less

Dose-dependent high-resolution electron ptychography

NASA Astrophysics Data System (ADS)

D'Alfonso, A. J.; Allen, L. J.; Sawada, H.; Kirkland, A. I.

2016-02-01

Recent reports of electron ptychography at atomic resolution have ushered in a new era of coherent diffractive imaging in the context of electron microscopy. We report and discuss electron ptychography under variable electron dose conditions, exploring the prospects of an approach which has considerable potential for imaging where low dose is needed.

Multi-pass transmission electron microscopy

DOE PAGES

Juffmann, Thomas; Koppell, Stewart A.; Klopfer, Brannon B.; ...

2017-05-10

Feynman once asked physicists to build better electron microscopes to be able to watch biology at work. While electron microscopes can now provide atomic resolution, electron beam induced specimen damage precludes high resolution imaging of sensitive materials, such as single proteins or polymers. Here, we use simulations to show that an electron microscope based on a multi-pass measurement protocol enables imaging of single proteins, without averaging structures over multiple images. While we demonstrate the method for particular imaging targets, the approach is broadly applicable and is expected to improve resolution and sensitivity for a range of electron microscopy imaging modalities,more » including, for example, scanning and spectroscopic techniques. The approach implements a quantum mechanically optimal strategy which under idealized conditions can be considered interaction-free.« less

Breaking resolution limits in ultrafast electron diffraction and microscopy

PubMed Central

Baum, Peter; Zewail, Ahmed H.

2006-01-01

Ultrafast electron microscopy and diffraction are powerful techniques for the study of the time-resolved structures of molecules, materials, and biological systems. Central to these approaches is the use of ultrafast coherent electron packets. The electron pulses typically have an energy of 30 keV for diffraction and 100–200 keV for microscopy, corresponding to speeds of 33–70% of the speed of light. Although the spatial resolution can reach the atomic scale, the temporal resolution is limited by the pulse width and by the difference in group velocities of electrons and the light used to initiate the dynamical change. In this contribution, we introduce the concept of tilted optical pulses into diffraction and imaging techniques and demonstrate the methodology experimentally. These advances allow us to reach limits of time resolution down to regimes of a few femtoseconds and, possibly, attoseconds. With tilted pulses, every part of the sample is excited at precisely the same time as when the electrons arrive at the specimen. Here, this approach is demonstrated for the most unfavorable case of ultrafast crystallography. We also present a method for measuring the duration of electron packets by autocorrelating electron pulses in free space and without streaking, and we discuss the potential of tilting the electron pulses themselves for applications in domains involving nuclear and electron motions. PMID:17056711

A simple energy filter for low energy electron microscopy/photoelectron emission microscopy instruments.

PubMed

Tromp, R M; Fujikawa, Y; Hannon, J B; Ellis, A W; Berghaus, A; Schaff, O

2009-08-05

Addition of an electron energy filter to low energy electron microscopy (LEEM) and photoelectron emission microscopy (PEEM) instruments greatly improves their analytical capabilities. However, such filters tend to be quite complex, both electron optically and mechanically. Here we describe a simple energy filter for the existing IBM LEEM/PEEM instrument, which is realized by adding a single scanning aperture slit to the objective transfer optics, without any further modifications to the microscope. This energy filter displays a very high energy resolution ΔE/E = 2 × 10(-5), and a non-isochromaticity of ∼0.5 eV/10 µm. The setup is capable of recording selected area electron energy spectra and angular distributions at 0.15 eV energy resolution, as well as energy filtered images with a 1.5 eV energy pass band at an estimated spatial resolution of ∼10 nm. We demonstrate the use of this energy filter in imaging and spectroscopy of surfaces using a laboratory-based He I (21.2 eV) light source, as well as imaging of Ag nanowires on Si(001) using the 4 eV energy loss Ag plasmon.

Diffraction and microscopy with attosecond electron pulse trains

NASA Astrophysics Data System (ADS)

Morimoto, Yuya; Baum, Peter

2018-03-01

Attosecond spectroscopy1-7 can resolve electronic processes directly in time, but a movie-like space-time recording is impeded by the too long wavelength ( 100 times larger than atomic distances) or the source-sample entanglement in re-collision techniques8-11. Here we advance attosecond metrology to picometre wavelength and sub-atomic resolution by using free-space electrons instead of higher-harmonic photons1-7 or re-colliding wavepackets8-11. A beam of 70-keV electrons at 4.5-pm de Broglie wavelength is modulated by the electric field of laser cycles into a sequence of electron pulses with sub-optical-cycle duration. Time-resolved diffraction from crystalline silicon reveals a < 10-as delay of Bragg emission and demonstrates the possibility of analytic attosecond-ångström diffraction. Real-space electron microscopy visualizes with sub-light-cycle resolution how an optical wave propagates in space and time. This unification of attosecond science with electron microscopy and diffraction enables space-time imaging of light-driven processes in the entire range of sample morphologies that electron microscopy can access.

Immunogold labeling reveals subcellular localisation of silica nanoparticles in a human blood-brain barrier model

NASA Astrophysics Data System (ADS)

Ye, Dong; Anguissola, Sergio; O'Neill, Tiina; Dawson, Kenneth A.

2015-05-01

Subcellular location of nanoparticles has been widely investigated with fluorescence microscopy, via fluorescently labeled antibodies to visualise target antigens in cells. However, fluorescence microscopy, such as confocal or live cell imaging, has generally limited 3D spatial resolution. Conventional electron microscopy can be useful in bridging resolution gap, but still not ideal in resolving subcellular organelle identities. Using the pre-embedding immunogold electron microscopic imaging, we performed accurate examination of the intracellular trafficking and gathered further evidence of transport mechanisms of silica nanoparticles across a human in vitro blood-brain barrier model. Our approach can effectively immunolocalise a variety of intracellular compartments and provide new insights into the uptake and subcellular transport of nanoparticles.Subcellular location of nanoparticles has been widely investigated with fluorescence microscopy, via fluorescently labeled antibodies to visualise target antigens in cells. However, fluorescence microscopy, such as confocal or live cell imaging, has generally limited 3D spatial resolution. Conventional electron microscopy can be useful in bridging resolution gap, but still not ideal in resolving subcellular organelle identities. Using the pre-embedding immunogold electron microscopic imaging, we performed accurate examination of the intracellular trafficking and gathered further evidence of transport mechanisms of silica nanoparticles across a human in vitro blood-brain barrier model. Our approach can effectively immunolocalise a variety of intracellular compartments and provide new insights into the uptake and subcellular transport of nanoparticles. Electronic supplementary information (ESI) available: Nanoparticle characterisation data, preservation of cellular structures, staining controls, optimisation of size amplification via the silver enhancement, and more imaging results from anti-clathrin and anti-caveolin 1 immunolabeling. See DOI: 10.1039/c5nr01539a

Reflections on the value of electron microscopy in the study of heterogeneous catalysts

PubMed Central

2017-01-01

Electron microscopy (EM) is arguably the single most powerful method of characterizing heterogeneous catalysts. Irrespective of whether they are bulk and multiphasic, or monophasic and monocrystalline, or nanocluster and even single-atom and on a support, their structures in atomic detail can be visualized in two or three dimensions, thanks to high-resolution instruments, with sub-Ångstrom spatial resolutions. Their topography, tomography, phase-purity, composition, as well as the bonding, and valence-states of their constituent atoms and ions and, in favourable circumstances, the short-range and long-range atomic order and dynamics of the catalytically active sites, can all be retrieved by the panoply of variants of modern EM. The latter embrace electron crystallography, rotation and precession electron diffraction, X-ray emission and high-resolution electron energy-loss spectra (EELS). Aberration-corrected (AC) transmission (TEM) and scanning transmission electron microscopy (STEM) have led to a revolution in structure determination. Environmental EM is already playing an increasing role in catalyst characterization, and new advances, involving special cells for the study of solid catalysts in contact with liquid reactants, have recently been deployed. PMID:28265196

Sub-nanometre resolution imaging of polymer-fullerene photovoltaic blends using energy-filtered scanning electron microscopy.

PubMed

Masters, Robert C; Pearson, Andrew J; Glen, Tom S; Sasam, Fabian-Cyril; Li, Letian; Dapor, Maurizio; Donald, Athene M; Lidzey, David G; Rodenburg, Cornelia

2015-04-24

The resolution capability of the scanning electron microscope has increased immensely in recent years, and is now within the sub-nanometre range, at least for inorganic materials. An equivalent advance has not yet been achieved for imaging the morphologies of nanostructured organic materials, such as organic photovoltaic blends. Here we show that energy-selective secondary electron detection can be used to obtain high-contrast, material-specific images of an organic photovoltaic blend. We also find that we can differentiate mixed phases from pure material phases in our data. The lateral resolution demonstrated is twice that previously reported from secondary electron imaging. Our results suggest that our energy-filtered scanning electron microscopy approach will be able to make major inroads into the understanding of complex, nano-structured organic materials.

Imaging interactions of metal oxide nanoparticles with macrophage cells by ultra-high resolution scanning electron microscopy techniques.

PubMed

Plascencia-Villa, Germán; Starr, Clarise R; Armstrong, Linda S; Ponce, Arturo; José-Yacamán, Miguel

2012-11-01

Use of engineered metal oxide nanoparticles in a plethora of biological applications and custom products has warned about some possible dose-dependent cytotoxic effects. Macrophages are key components of the innate immune system used to study possible toxic effects and internalization of different nanoparticulate materials. In this work, ultra-high resolution field emission scanning electron microscopy (FE-SEM) was used to offer new insights into the dynamical processes of interaction of nanomaterials with macrophage cells dosed with different concentrations of metal oxide nanoparticles (CeO(2), TiO(2) and ZnO). The versatility of FE-SEM has allowed obtaining a detailed characterization of processes of adsorption and endocytosis of nanoparticles, by using advanced analytical and imaging techniques on complete unstained uncoated cells, including secondary electron imaging, high-sensitive backscattered electron imaging, X-ray microanalysis and stereoimaging. Low voltage BF/DF-STEM confirmed nanoparticle adsorption and internalization into endosomes of CeO(2) and TiO(2), whereas ZnO develop apoptosis after 24 h of interaction caused by dissolution and invasion of cell nucleus. Ultra-high resolution scanning electron microscopy techniques provided new insights into interactions of inorganic nanoparticles with macrophage cells with high spatial resolution.

Filling the gap: adding super-resolution to array tomography for correlated ultrastructural and molecular identification of electrical synapses at the C. elegans connectome.

PubMed

Markert, Sebastian Matthias; Britz, Sebastian; Proppert, Sven; Lang, Marietta; Witvliet, Daniel; Mulcahy, Ben; Sauer, Markus; Zhen, Mei; Bessereau, Jean-Louis; Stigloher, Christian

2016-10-01

Correlating molecular labeling at the ultrastructural level with high confidence remains challenging. Array tomography (AT) allows for a combination of fluorescence and electron microscopy (EM) to visualize subcellular protein localization on serial EM sections. Here, we describe an application for AT that combines near-native tissue preservation via high-pressure freezing and freeze substitution with super-resolution light microscopy and high-resolution scanning electron microscopy (SEM) analysis on the same section. We established protocols that combine SEM with structured illumination microscopy (SIM) and direct stochastic optical reconstruction microscopy (dSTORM). We devised a method for easy, precise, and unbiased correlation of EM images and super-resolution imaging data using endogenous cellular landmarks and freely available image processing software. We demonstrate that these methods allow us to identify and label gap junctions in Caenorhabditis elegans with precision and confidence, and imaging of even smaller structures is feasible. With the emergence of connectomics, these methods will allow us to fill in the gap-acquiring the correlated ultrastructural and molecular identity of electrical synapses.

Correlative Fluorescence and Electron Microscopy in 3D-Scanning Electron Microscope Perspective.

PubMed

Franks, Jonathan; Wallace, Callen T; Shibata, Masateru; Suga, Mitsuo; Erdman, Natasha; Stolz, Donna B; Watkins, Simon C

2017-04-03

The ability to correlate fluorescence microscopy (FM) and electron microscopy (EM) data obtained on biological (cell and tissue) specimens is essential to bridge the resolution gap between the data obtained by these different imaging techniques. In the past such correlations were limited to either EM navigation in two dimensions to the locations previously highlighted by fluorescence markers, or subsequent high-resolution acquisition of tomographic information using a TEM. We present a novel approach whereby a sample previously investigated by FM is embedded and subjected to sequential mechanical polishing and backscatter imaging by scanning electron microscope. The resulting three dimensional EM tomogram of the sample can be directly correlated to the FM data. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

High-resolution Single Particle Analysis from Electron Cryo-microscopy Images Using SPHIRE

PubMed Central

Moriya, Toshio; Saur, Michael; Stabrin, Markus; Merino, Felipe; Voicu, Horatiu; Huang, Zhong; Penczek, Pawel A.; Raunser, Stefan; Gatsogiannis, Christos

2017-01-01

SPHIRE (SPARX for High-Resolution Electron Microscopy) is a novel open-source, user-friendly software suite for the semi-automated processing of single particle electron cryo-microscopy (cryo-EM) data. The protocol presented here describes in detail how to obtain a near-atomic resolution structure starting from cryo-EM micrograph movies by guiding users through all steps of the single particle structure determination pipeline. These steps are controlled from the new SPHIRE graphical user interface and require minimum user intervention. Using this protocol, a 3.5 Å structure of TcdA1, a Tc toxin complex from Photorhabdus luminescens, was derived from only 9500 single particles. This streamlined approach will help novice users without extensive processing experience and a priori structural information, to obtain noise-free and unbiased atomic models of their purified macromolecular complexes in their native state. PMID:28570515

Nanoscale deformation analysis with high-resolution transmission electron microscopy and digital image correlation

DOE PAGES

Wang, Xueju; Pan, Zhipeng; Fan, Feifei; ...

2015-09-10

We present an application of the digital image correlation (DIC) method to high-resolution transmission electron microscopy (HRTEM) images for nanoscale deformation analysis. The combination of DIC and HRTEM offers both the ultrahigh spatial resolution and high displacement detection sensitivity that are not possible with other microscope-based DIC techniques. We demonstrate the accuracy and utility of the HRTEM-DIC technique through displacement and strain analysis on amorphous silicon. Two types of error sources resulting from the transmission electron microscopy (TEM) image noise and electromagnetic-lens distortions are quantitatively investigated via rigid-body translation experiments. The local and global DIC approaches are applied for themore » analysis of diffusion- and reaction-induced deformation fields in electrochemically lithiated amorphous silicon. As a result, the DIC technique coupled with HRTEM provides a new avenue for the deformation analysis of materials at the nanometer length scales.« less

Nano-fEM: protein localization using photo-activated localization microscopy and electron microscopy.

PubMed

Watanabe, Shigeki; Richards, Jackson; Hollopeter, Gunther; Hobson, Robert J; Davis, Wayne M; Jorgensen, Erik M

2012-12-03

Mapping the distribution of proteins is essential for understanding the function of proteins in a cell. Fluorescence microscopy is extensively used for protein localization, but subcellular context is often absent in fluorescence images. Immuno-electron microscopy, on the other hand, can localize proteins, but the technique is limited by a lack of compatible antibodies, poor preservation of morphology and because most antigens are not exposed to the specimen surface. Correlative approaches can acquire the fluorescence image from a whole cell first, either from immuno-fluorescence or genetically tagged proteins. The sample is then fixed and embedded for electron microscopy, and the images are correlated (1-3). However, the low-resolution fluorescence image and the lack of fiducial markers preclude the precise localization of proteins. Alternatively, fluorescence imaging can be done after preserving the specimen in plastic. In this approach, the block is sectioned, and fluorescence images and electron micrographs of the same section are correlated (4-7). However, the diffraction limit of light in the correlated image obscures the locations of individual molecules, and the fluorescence often extends beyond the boundary of the cell. Nano-resolution fluorescence electron microscopy (nano-fEM) is designed to localize proteins at nano-scale by imaging the same sections using photo-activated localization microscopy (PALM) and electron microscopy. PALM overcomes the diffraction limit by imaging individual fluorescent proteins and subsequently mapping the centroid of each fluorescent spot (8-10). We outline the nano-fEM technique in five steps. First, the sample is fixed and embedded using conditions that preserve the fluorescence of tagged proteins. Second, the resin blocks are sectioned into ultrathin segments (70-80 nm) that are mounted on a cover glass. Third, fluorescence is imaged in these sections using the Zeiss PALM microscope. Fourth, electron dense structures are imaged in these same sections using a scanning electron microscope. Fifth, the fluorescence and electron micrographs are aligned using gold particles as fiducial markers. In summary, the subcellular localization of fluorescently tagged proteins can be determined at nanometer resolution in approximately one week.

High spatial resolution detection of low-energy electrons using an event-counting method, application to point projection microscopy

NASA Astrophysics Data System (ADS)

Salançon, Evelyne; Degiovanni, Alain; Lapena, Laurent; Morin, Roger

2018-04-01

An event-counting method using a two-microchannel plate stack in a low-energy electron point projection microscope is implemented. 15 μm detector spatial resolution, i.e., the distance between first-neighbor microchannels, is demonstrated. This leads to a 7 times better microscope resolution. Compared to previous work with neutrons [Tremsin et al., Nucl. Instrum. Methods Phys. Res., Sect. A 592, 374 (2008)], the large number of detection events achieved with electrons shows that the local response of the detector is mainly governed by the angle between the hexagonal structures of the two microchannel plates. Using this method in point projection microscopy offers the prospect of working with a greater source-object distance (350 nm instead of 50 nm), advancing toward atomic resolution.

Four-dimensional ultrafast electron microscopy of phase transitions

PubMed Central

Grinolds, Michael S.; Lobastov, Vladimir A.; Weissenrieder, Jonas; Zewail, Ahmed H.

2006-01-01

Reported here is direct imaging (and diffraction) by using 4D ultrafast electron microscopy (UEM) with combined spatial and temporal resolutions. In the first phase of UEM, it was possible to obtain snapshot images by using timed, single-electron packets; each packet is free of space–charge effects. Here, we demonstrate the ability to obtain sequences of snapshots (“movies”) with atomic-scale spatial resolution and ultrashort temporal resolution. Specifically, it is shown that ultrafast metal–insulator phase transitions can be studied with these achieved spatial and temporal resolutions. The diffraction (atomic scale) and images (nanometer scale) we obtained manifest the structural phase transition with its characteristic hysteresis, and the time scale involved (100 fs) is now studied by directly monitoring coordinates of the atoms themselves. PMID:17130445

Sub-nanometre resolution imaging of polymer–fullerene photovoltaic blends using energy-filtered scanning electron microscopy

PubMed Central

Masters, Robert C.; Pearson, Andrew J.; Glen, Tom S.; Sasam, Fabian-Cyril; Li, Letian; Dapor, Maurizio; Donald, Athene M.; Lidzey, David G.; Rodenburg, Cornelia

2015-01-01

The resolution capability of the scanning electron microscope has increased immensely in recent years, and is now within the sub-nanometre range, at least for inorganic materials. An equivalent advance has not yet been achieved for imaging the morphologies of nanostructured organic materials, such as organic photovoltaic blends. Here we show that energy-selective secondary electron detection can be used to obtain high-contrast, material-specific images of an organic photovoltaic blend. We also find that we can differentiate mixed phases from pure material phases in our data. The lateral resolution demonstrated is twice that previously reported from secondary electron imaging. Our results suggest that our energy-filtered scanning electron microscopy approach will be able to make major inroads into the understanding of complex, nano-structured organic materials. PMID:25906738

Three-Dimensional Intercalated Porous Graphene on Si(111)

NASA Astrophysics Data System (ADS)

Pham, Trung T.; Sporken, Robert

2018-02-01

Three-dimensional intercalated porous graphene has been formed on Si(111) by electron beam evaporation under appropriate conditions and its structural and electronic properties investigated in detail by reflection high-energy electron diffraction, x-ray photoemission spectroscopy, Raman spectroscopy, high-resolution scanning electron microscopy, atomic force microscopy, and scanning tunneling microscopy. The results show that the crystalline quality of the porous graphene depended not only on the substrate temperature but also on the SiC layer thickness during carbon atom deposition.

Correlative Super-Resolution Microscopy: New Dimensions and New Opportunities.

PubMed

Hauser, Meghan; Wojcik, Michal; Kim, Doory; Mahmoudi, Morteza; Li, Wan; Xu, Ke

2017-06-14

Correlative microscopy, the integration of two or more microscopy techniques performed on the same sample, produces results that emphasize the strengths of each technique while offsetting their individual weaknesses. Light microscopy has historically been a central method in correlative microscopy due to its widespread availability, compatibility with hydrated and live biological samples, and excellent molecular specificity through fluorescence labeling. However, conventional light microscopy can only achieve a resolution of ∼300 nm, undercutting its advantages in correlations with higher-resolution methods. The rise of super-resolution microscopy (SRM) over the past decade has drastically improved the resolution of light microscopy to ∼10 nm, thus creating exciting new opportunities and challenges for correlative microscopy. Here we review how these challenges are addressed to effectively correlate SRM with other microscopy techniques, including light microscopy, electron microscopy, cryomicroscopy, atomic force microscopy, and various forms of spectroscopy. Though we emphasize biological studies, we also discuss the application of correlative SRM to materials characterization and single-molecule reactions. Finally, we point out current limitations and discuss possible future improvements and advances. We thus demonstrate how a correlative approach adds new dimensions of information and provides new opportunities in the fast-growing field of SRM.

Scanning ultrafast electron microscopy.

PubMed

Yang, Ding-Shyue; Mohammed, Omar F; Zewail, Ahmed H

2010-08-24

Progress has been made in the development of four-dimensional ultrafast electron microscopy, which enables space-time imaging of structural dynamics in the condensed phase. In ultrafast electron microscopy, the electrons are accelerated, typically to 200 keV, and the microscope operates in the transmission mode. Here, we report the development of scanning ultrafast electron microscopy using a field-emission-source configuration. Scanning of pulses is made in the single-electron mode, for which the pulse contains at most one or a few electrons, thus achieving imaging without the space-charge effect between electrons, and still in ten(s) of seconds. For imaging, the secondary electrons from surface structures are detected, as demonstrated here for material surfaces and biological specimens. By recording backscattered electrons, diffraction patterns from single crystals were also obtained. Scanning pulsed-electron microscopy with the acquired spatiotemporal resolutions, and its efficient heat-dissipation feature, is now poised to provide in situ 4D imaging and with environmental capability.

An historical account of the development and applications of the negative staining technique to the electron microscopy of viruses.

PubMed

Horne, R W; Wildy, P

1979-09-01

A brief historical account of the development and applications of the negative staining techniques to the study of the structure of viruses and their components as observed in the electron microscope is presented. Although the basic method of surrounding or embedding specimens in opaque dyes was used in light microscopy dating from about 1884, the equivalent preparative techniques applied to electron microscopy were comparatively recent. The combination of experiments on a sophisticated bacterial virus and the installation of a high resolution electron microscope in the Cavendish Laboratory, Cambridge, during 1954, subsequently led to the analysis of several important morphological features of animal, plant and bacterial viruses. The implications of the results from these early experiments on viruses and recent developments in negative staining methods for high resolution image analysis of electron micrographs are also discussed.

A graphene oxide-carbon nanotube grid for high-resolution transmission electron microscopy of nanomaterials.

PubMed

Zhang, Lina; Zhang, Haoxu; Zhou, Ruifeng; Chen, Zhuo; Li, Qunqing; Fan, Shoushan; Ge, Guanglu; Liu, Renxiao; Jiang, Kaili

2011-09-23

A novel grid for use in transmission electron microscopy is developed. The supporting film of the grid is composed of thin graphene oxide films overlying a super-aligned carbon nanotube network. The composite film combines the advantages of graphene oxide and carbon nanotube networks and has the following properties: it is ultra-thin, it has a large flat and smooth effective supporting area with a homogeneous amorphous appearance, high stability, and good conductivity. The graphene oxide-carbon nanotube grid has a distinct advantage when characterizing the fine structure of a mass of nanomaterials over conventional amorphous carbon grids. Clear high-resolution transmission electron microscopy images of various nanomaterials are obtained easily using the new grids.

A fast image simulation algorithm for scanning transmission electron microscopy.

PubMed

Ophus, Colin

2017-01-01

Image simulation for scanning transmission electron microscopy at atomic resolution for samples with realistic dimensions can require very large computation times using existing simulation algorithms. We present a new algorithm named PRISM that combines features of the two most commonly used algorithms, namely the Bloch wave and multislice methods. PRISM uses a Fourier interpolation factor f that has typical values of 4-20 for atomic resolution simulations. We show that in many cases PRISM can provide a speedup that scales with f 4 compared to multislice simulations, with a negligible loss of accuracy. We demonstrate the usefulness of this method with large-scale scanning transmission electron microscopy image simulations of a crystalline nanoparticle on an amorphous carbon substrate.

A fast image simulation algorithm for scanning transmission electron microscopy

DOE PAGES

Ophus, Colin

2017-05-10

Image simulation for scanning transmission electron microscopy at atomic resolution for samples with realistic dimensions can require very large computation times using existing simulation algorithms. Here, we present a new algorithm named PRISM that combines features of the two most commonly used algorithms, namely the Bloch wave and multislice methods. PRISM uses a Fourier interpolation factor f that has typical values of 4-20 for atomic resolution simulations. We show that in many cases PRISM can provide a speedup that scales with f 4 compared to multislice simulations, with a negligible loss of accuracy. We demonstrate the usefulness of this methodmore » with large-scale scanning transmission electron microscopy image simulations of a crystalline nanoparticle on an amorphous carbon substrate.« less

Atomic resolution elemental mapping using energy-filtered imaging scanning transmission electron microscopy with chromatic aberration correction.

PubMed

Krause, F F; Rosenauer, A; Barthel, J; Mayer, J; Urban, K; Dunin-Borkowski, R E; Brown, H G; Forbes, B D; Allen, L J

2017-10-01

This paper addresses a novel approach to atomic resolution elemental mapping, demonstrating a method that produces elemental maps with a similar resolution to the established method of electron energy-loss spectroscopy in scanning transmission electron microscopy. Dubbed energy-filtered imaging scanning transmission electron microscopy (EFISTEM) this mode of imaging is, by the quantum mechanical principle of reciprocity, equivalent to tilting the probe in energy-filtered transmission electron microscopy (EFTEM) through a cone and incoherently averaging the results. In this paper we present a proof-of-principle EFISTEM experimental study on strontium titanate. The present approach, made possible by chromatic aberration correction, has the advantage that it provides elemental maps which are immune to spatial incoherence in the electron source, coherent aberrations in the probe-forming lens and probe jitter. The veracity of the experiment is supported by quantum mechanical image simulations, which provide an insight into the image-forming process. Elemental maps obtained in EFTEM suffer from the effect known as preservation of elastic contrast, which, for example, can lead to a given atomic species appearing to be in atomic columns where it is not to be found. EFISTEM very substantially reduces the preservation of elastic contrast and yields images which show stability of contrast with changing thickness. The experimental application is demonstrated in a proof-of-principle study on strontium titanate. Copyright © 2017 Elsevier B.V. All rights reserved.

Navigating 3D electron microscopy maps with EM-SURFER.

PubMed

Esquivel-Rodríguez, Juan; Xiong, Yi; Han, Xusi; Guang, Shuomeng; Christoffer, Charles; Kihara, Daisuke

2015-05-30

The Electron Microscopy DataBank (EMDB) is growing rapidly, accumulating biological structural data obtained mainly by electron microscopy and tomography, which are emerging techniques for determining large biomolecular complex and subcellular structures. Together with the Protein Data Bank (PDB), EMDB is becoming a fundamental resource of the tertiary structures of biological macromolecules. To take full advantage of this indispensable resource, the ability to search the database by structural similarity is essential. However, unlike high-resolution structures stored in PDB, methods for comparing low-resolution electron microscopy (EM) density maps in EMDB are not well established. We developed a computational method for efficiently searching low-resolution EM maps. The method uses a compact fingerprint representation of EM maps based on the 3D Zernike descriptor, which is derived from a mathematical series expansion for EM maps that are considered as 3D functions. The method is implemented in a web server named EM-SURFER, which allows users to search against the entire EMDB in real-time. EM-SURFER compares the global shapes of EM maps. Examples of search results from different types of query structures are discussed. We developed EM-SURFER, which retrieves structurally relevant matches for query EM maps from EMDB within seconds. The unique capability of EM-SURFER to detect 3D shape similarity of low-resolution EM maps should prove invaluable in structural biology.

Water without windows: Evaluating the performance of open cell transmission electron microscopy under saturated water vapor conditions, and assessing its potential for microscopy of hydrated biological specimens.

PubMed

Cassidy, Cathal; Yamashita, Masao; Cheung, Martin; Kalale, Chola; Adaniya, Hidehito; Kuwahara, Ryusuke; Shintake, Tsumoru

2017-01-01

We have performed open cell transmission electron microscopy experiments through pure water vapor in the saturation pressure regime (>0.6 kPa), in a modern microscope capable of sub-Å resolution. We have systematically studied achievable pressure levels, stability and gas purity, effective thickness of the water vapor column and associated electron scattering processes, and the effect of gas pressure on electron optical resolution and image contrast. For example, for 1.3 kPa pure water vapor and 300kV electrons, we report pressure stability of ± 20 Pa over tens of minutes, effective thickness of 0.57 inelastic mean free paths, lattice resolution of 0.14 nm on a reference Au specimen, and no significant degradation in contrast or stability of a biological specimen (M13 virus, with 6 nm body diameter). We have also done some brief experiments to confirm feasibility of loading specimens into an in situ water vapor ambient without exposure to intermediate desiccating conditions. Finally, we have also checked if water experiments had any discernible impact on the microscope performance, and report pertinent vacuum and electron optical data, for reference purposes.

Direct observation and analysis of york-shell materials using low-voltage high-resolution scanning electron microscopy: Nanometal-particles encapsulated in metal-oxide, carbon, and polymer

NASA Astrophysics Data System (ADS)

Asahina, Shunsuke; Suga, Mitsuo; Takahashi, Hideyuki; Young Jeong, Hu; Galeano, Carolina; Schüth, Ferdi; Terasaki, Osamu

2014-11-01

Nanometal particles show characteristic features in chemical and physical properties depending on their sizes and shapes. For keeping and further enhancing their features, the particles should be protected from coalescence or degradation. One approach is to encapsulate the nanometal particles inside pores with chemically inert or functional materials, such as carbon, polymer, and metal oxides, which contain mesopores to allow permeation of only chemicals not the nanometal particles. Recently developed low-voltage high-resolution scanning electron microscopy was applied to the study of structural, chemical, and electron state of both nanometal particles and encapsulating materials in yolk-shell materials of Au@C, Ru/Pt@C, Au@TiO2, and Pt@Polymer. Progresses in the following categories were shown for the yolk-shell materials: (i) resolution of topographic image contrast by secondary electrons, of atomic-number contrast by back-scattered electrons, and of elemental mapping by X-ray energy dispersive spectroscopy; (ii) sample preparation for observing internal structures; and (iii) X-ray spectroscopy such as soft X-ray emission spectroscopy. Transmission electron microscopy was also used for characterization of Au@C.

Helium ion microscopy and ultra-high-resolution scanning electron microscopy analysis of membrane-extracted cells reveals novel characteristics of the cytoskeleton of Giardia intestinalis.

PubMed

Gadelha, Ana Paula Rocha; Benchimol, Marlene; de Souza, Wanderley

2015-06-01

Giardia intestinalis presents a complex microtubular cytoskeleton formed by specialized structures, such as the adhesive disk, four pairs of flagella, the funis and the median body. The ultrastructural organization of the Giardia cytoskeleton has been analyzed using different microscopic techniques, including high-resolution scanning electron microscopy. Recent advances in scanning microscopy technology have opened a new venue for the characterization of cellular structures and include scanning probe microscopy techniques such as ultra-high-resolution scanning electron microscopy (UHRSEM) and helium ion microscopy (HIM). Here, we studied the organization of the cytoskeleton of G. intestinalis trophozoites using UHRSEM and HIM in membrane-extracted cells. The results revealed a number of new cytoskeletal elements associated with the lateral crest and the dorsal surface of the parasite. The fine structure of the banded collar was also observed. The marginal plates were seen linked to a network of filaments, which were continuous with filaments parallel to the main cell axis. Cytoplasmic filaments that supported the internal structures were seen by the first time. Using anti-actin antibody, we observed a labeling in these filamentous structures. Taken together, these data revealed new surface characteristics of the cytoskeleton of G. intestinalis and may contribute to an improved understanding of the structural organization of trophozoites. Copyright © 2015 Elsevier Inc. All rights reserved.

Imaging interactions of metal oxide nanoparticles with macrophage cells by ultra-high resolution scanning electron microscopy techniques†

PubMed Central

Plascencia-Villa, Germán; Starr, Clarise R.; Armstrong, Linda S.; Ponce, Arturo

2016-01-01

Use of engineered metal oxide nanoparticles in a plethora of biological applications and custom products has warned about some possible dose-dependent cytotoxic effects. Macrophages are key components of the innate immune system used to study possible toxic effects and internalization of different nanoparticulate materials. In this work, ultra-high resolution field emission scanning electron microscopy (FE-SEM) was used to offer new insights into the dynamical processes of interaction of nanomaterials with macrophage cells dosed with different concentrations of metal oxide nanoparticles (CeO2, TiO2 and ZnO). The versatility of FE-SEM has allowed obtaining a detailed characterization of processes of adsorption and endocytosis of nanoparticles, by using advanced analytical and imaging techniques on complete unstained uncoated cells, including secondary electron imaging, high-sensitive backscattered electron imaging, X-ray microanalysis and stereoimaging. Low voltage BF/DF-STEM confirmed nanoparticle adsorption and internalization into endosomes of CeO2 and TiO2, whereas ZnO develop apoptosis after 24 h of interaction caused by dissolution and invasion of cell nucleus. Ultra-high resolution scanning electron microscopy techniques provided new insights into interactions of inorganic nanoparticles with macrophage cells with high spatial resolution. PMID:23023106

Transmission/Scanning Transmission Electron Microscopy | Materials Science

Science.gov Websites

imaging such as high resolution TEM. Transmission electron diffraction patterns help to determine the microstructure of a material and its defects. Phase-contrast imaging or high-resolution (HR) TEM imaging gives high scattering angle can be collected to form high-resolution, chemically sensitive, atomic number (Z

Quantum dot immunocytochemical localization of somatostatin in somatostatinoma by Widefield Epifluorescence, super-resolution light, and immunoelectron microscopy.

PubMed

Killingsworth, Murray C; Lai, Ken; Wu, Xiaojuan; Yong, Jim L C; Lee, C Soon

2012-11-01

Quantum dot nanocrystal probes (QDs) have been used for detection of somatostatin hormone in secretory granules of somatostatinoma tumor cells by immunofluorescence light microscopy, super-resolution light microscopy, and immunoelectron microscopy. Immunostaining for all modalities was done using sections taken from an epoxy resin-embedded tissue specimen and a similar labeling protocol. This approach allowed assessment of labeling at light microscopy level before examination at super-resolution and electron microscopy level and was a significant aid in interpretation. Etching of ultrathin sections with saturated sodium metaperiodate was a critical step presumably able to retrieve some tissue antigenicity masked by processing in epoxy resin. Immunofluorescence microscopy of QD-immunolabeled sections showed somatostatin hormone localization in cytoplasmic granules. Some variable staining of tumor gland-like structures appeared related to granule maturity and dispersal of granule contents within the tumor cell cytoplasm. Super-resolution light microscopy demonstrated localization of somatostatin within individual secretory granules to be heterogeneous, and this staining pattern was confirmed by immunoelectron microscopy.

Quantum Dot Immunocytochemical Localization of Somatostatin in Somatostatinoma by Widefield Epifluorescence, Super-resolution Light, and Immunoelectron Microscopy

PubMed Central

Lai, Ken; Wu, Xiaojuan; Yong, Jim L. C.; Lee, C. Soon

2012-01-01

Quantum dot nanocrystal probes (QDs) have been used for detection of somatostatin hormone in secretory granules of somatostatinoma tumor cells by immunofluorescence light microscopy, super-resolution light microscopy, and immunoelectron microscopy. Immunostaining for all modalities was done using sections taken from an epoxy resin-embedded tissue specimen and a similar labeling protocol. This approach allowed assessment of labeling at light microscopy level before examination at super-resolution and electron microscopy level and was a significant aid in interpretation. Etching of ultrathin sections with saturated sodium metaperiodate was a critical step presumably able to retrieve some tissue antigenicity masked by processing in epoxy resin. Immunofluorescence microscopy of QD-immunolabeled sections showed somatostatin hormone localization in cytoplasmic granules. Some variable staining of tumor gland-like structures appeared related to granule maturity and dispersal of granule contents within the tumor cell cytoplasm. Super-resolution light microscopy demonstrated localization of somatostatin within individual secretory granules to be heterogeneous, and this staining pattern was confirmed by immunoelectron microscopy. PMID:22899862

Evaluation of a hybrid pixel detector for electron microscopy.

PubMed

Faruqi, A R; Cattermole, D M; Henderson, R; Mikulec, B; Raeburn, C

2003-04-01

We describe the application of a silicon hybrid pixel detector, containing 64 by 64 pixels, each 170 microm(2), in electron microscopy. The device offers improved resolution compared to CCDs along with faster and noiseless readout. Evaluation of the detector, carried out on a 120 kV electron microscope, demonstrates the potential of the device.

EMRinger: side chain–directed model and map validation for 3D cryo-electron microscopy

DOE PAGES

Barad, Benjamin A.; Echols, Nathaniel; Wang, Ray Yu-Ruei; ...

2015-08-17

Advances in high-resolution cryo-electron microscopy (cryo-EM) require the development of validation metrics to independently assess map quality and model geometry. We report that EMRinger is a tool that assesses the precise fitting of an atomic model into the map during refinement and shows how radiation damage alters scattering from negatively charged amino acids. EMRinger (https://github.com/fraser-lab/EMRinger) will be useful for monitoring progress in resolving and modeling high-resolution features in cryo-EM.

The significance of Bragg's law in electron diffraction and microscopy, and Bragg's second law.

PubMed

Humphreys, C J

2013-01-01

Bragg's second law, which deserves to be more widely known, is recounted. The significance of Bragg's law in electron diffraction and microscopy is then discussed, with particular emphasis on differences between X-ray and electron diffraction. As an example of such differences, the critical voltage effect in electron diffraction is described. It is then shown that the lattice imaging of crystals in high-resolution electron microscopy directly reveals the Bragg planes used for the imaging process, exactly as visualized by Bragg in his real-space law. Finally, it is shown how in 2012, for the first time, on the centennial anniversary of Bragg's law, single atoms have been identified in an electron microscope using X-rays emitted from the specimen. Hence atomic resolution X-ray maps of a crystal in real space can be formed which give the positions and identities of the different atoms in the crystal, or of a single impurity atom in the crystal.

Techniques for super-resolution microscopy using NV-diamond

NASA Astrophysics Data System (ADS)

Trifonov, Alexei; Glenn, David; Bar-Gill, Nir; Le Sage, David; Walsworth, Ronald

2011-05-01

We discuss the development and application of techniques for super-resolution microscopy using NV centers in diamond: stimulated emission depletion (STED), metastable ground state depletion (GSD), and stochastic optical reconstruction microscopy (STORM). NV centers do not bleach under optical excitation, are not biotoxic, and have long-lived electronic spin coherence and spin-state-dependent fluorescence. Thus NV-diamond has great potential as a fluorescent biomarker and as a magnetic biosensor.

Fabrication of ZnS nanoparticle chains on a protein template

PubMed Central

Hulleman, J.; Kim, S. M.; Tumkur, T.; Rochet, J.-C.; Stach, E.; Stanciu, L.

2011-01-01

In the present study, we have exploited the properties of a fibrillar protein for the template synthesis of zinc sulfide (ZnS) nanoparticle chains. The diameter of the ZnS nanoparticle chains was tuned in range of ~30 to ~165 nm by varying the process variables. The nanoparticle chains were characterized by field emission scanning electron microscopy, UV–Visible spectroscopy, transmission electron microscopy, electron energy loss spectroscopy, and high-resolution transmission electron microscopy. The effect of incubation temperature on the morphology of the nanoparticle chains was also studied. PMID:21804765

Imaging plasmodesmata with high-resolution scanning electron microscopy.

PubMed

Barton, Deborah A; Overall, Robyn L

2015-01-01

High-resolution scanning electron microscopy (HRSEM) is an effective tool to investigate the distribution of plasmodesmata within plant cell walls as well as to probe their complex, three-dimensional architecture. It is a useful alternative to traditional transmission electron microscopy (TEM) in which plasmodesmata are sectioned to reveal their internal substructures. Benefits of adopting an HRSEM approach to studies of plasmodesmata are that the specimen preparation methods are less complex and time consuming than for TEM, many plasmodesmata within a large region of tissue can be imaged in a single session, and three-dimensional information is readily available without the need for reconstructing TEM serial sections or employing transmission electron tomography, both of which are lengthy processes. Here we describe methods to prepare plant samples for HRSEM using pre- or postfixation extraction of cellular material in order to visualize plasmodesmata embedded within plant cell walls.

4D imaging of transient structures and morphologies in ultrafast electron microscopy.

PubMed

Barwick, Brett; Park, Hyun Soon; Kwon, Oh-Hoon; Baskin, J Spencer; Zewail, Ahmed H

2008-11-21

With advances in spatial resolution reaching the atomic scale, two-dimensional (2D) and 3D imaging in electron microscopy has become an essential methodology in various fields of study. Here, we report 4D imaging, with in situ spatiotemporal resolutions, in ultrafast electron microscopy (UEM). The ability to capture selected-area-image dynamics with pixel resolution and to control the time separation between pulses for temporal cooling of the specimen made possible studies of fleeting structures and morphologies. We demonstrate the potential for applications with two examples, gold and graphite. For gold, after thermally induced stress, we determined the atomic structural expansion, the nonthermal lattice temperature, and the ultrafast transients of warping/bulging. In contrast, in graphite, striking coherent transients of the structure were observed in both image and diffraction, directly measuring, on the nanoscale, the longitudinal resonance period governed by Young's elastic modulus. The success of these studies demonstrates the promise of UEM in real-space imaging of dynamics.

High yield production of long branched Au nanoparticles characterized by atomic resolution transmission electron microscopy

PubMed Central

Mayoral, Alvaro; Magen, Cesar; Jose-Yacaman, Miguel

2011-01-01

Long multi-branched gold nanoparticles have been synthesized in a very high yield through a facile synthesis combining two different capping agents. The stability of these materials with the time has been tested and their characterization have been performed by diverse advanced electron microscopy techniques, paying special attention to aberration corrected transmission electron microscopy in order to unambiguously analyze the surface structure of the branches and provide insights for the formation of stellated gold nanoparticles. PMID:22125420

Tackling the Challenges of Dynamic Experiments Using Liquid-Cell Transmission Electron Microscopy.

PubMed

Parent, Lucas R; Bakalis, Evangelos; Proetto, Maria; Li, Yiwen; Park, Chiwoo; Zerbetto, Francesco; Gianneschi, Nathan C

2018-01-16

Revolutions in science and engineering frequently result from the development, and wide adoption, of a new, powerful characterization or imaging technique. Beginning with the first glass lenses and telescopes in astronomy, to the development of visual-light microscopy, staining techniques, confocal microscopy, and fluorescence super-resolution microscopy in biology, and most recently aberration-corrected, cryogenic, and ultrafast (4D) electron microscopy, X-ray microscopy, and scanning probe microscopy in nanoscience. Through these developments, our perception and understanding of the physical nature of matter at length-scales beyond ordinary perception have been fundamentally transformed. Despite this progression in microscopy, techniques for observing nanoscale chemical processes and solvated/hydrated systems are limited, as the necessary spatial and temporal resolution presents significant technical challenges. However, the standard reliance on indirect or bulk phase characterization of nanoscale samples in liquids is undergoing a shift in recent times with the realization ( Williamson et al. Nat. Mater . 2003 , 2 , 532 - 536 ) of liquid-cell (scanning) transmission electron microscopy, LC(S)TEM, where picoliters of solution are hermetically sealed between electron-transparent "windows," which can be directly imaged or videoed at the nanoscale using conventional transmission electron microscopes. This Account seeks to open a discussion on the topic of standardizing strategies for conducting imaging experiments with a view to characterizing dynamics and motion of nanoscale materials. This is a challenge that could be described by critics and proponents alike, as analogous to doing chemistry in a lightning storm; where the nature of the solution, the nanomaterial, and the dynamic behaviors are all potentially subject to artifactual influence by the very act of our observation.

Hollow Cone Electron Imaging for Single Particle 3D Reconstruction of Proteins

PubMed Central

Tsai, Chun-Ying; Chang, Yuan-Chih; Lobato, Ivan; Van Dyck, Dirk; Chen, Fu-Rong

2016-01-01

The main bottlenecks for high-resolution biological imaging in electron microscopy are radiation sensitivity and low contrast. The phase contrast at low spatial frequencies can be enhanced by using a large defocus but this strongly reduces the resolution. Recently, phase plates have been developed to enhance the contrast at small defocus but electrical charging remains a problem. Single particle cryo-electron microscopy is mostly used to minimize the radiation damage and to enhance the resolution of the 3D reconstructions but it requires averaging images of a massive number of individual particles. Here we present a new route to achieve the same goals by hollow cone dark field imaging using thermal diffuse scattered electrons giving about a 4 times contrast increase as compared to bright field imaging. We demonstrate the 3D reconstruction of a stained GroEL particle can yield about 13.5 Å resolution but using a strongly reduced number of images. PMID:27292544

Scanning ultrafast electron microscopy

PubMed Central

Yang, Ding-Shyue; Mohammed, Omar F.; Zewail, Ahmed H.

2010-01-01

Progress has been made in the development of four-dimensional ultrafast electron microscopy, which enables space-time imaging of structural dynamics in the condensed phase. In ultrafast electron microscopy, the electrons are accelerated, typically to 200 keV, and the microscope operates in the transmission mode. Here, we report the development of scanning ultrafast electron microscopy using a field-emission-source configuration. Scanning of pulses is made in the single-electron mode, for which the pulse contains at most one or a few electrons, thus achieving imaging without the space-charge effect between electrons, and still in ten(s) of seconds. For imaging, the secondary electrons from surface structures are detected, as demonstrated here for material surfaces and biological specimens. By recording backscattered electrons, diffraction patterns from single crystals were also obtained. Scanning pulsed-electron microscopy with the acquired spatiotemporal resolutions, and its efficient heat-dissipation feature, is now poised to provide in situ 4D imaging and with environmental capability. PMID:20696933

High-resolution x-ray diffraction microscopy of specifically labeled yeast cells

PubMed Central

Nelson, Johanna; Huang, Xiaojing; Steinbrener, Jan; Shapiro, David; Kirz, Janos; Marchesini, Stefano; Neiman, Aaron M.; Turner, Joshua J.; Jacobsen, Chris

2010-01-01

X-ray diffraction microscopy complements other x-ray microscopy methods by being free of lens-imposed radiation dose and resolution limits, and it allows for high-resolution imaging of biological specimens too thick to be viewed by electron microscopy. We report here the highest resolution (11–13 nm) x-ray diffraction micrograph of biological specimens, and a demonstration of molecular-specific gold labeling at different depths within cells via through-focus propagation of the reconstructed wavefield. The lectin concanavalin A conjugated to colloidal gold particles was used to label the α-mannan sugar in the cell wall of the yeast Saccharomyces cerevisiae. Cells were plunge-frozen in liquid ethane and freeze-dried, after which they were imaged whole using x-ray diffraction microscopy at 750 eV photon energy. PMID:20368463

High-resolution x-ray diffraction microscopy of specifically labeled yeast cells

DOE PAGES

Nelson, Johanna; Huang, Xiaojing; Steinbrener, Jan; ...

2010-04-20

X-ray diffraction microscopy complements other x-ray microscopy methods by being free of lens-imposed radiation dose and resolution limits, and it allows for high-resolution imaging of biological specimens too thick to be viewed by electron microscopy. We report here the highest resolution (11-13 nm) x-ray diffraction micrograph of biological specimens, and a demonstration of molecular-specific gold labeling at different depths within cells via through-focus propagation of the reconstructed wavefield. The lectin concanavalin A conjugated to colloidal gold particles was used to label the α-mannan sugar in the cell wall of the yeast Saccharomyces cerevisiae. Cells were plunge-frozen in liquid ethane andmore » freeze-dried, after which they were imaged whole using x-ray diffraction microscopy at 750 eV photon energy.« less

A direct electron detector for time-resolved MeV electron microscopy

DOE PAGES

Vecchione, T.; Denes, P.; Jobe, R. K.; ...

2017-03-15

The introduction of direct electron detectors enabled the structural biology revolution of cryogenic electron microscopy. Direct electron detectors are now expected to have a similarly dramatic impact on time-resolved MeV electron microscopy, particularly by enabling both spatial and temporal jitter correction. Here in this paper, we report on the commissioning of a direct electron detector for time-resolved MeV electron microscopy. The direct electron detector demonstrated MeV single electron sensitivity and is capable of recording megapixel images at 180 Hz. The detector has a 15-bit dynamic range, better than 30-μm spatial resolution and less than 20 analogue-to-digital converter count RMS pixelmore » noise. The unique capabilities of the direct electron detector and the data analysis required to take advantage of these capabilities are presented. The technical challenges associated with generating and processing large amounts of data are also discussed.« less

A direct electron detector for time-resolved MeV electron microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vecchione, T.; Denes, P.; Jobe, R. K.

The introduction of direct electron detectors enabled the structural biology revolution of cryogenic electron microscopy. Direct electron detectors are now expected to have a similarly dramatic impact on time-resolved MeV electron microscopy, particularly by enabling both spatial and temporal jitter correction. Here we report on the commissioning of a direct electron detector for time-resolved MeV electron microscopy. The direct electron detector demonstrated MeV single electron sensitivity and is capable of recording megapixel images at 180 Hz. The detector has a 15-bit dynamic range, better than 30-μmμm spatial resolution and less than 20 analogue-to-digital converter count RMS pixel noise. The uniquemore » capabilities of the direct electron detector and the data analysis required to take advantage of these capabilities are presented. The technical challenges associated with generating and processing large amounts of data are also discussed.« less

A direct electron detector for time-resolved MeV electron microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vecchione, T.; Denes, P.; Jobe, R. K.

The introduction of direct electron detectors enabled the structural biology revolution of cryogenic electron microscopy. Direct electron detectors are now expected to have a similarly dramatic impact on time-resolved MeV electron microscopy, particularly by enabling both spatial and temporal jitter correction. Here in this paper, we report on the commissioning of a direct electron detector for time-resolved MeV electron microscopy. The direct electron detector demonstrated MeV single electron sensitivity and is capable of recording megapixel images at 180 Hz. The detector has a 15-bit dynamic range, better than 30-μm spatial resolution and less than 20 analogue-to-digital converter count RMS pixelmore » noise. The unique capabilities of the direct electron detector and the data analysis required to take advantage of these capabilities are presented. The technical challenges associated with generating and processing large amounts of data are also discussed.« less

Water without windows: Evaluating the performance of open cell transmission electron microscopy under saturated water vapor conditions, and assessing its potential for microscopy of hydrated biological specimens

PubMed Central

Yamashita, Masao; Cheung, Martin; Kalale, Chola; Adaniya, Hidehito; Kuwahara, Ryusuke; Shintake, Tsumoru

2017-01-01

We have performed open cell transmission electron microscopy experiments through pure water vapor in the saturation pressure regime (>0.6 kPa), in a modern microscope capable of sub-Å resolution. We have systematically studied achievable pressure levels, stability and gas purity, effective thickness of the water vapor column and associated electron scattering processes, and the effect of gas pressure on electron optical resolution and image contrast. For example, for 1.3 kPa pure water vapor and 300kV electrons, we report pressure stability of ± 20 Pa over tens of minutes, effective thickness of 0.57 inelastic mean free paths, lattice resolution of 0.14 nm on a reference Au specimen, and no significant degradation in contrast or stability of a biological specimen (M13 virus, with 6 nm body diameter). We have also done some brief experiments to confirm feasibility of loading specimens into an in situ water vapor ambient without exposure to intermediate desiccating conditions. Finally, we have also checked if water experiments had any discernible impact on the microscope performance, and report pertinent vacuum and electron optical data, for reference purposes. PMID:29099843

Influence of total beam current on HRTEM image resolution in differentially pumped ETEM with nitrogen gas.

PubMed

Bright, A N; Yoshida, K; Tanaka, N

2013-01-01

Environmental transmission electron microscopy (ETEM) enables the study of catalytic and other reaction processes as they occur with Angstrom-level resolution. The microscope used is a dedicated ETEM (Titan ETEM, FEI Company) with a differential pumping vacuum system and apertures, allowing aberration corrected high-resolution transmission electron microscopy (HRTEM) imaging to be performed with gas pressures up to 20 mbar in the sample area and with significant advantages over membrane-type E-cell holders. The effect on image resolution of varying the nitrogen gas pressure, electron beam current density and total beam current were measured using information limit (Young's fringes) on a standard cross grating sample and from silicon crystal lattice imaging. As expected, increasing gas pressure causes a decrease in HRTEM image resolution. However, the total electron beam current also causes big changes in the image resolution (lower beam current giving better resolution), whereas varying the beam current density has almost no effect on resolution, a result that has not been reported previously. This behavior is seen even with zero-loss filtered imaging, which we believe shows that the drop in resolution is caused by elastic scattering at gas ions created by the incident electron beam. Suitable conditions for acquiring high resolution images in a gas environment are discussed. Lattice images at nitrogen pressures up to 16 mbar are shown, with 0.12 nm information transfer at 4 mbar. Copyright © 2012 Elsevier B.V. All rights reserved.

Scanning transmission electron microscopy and its application to the study of nanoparticles and nanoparticle systems.

PubMed

Liu, Jingyue

2005-06-01

Scanning transmission electron microscopy (STEM) techniques can provide imaging, diffraction and spectroscopic information, either simultaneously or in a serial manner, of the specimen with an atomic or a sub-nanometer spatial resolution. High-resolution STEM imaging, when combined with nanodiffraction, atomic resolution electron energy-loss spectroscopy and nanometer resolution X-ray energy dispersive spectroscopy techniques, is critical to the fundamental studies of importance to nanoscience and nanotechnology. The availability of sub-nanometer or sub-angstrom electron probes in a STEM instrument, due to the use of a field emission gun and aberration correctors, ensures the greatest capabilities for studies of sizes, shapes, defects, crystal and surface structures, and compositions and electronic states of nanometer-size regions of thin films, nanoparticles and nanoparticle systems. The various imaging, diffraction and spectroscopy modes available in a dedicated STEM or a field emission TEM/STEM instrument are reviewed and the application of these techniques to the study of nanoparticles and nanostructured catalysts is used as an example to illustrate the critical role of the various STEM techniques in nanotechnology and nanoscience research.

Relative merits and limiting factors for x-ray and electron microscopy of thick, hydrated organic materials

DOE PAGES

Du, Ming; Jacobsen, Chris

2017-10-07

Electron and x-ray microscopes allow one to image the entire, unlabeled structure of hydrated materials at a resolution well beyond what visible light microscopes can achieve. However, both approaches involve ionizing radiation, so that radiation damage must be considered as one of the limits to imaging. Drawing upon earlier work, we describe here a unified approach to estimating the image contrast (and thus the required exposure and corresponding radiation dose) in both x-ray and electron microscopy. This approach accounts for factors such as plural and inelastic scattering, and (in electron microscopy) the use of energy filters to obtain so-called "zeromore » loss" images. As expected, it shows that electron microscopy offers lower dose for specimens thinner than about 1 mu m (such as for studies of macromolecules, viruses, bacteria and archaebacteria, and thin sectioned material), while x-ray microscopy offers superior characteristics for imaging thicker specimen such as whole eukaryotic cells, thick-sectioned tissues, and organs. The required radiation dose scales strongly as a function of the desired spatial resolution, allowing one to understand the limits of live and frozen hydrated specimen imaging. Lastly, we consider the factors limiting x-ray microscopy of thicker materials, suggesting that specimens as thick as a whole mouse brain can be imaged with x-ray microscopes without significant image degradation should appropriate image reconstruction methods be identified.« less

Relative merits and limiting factors for x-ray and electron microscopy of thick, hydrated organic materials

DOE Office of Scientific and Technical Information (OSTI.GOV)

Du, Ming; Jacobsen, Chris

Electron and x-ray microscopes allow one to image the entire, unlabeled structure of hydrated materials at a resolution well beyond what visible light microscopes can achieve. However, both approaches involve ionizing radiation, so that radiation damage must be considered as one of the limits to imaging. Drawing upon earlier work, we describe here a unified approach to estimating the image contrast (and thus the required exposure and corresponding radiation dose) in both x-ray and electron microscopy. This approach accounts for factors such as plural and inelastic scattering, and (in electron microscopy) the use of energy filters to obtain so-called "zeromore » loss" images. As expected, it shows that electron microscopy offers lower dose for specimens thinner than about 1 mu m (such as for studies of macromolecules, viruses, bacteria and archaebacteria, and thin sectioned material), while x-ray microscopy offers superior characteristics for imaging thicker specimen such as whole eukaryotic cells, thick-sectioned tissues, and organs. The required radiation dose scales strongly as a function of the desired spatial resolution, allowing one to understand the limits of live and frozen hydrated specimen imaging. Lastly, we consider the factors limiting x-ray microscopy of thicker materials, suggesting that specimens as thick as a whole mouse brain can be imaged with x-ray microscopes without significant image degradation should appropriate image reconstruction methods be identified.« less

Transmission electron microscopy of amyloid fibrils.

PubMed

Gras, Sally L; Waddington, Lynne J; Goldie, Kenneth N

2011-01-01

Transmission Electron Microscopy of negatively stained and cryo-prepared specimens allows amyloid fibrils to be visualised at high resolution in a dried or a hydrated state, and is an essential method for characterising the morphology of fibrils and pre-fibrillar species. We outline the key steps involved in the preparation and observation of samples using negative staining and cryo-electron preservation. We also discuss methods to measure fibril characteristics, such as fibril width, from electron micrographs.

A Dose-Rate Effect in Single-Particle Electron Microscopy

PubMed Central

Chen, James Z.; Sachse, Carsten; Xu, Chen; Mielke, Thorsten; Spahn, Christian M. T.; Grigorieff, Nikolaus

2008-01-01

A low beam-intensity, low electron-dose imaging method has been developed for single-particle electron cryo-microscopy (cryo-EM). Experiments indicate that the new technique can reduce beam-induced specimen movement and secondary radiolytic effects, such as “bubbling”. The improvement in image quality, especially for multiple-exposure data collection, will help single-particle cryo-EM to reach higher resolution. PMID:17977018

Diverse protocols for correlative super-resolution fluorescence imaging and electron microscopy of chemically fixed samples

PubMed Central

Kopek, Benjamin G.; Paez-Segala, Maria G.; Shtengel, Gleb; Sochacki, Kem A.; Sun, Mei G.; Wang, Yalin; Xu, C. Shan; van Engelenburg, Schuyler B.; Taraska, Justin W.; Looger, Loren L.; Hess, Harald F.

2017-01-01

Our groups have recently developed related approaches for sample preparation for super-resolution imaging within endogenous cellular environments using correlative light and electron microscopy (CLEM). Four distinct techniques for preparing and acquiring super-resolution CLEM datasets on aldehyde-fixed specimens are provided, including Tokuyasu cryosectioning, whole-cell mount, cell unroofing and platinum replication, and resin embedding and sectioning. Choice of the best protocol for a given application depends on a number of criteria that are discussed in detail. Tokuyasu cryosectioning is relatively rapid but is limited to small, delicate specimens. Whole-cell mount has the simplest sample preparation but is restricted to surface structures. Cell unroofing and platinum replica creates high-contrast, 3-dimensional images of the cytoplasmic surface of the plasma membrane, but is more challenging than whole-cell mount. Resin embedding permits serial sectioning of large samples, but is limited to osmium-resistant probes, and is technically difficult. Expected results from these protocols include super-resolution localization (~10–50 nm) of fluorescent targets within the context of electron microscopy ultrastructure, which can help address cell biological questions. These protocols can be completed in 2–7 days, are compatible with a number of super-resolution imaging protocols, and are broadly applicable across biology. PMID:28384138

Image improvement and three-dimensional reconstruction using holographic image processing

NASA Technical Reports Server (NTRS)

Stroke, G. W.; Halioua, M.; Thon, F.; Willasch, D. H.

1977-01-01

Holographic computing principles make possible image improvement and synthesis in many cases of current scientific and engineering interest. Examples are given for the improvement of resolution in electron microscopy and 3-D reconstruction in electron microscopy and X-ray crystallography, following an analysis of optical versus digital computing in such applications.

Aberrated electron probes for magnetic spectroscopy with atomic resolution: Theory and practical aspects

DOE PAGES

Rusz, Ján; Idrobo, Juan Carlos

2016-03-24

It was recently proposed that electron magnetic circular dichroism (EMCD) can be measured in scanning transmission electron microscopy (STEM) with atomic resolution by tuning the phase distribution of a electron beam. Here, we describe the theoretical and practical aspects for the detection of out-of-plane and in-plane magnetization utilizing atomic size electron probes. Here we present the calculated optimized astigmatic probes and discuss how to achieve them experimentally.

Three phase crystallography and solute distribution analysis during residual austenite decomposition in tempered nanocrystalline bainitic steels

DOE Office of Scientific and Technical Information (OSTI.GOV)

Caballero, F.G.; Yen, Hung-Wei; Australian Centre for Microscopy and Microanalysis, The University of Sydney, NSW 2006

2014-02-15

Interphase carbide precipitation due to austenite decomposition was investigated by high resolution transmission electron microscopy and atom probe tomography in tempered nanostructured bainitic steels. Results showed that cementite (θ) forms by a paraequilibrium transformation mechanism at the bainitic ferrite–austenite interface with a simultaneous three phase crystallographic orientation relationship. - Highlights: • Interphase carbide precipitation due to austenite decomposition • Tempered nanostructured bainitic steels • High resolution transmission electron microscopy and atom probe tomography • Paraequilibrium θ with three phase crystallographic orientation relationship.

7A projection map of the S-layer protein sbpA obtained with trehalose-embedded monolayer crystals.

PubMed

Norville, Julie E; Kelly, Deborah F; Knight, Thomas F; Belcher, Angela M; Walz, Thomas

2007-12-01

Two-dimensional crystallization on lipid monolayers is a versatile tool to obtain structural information of proteins by electron microscopy. An inherent problem with this approach is to prepare samples in a way that preserves the crystalline order of the protein array and produces specimens that are sufficiently flat for high-resolution data collection at high tilt angles. As a test specimen to optimize the preparation of lipid monolayer crystals for electron microscopy imaging, we used the S-layer protein sbpA, a protein with potential for designing arrays of both biological and inorganic materials with engineered properties for a variety of nanotechnology applications. Sugar embedding is currently considered the best method to prepare two-dimensional crystals of membrane proteins reconstituted into lipid bilayers. We found that using a loop to transfer lipid monolayer crystals to an electron microscopy grid followed by embedding in trehalose and quick-freezing in liquid ethane also yielded the highest resolution images for sbpA lipid monolayer crystals. Using images of specimens prepared in this way we could calculate a projection map of sbpA at 7A resolution, one of the highest resolution projection structures obtained with lipid monolayer crystals to date.

Copper Decoration of Carbon Nanotubes and High Resolution Electron Microscopy

NASA Astrophysics Data System (ADS)

Probst, Camille

A new process of decorating carbon nanotubes with copper was developed for the fabrication of nanocomposite aluminum-nanotubes. The process consists of three stages: oxidation, activation and electroless copper plating on the nanotubes. The oxidation step was required to create chemical function on the nanotubes, essential for the activation step. Then, catalytic nanoparticles of tin-palladium were deposited on the tubes. Finally, during the electroless copper plating, copper particles with a size between 20 and 60 nm were uniformly deposited on the nanotubes surface. The reproducibility of the process was shown by using another type of carbon nanotube. The fabrication of nanocomposites aluminum-nanotubes was tested by aluminum vacuum infiltration. Although the infiltration of carbon nanotubes did not produce the expected results, an interesting electron microscopy sample was discovered during the process development: the activated carbon nanotubes. Secondly, scanning transmitted electron microscopy (STEM) imaging in SEM was analysed. The images were obtained with a new detector on the field emission scanning electron microscope (Hitachi S-4700). Various parameters were analysed with the use of two different samples: the activated carbon nanotubes (previously obtained) and gold-palladium nanodeposits. Influences of working distance, accelerating voltage or sample used on the spatial resolution of images obtained with SMART (Scanning Microscope Assessment and Resolution Testing) were analysed. An optimum working distance for the best spatial resolution related to the sample analysed was found for the imaging in STEM mode. Finally, relation between probe size and spatial resolution of backscattered electrons (BSE) images was studied. An image synthesis method was developed to generate the BSE images from backscattered electrons coefficients obtained with CASINO software. Spatial resolution of images was determined using SMART. The analysis shown that using a probe size smaller than the size of the observed object (sample features) does not improve the spatial resolution. In addition, the effects of the accelerating voltage, the current intensity and the sample geometry and composition were analysed.

Application of environmental scanning electron microscopy to determine biological surface structure.

PubMed

Kirk, S E; Skepper, J N; Donald, A M

2009-02-01

The use of environmental scanning electron microscopy in biology is growing as more becomes understood about the advantages and limitations of the technique. These are discussed and we include new evidence about the effect of environmental scanning electron microscopy imaging on the viability of mammalian cells. We show that although specimen preparation for high-vacuum scanning electron microscopy introduces some artefacts, there are also challenges in the use of environmental scanning electron microscopy, particularly at higher resolutions. This suggests the two technologies are best used in combination. We have used human monocyte-derived macrophages as a test sample, imaging their complicated and delicate membrane ruffles and protrusions. We have also explored the possibility of using environmental scanning electron microscopy for dynamic experiments, finding that mammalian cells cannot be imaged and kept alive in the environmental scanning electron microscopy. The dehydration step in which the cell surface is exposed causes irreversible damage, probably via loss of membrane integrity during liquid removal in the specimen chamber. Therefore, mammalian cells should be imaged after fixation where possible to protect against damage as a result of chamber conditions.

Heat- and electron-beam-induced transport of gold particles into silicon oxide and silicon studied by in situ high-resolution transmission electron microscopy.

PubMed

Biskupek, Johannes; Kaiser, Ute; Falk, Fritz

2008-06-01

In this study, we describe the transport of gold (Au) nanoparticles from the surface into crystalline silicon (Si) covered by silicon oxide (SiO(2)) as revealed by in situ high-resolution transmission electron microscopy. Complete crystalline Au nanoparticles sink through the SiO(2) layer into the Si substrate when high-dose electron irradiation is applied and temperature is raised above 150 degrees C. Above temperatures of 250 degrees C, the Au nanoparticles finally dissolve into fragments accompanied by crystallization of the amorphized Si substrate around these fragments. The transport process is explained by a wetting process followed by Stokes motion. Modelling this process yields boundaries for the interface energies involved.

Resonant tunneling through electronic trapping states in thin MgO magnetic junctions.

PubMed

Teixeira, J M; Ventura, J; Araujo, J P; Sousa, J B; Wisniowski, P; Cardoso, S; Freitas, P P

2011-05-13

We report an inelastic electron tunneling spectroscopy study on MgO magnetic junctions with thin barriers (0.85-1.35 nm). Inelastic electron tunneling spectroscopy reveals resonant electronic trapping within the barrier for voltages V>0.15  V. These trapping features are associated with defects in the barrier crystalline structure, as confirmed by high-resolution transmission electron microscopy. Such defects are responsible for resonant tunneling due to energy levels that are formed in the barrier. A model was applied to determine the average location and energy level of the traps, indicating that they are mostly located in the middle of the MgO barrier, in accordance with the high-resolution transmission electron microscopy data and trap-assisted tunneling conductance theory. Evidence of the influence of trapping on the voltage dependence of tunnel magnetoresistance is shown.

DMD-based LED-illumination super-resolution and optical sectioning microscopy.

PubMed

Dan, Dan; Lei, Ming; Yao, Baoli; Wang, Wen; Winterhalder, Martin; Zumbusch, Andreas; Qi, Yujiao; Xia, Liang; Yan, Shaohui; Yang, Yanlong; Gao, Peng; Ye, Tong; Zhao, Wei

2013-01-01

Super-resolution three-dimensional (3D) optical microscopy has incomparable advantages over other high-resolution microscopic technologies, such as electron microscopy and atomic force microscopy, in the study of biological molecules, pathways and events in live cells and tissues. We present a novel approach of structured illumination microscopy (SIM) by using a digital micromirror device (DMD) for fringe projection and a low-coherence LED light for illumination. The lateral resolution of 90 nm and the optical sectioning depth of 120 μm were achieved. The maximum acquisition speed for 3D imaging in the optical sectioning mode was 1.6×10(7) pixels/second, which was mainly limited by the sensitivity and speed of the CCD camera. In contrast to other SIM techniques, the DMD-based LED-illumination SIM is cost-effective, ease of multi-wavelength switchable and speckle-noise-free. The 2D super-resolution and 3D optical sectioning modalities can be easily switched and applied to either fluorescent or non-fluorescent specimens.

DMD-based LED-illumination Super-resolution and optical sectioning microscopy

PubMed Central

Dan, Dan; Lei, Ming; Yao, Baoli; Wang, Wen; Winterhalder, Martin; Zumbusch, Andreas; Qi, Yujiao; Xia, Liang; Yan, Shaohui; Yang, Yanlong; Gao, Peng; Ye, Tong; Zhao, Wei

2013-01-01

Super-resolution three-dimensional (3D) optical microscopy has incomparable advantages over other high-resolution microscopic technologies, such as electron microscopy and atomic force microscopy, in the study of biological molecules, pathways and events in live cells and tissues. We present a novel approach of structured illumination microscopy (SIM) by using a digital micromirror device (DMD) for fringe projection and a low-coherence LED light for illumination. The lateral resolution of 90 nm and the optical sectioning depth of 120 μm were achieved. The maximum acquisition speed for 3D imaging in the optical sectioning mode was 1.6×107 pixels/second, which was mainly limited by the sensitivity and speed of the CCD camera. In contrast to other SIM techniques, the DMD-based LED-illumination SIM is cost-effective, ease of multi-wavelength switchable and speckle-noise-free. The 2D super-resolution and 3D optical sectioning modalities can be easily switched and applied to either fluorescent or non-fluorescent specimens. PMID:23346373

Phase contrast scanning transmission electron microscopy imaging of light and heavy atoms at the limit of contrast and resolution.

PubMed

Yücelen, Emrah; Lazić, Ivan; Bosch, Eric G T

2018-02-08

Using state of the art scanning transmission electron microscopy (STEM) it is nowadays possible to directly image single atomic columns at sub-Å resolution. In standard (high angle) annular dark field STEM ((HA)ADF-STEM), however, light elements are usually invisible when imaged together with heavier elements in one image. Here we demonstrate the capability of the recently introduced Integrated Differential Phase Contrast STEM (iDPC-STEM) technique to image both light and heavy atoms in a thin sample at sub-Å resolution. We use the technique to resolve both the Gallium and Nitrogen dumbbells in a GaN crystal in [[Formula: see text

Spectral Interferometry with Electron Microscopes

PubMed Central

Talebi, Nahid

2016-01-01

Interference patterns are not only a defining characteristic of waves, but also have several applications; characterization of coherent processes and holography. Spatial holography with electron waves, has paved the way towards space-resolved characterization of magnetic domains and electrostatic potentials with angstrom spatial resolution. Another impetus in electron microscopy has been introduced by ultrafast electron microscopy which uses pulses of sub-picosecond durations for probing a laser induced excitation of the sample. However, attosecond temporal resolution has not yet been reported, merely due to the statistical distribution of arrival times of electrons at the sample, with respect to the laser time reference. This is however, the very time resolution which will be needed for performing time-frequency analysis. These difficulties are addressed here by proposing a new methodology to improve the synchronization between electron and optical excitations through introducing an efficient electron-driven photon source. We use focused transition radiation of the electron as a pump for the sample. Due to the nature of transition radiation, the process is coherent. This technique allows us to perform spectral interferometry with electron microscopes, with applications in retrieving the phase of electron-induced polarizations and reconstructing dynamics of the induced vector potential. PMID:27649932

Correlative light-electron fractography for fatigue striations characterization in metallic alloys.

PubMed

Hein, Luis Rogerio de Oliveira; de Oliveira, José Alberto; de Campos, Kamila Amato

2013-09-01

The correlative light-electron fractography technique combines correlative microscopy concepts to the extended depth-from-focus reconstruction method, associating the reliable topographic information of 3-D maps from light microscopy ordered Z-stacks to the finest lateral resolution and large focus depth from scanning electron microscopy. Fatigue striations spacing analysis can be precisely measured, by correcting the mean surface tilting with the knowledge of local elevation data from elevation maps. This new technique aims to improve the accuracy of quantitative fractography in fatigue fracture investigations. Copyright © 2013 Wiley Periodicals, Inc.

Transmission Electron Microscopy Analysis of Skin Lesions from Sporotrichosis Epidemic in Rio de Janeiro, Brazil

PubMed Central

Porto Ferreira, Cassio; Oliveira de Almeida, Ana Cristina; Corte-Real, Suzana

2015-01-01

Transmission electron microscopy can yield useful information in a range of scientific fields; it is capable of imaging at a significantly higher resolution than light microscopes and has been a very useful tool in the identification of morphological changes of the dermis as well as assessment of changes in the extracellular matrix. Our aim is to characterize by electron microscopy the cellular profile of lesions caused by Sporothrix schenckii from the sporotrichosis epidemic in its zoonotic form that occurs in Rio de Janeiro, Brazil. PMID:25653392

Development of an environmental high-voltage electron microscope for reaction science.

PubMed

Tanaka, Nobuo; Usukura, Jiro; Kusunoki, Michiko; Saito, Yahachi; Sasaki, Katuhiro; Tanji, Takayoshi; Muto, Shunsuke; Arai, Shigeo

2013-02-01

Environmental transmission electron microscopy and ultra-high resolution electron microscopic observation using aberration correctors have recently emerged as topics of great interest. The former method is an extension of the so-called in situ electron microscopy that has been performed since the 1970s. Current research in this area has been focusing on dynamic observation with atomic resolution under gaseous atmospheres and in liquids. Since 2007, Nagoya University has been developing a new 1-MV high voltage (scanning) transmission electron microscope that can be used to observe nanomaterials under conditions that include the presence of gases, liquids and illuminating lights, and it can be also used to perform mechanical operations to nanometre-sized areas as well as electron tomography and elemental analysis by electron energy loss spectroscopy. The new instrument has been used to image and analyse various types of samples including biological ones.

Overview of electron crystallography of membrane proteins: crystallization and screening strategies using negative stain electron microscopy.

PubMed

Nannenga, Brent L; Iadanza, Matthew G; Vollmar, Breanna S; Gonen, Tamir

2013-01-01

Electron cryomicroscopy, or cryoEM, is an emerging technique for studying the three-dimensional structures of proteins and large macromolecular machines. Electron crystallography is a branch of cryoEM in which structures of proteins can be studied at resolutions that rival those achieved by X-ray crystallography. Electron crystallography employs two-dimensional crystals of a membrane protein embedded within a lipid bilayer. The key to a successful electron crystallographic experiment is the crystallization, or reconstitution, of the protein of interest. This unit describes ways in which protein can be expressed, purified, and reconstituted into well-ordered two-dimensional crystals. A protocol is also provided for negative stain electron microscopy as a tool for screening crystallization trials. When large and well-ordered crystals are obtained, the structures of both protein and its surrounding membrane can be determined to atomic resolution.

Advanced electron microscopy characterization of nanomaterials for catalysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Su, Dong

Transmission electron microscopy (TEM) has become one of the most powerful techniques in the fields of material science, inorganic chemistry and nanotechnology. In terms of resolutions, advanced TEM may reach a high spatial resolution of 0.05 nm, a high energy-resolution of 7 meV. In addition, in situ TEM can help researcher to image the process happened within 1 ms. This paper reviews the recent technical approaches of applying advanced TEM characterization on nanomaterials for catalysis. The text is organized according to the demanded information of nanocrystals from the perspective of application: for example, size, composition, phase, strain, and morphology. Themore » electron beam induced effect and in situ TEM are also introduced. As a result, I hope this review can help the scientists in related fields to take advantage of advanced TEM to their own researches.« less

Advanced electron microscopy characterization of nanomaterials for catalysis

DOE PAGES

Su, Dong

2017-04-01

Transmission electron microscopy (TEM) has become one of the most powerful techniques in the fields of material science, inorganic chemistry and nanotechnology. In terms of resolutions, advanced TEM may reach a high spatial resolution of 0.05 nm, a high energy-resolution of 7 meV. In addition, in situ TEM can help researcher to image the process happened within 1 ms. This paper reviews the recent technical approaches of applying advanced TEM characterization on nanomaterials for catalysis. The text is organized according to the demanded information of nanocrystals from the perspective of application: for example, size, composition, phase, strain, and morphology. Themore » electron beam induced effect and in situ TEM are also introduced. As a result, I hope this review can help the scientists in related fields to take advantage of advanced TEM to their own researches.« less

Three-dimensional imaging of adherent cells using FIB/SEM and STEM.

PubMed

Villinger, Clarissa; Schauflinger, Martin; Gregorius, Heiko; Kranz, Christine; Höhn, Katharina; Nafeey, Soufi; Walther, Paul

2014-01-01

In this chapter we describe three different approaches for three-dimensional imaging of electron microscopic samples: serial sectioning transmission electron microscopy (TEM), scanning transmission electron microscopy (STEM) tomography, and focused ion beam/scanning electron microscopy (FIB/SEM) tomography. With these methods, relatively large volumes of resin-embedded biological structures can be analyzed at resolutions of a few nm within a reasonable expenditure of time. The traditional method is serial sectioning and imaging the same area in all sections. Another method is TEM tomography that involves tilting a section in the electron beam and then reconstruction of the volume by back projection of the images. When the scanning transmission (STEM) mode is used, thicker sections (up to 1 μm) can be analyzed. The third approach presented here is focused ion beam/scanning electron microscopy (FIB/SEM) tomography, in which a sample is repeatedly milled with a focused ion beam (FIB) and each newly produced block face is imaged with the scanning electron microscope (SEM). This process can be repeated ad libitum in arbitrary small increments allowing 3D analysis of relatively large volumes such as eukaryotic cells. We show that resolution of this approach is considerably improved when the secondary electron signal is used. However, the most important prerequisite for three-dimensional imaging is good specimen preparation. For all three imaging methods, cryo-fixed (high-pressure frozen) and freeze-substituted samples have been used.

Contribution of high-resolution correlative imaging techniques in the study of the liver sieve in three-dimensions.

PubMed

Braet, Filip; Wisse, Eddie; Bomans, Paul; Frederik, Peter; Geerts, Willie; Koster, Abraham; Soon, Lilian; Ringer, Simon

2007-03-01

Correlative microscopy has become increasingly important for the analysis of the structure, function, and dynamics of cells. This is largely due to the result of recent advances in light-, probe-, laser- and various electron microscopy techniques that facilitate three-dimensional studies. Furthermore, the improved understanding in the past decade of imaging cell compartments in the third dimension has resulted largely from the availability of powerful computers, fast high-resolution CCD cameras, specifically developed imaging analysis software, and various probes designed for labeling living and or fixed cells. In this paper, we review different correlative high-resolution imaging methodologies and how these microscopy techniques facilitated the accumulation of new insights in the morpho-functional and structural organization of the hepatic sieve. Various aspects of hepatic endothelial fenestrae regarding their structure, origin, dynamics, and formation will be explored throughout this paper by comparing the results of confocal laser scanning-, correlative fluorescence and scanning electron-, atomic force-, and whole-mount electron microscopy. Furthermore, the recent advances of vitrifying cells with the vitrobot in combination with the glove box for the preparation of cells for cryo-electron microscopic investigation will be discussed. Finally, the first transmission electron tomography data of the liver sieve in three-dimensions are presented. The obtained data unambiguously show the involvement of special domains in the de novo formation and disappearance of hepatic fenestrae, and focuses future research into the (supra)molecular structure of the fenestrae-forming center, defenestration center and fenestrae-, and sieve plate cytoskeleton ring by using advanced cryo-electron tomography. (c) 2007 Wiley-Liss, Inc.

Scanning superlens microscopy for non-invasive large field-of-view visible light nanoscale imaging

NASA Astrophysics Data System (ADS)

Wang, Feifei; Liu, Lianqing; Yu, Haibo; Wen, Yangdong; Yu, Peng; Liu, Zhu; Wang, Yuechao; Li, Wen Jung

2016-12-01

Nanoscale correlation of structural information acquisition with specific-molecule identification provides new insight for studying rare subcellular events. To achieve this correlation, scanning electron microscopy has been combined with super-resolution fluorescent microscopy, despite its destructivity when acquiring biological structure information. Here we propose time-efficient non-invasive microsphere-based scanning superlens microscopy that enables the large-area observation of live-cell morphology or sub-membrane structures with sub-diffraction-limited resolution and is demonstrated by observing biological and non-biological objects. This microscopy operates in both non-invasive and contact modes with ~200 times the acquisition efficiency of atomic force microscopy, which is achieved by replacing the point of an atomic force microscope tip with an imaging area of microspheres and stitching the areas recorded during scanning, enabling sub-diffraction-limited resolution. Our method marks a possible path to non-invasive cell imaging and simultaneous tracking of specific molecules with nanoscale resolution, facilitating the study of subcellular events over a total cell period.

Active control of bright electron beams with RF optics for femtosecond microscopy

DOE PAGES

Williams, J.; Zhou, F.; Sun, T.; ...

2017-08-01

A frontier challenge in implementing femtosecond electron microscopy is to gain precise optical control of intense beams to mitigate collective space charge effects for significantly improving the throughput. In this paper, we explore the flexible uses of an RF cavity as a longitudinal lens in a high-intensity beam column for condensing the electron beams both temporally and spectrally, relevant to the design of ultrafast electron microscopy. Through the introduction of a novel atomic grating approach for characterization of electron bunch phase space and control optics, we elucidate the principles for predicting and controlling the phase space dynamics to reach optimalmore » compressions at various electron densities and generating conditions. We provide strategies to identify high-brightness modes, achieving ~100 fs and ~1 eV resolutions with 10 6 electrons per bunch, and establish the scaling of performance for different bunch charges. These results benchmark the sensitivity and resolution from the fundamental beam brightness perspective and also validate the adaptive optics concept to enable delicate control of the density-dependent phase space structures to optimize the performance, including delivering ultrashort, monochromatic, high-dose, or coherent electron bunches.« less

Active control of bright electron beams with RF optics for femtosecond microscopy

PubMed Central

Williams, J.; Zhou, F.; Sun, T.; Tao, Z.; Chang, K.; Makino, K.; Berz, M.; Duxbury, P. M.; Ruan, C.-Y.

2017-01-01

A frontier challenge in implementing femtosecond electron microscopy is to gain precise optical control of intense beams to mitigate collective space charge effects for significantly improving the throughput. Here, we explore the flexible uses of an RF cavity as a longitudinal lens in a high-intensity beam column for condensing the electron beams both temporally and spectrally, relevant to the design of ultrafast electron microscopy. Through the introduction of a novel atomic grating approach for characterization of electron bunch phase space and control optics, we elucidate the principles for predicting and controlling the phase space dynamics to reach optimal compressions at various electron densities and generating conditions. We provide strategies to identify high-brightness modes, achieving ∼100 fs and ∼1 eV resolutions with 106 electrons per bunch, and establish the scaling of performance for different bunch charges. These results benchmark the sensitivity and resolution from the fundamental beam brightness perspective and also validate the adaptive optics concept to enable delicate control of the density-dependent phase space structures to optimize the performance, including delivering ultrashort, monochromatic, high-dose, or coherent electron bunches. PMID:28868325

Perspectives on in situ electron microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zheng, Haimei; Zhu, Yimei

In situ transmission electron microscopy (TEM) with the ability to reveal materials dynamic processes with high spatial and temporal resolution has attracted significant interest. The recent advances in in situ methods, including liquid and gas sample environment, pump-probe ultrafast microscopy, nanomechanics and ferroelectric domain switching the aberration corrected electron optics as well as fast electron detector has opened new opportunities to extend the impact of in situ TEM in broad areas of research ranging from materials science to chemistry, physics and biology. Here in this paper, we highlight the development of liquid environment electron microscopy and its applications in themore » study of colloidal nanoparticle growth, electrochemical processes and others; in situ study of topological vortices in ferroelectric and ferromagnetic materials. At the end, perspectives of future in situ TEM are provided.« less

Perspectives on in situ electron microscopy

DOE PAGES

Zheng, Haimei; Zhu, Yimei

2017-03-29

In situ transmission electron microscopy (TEM) with the ability to reveal materials dynamic processes with high spatial and temporal resolution has attracted significant interest. The recent advances in in situ methods, including liquid and gas sample environment, pump-probe ultrafast microscopy, nanomechanics and ferroelectric domain switching the aberration corrected electron optics as well as fast electron detector has opened new opportunities to extend the impact of in situ TEM in broad areas of research ranging from materials science to chemistry, physics and biology. Here in this paper, we highlight the development of liquid environment electron microscopy and its applications in themore » study of colloidal nanoparticle growth, electrochemical processes and others; in situ study of topological vortices in ferroelectric and ferromagnetic materials. At the end, perspectives of future in situ TEM are provided.« less

Photon-assisted electron energy loss spectroscopy and ultrafast imaging.

PubMed

Howie, Archie

2009-08-01

A variety of ways is described in which photons can be used not only for ultrafast electron microscopy but also to enormously widen the energy range of spatially-resolved electron spectroscopy. Periodic chains of femtosecond laser pulses are a particularly important and accurately timed source for single-shot imaging and diffraction as well as for several forms of pump-probe microscopy at even higher spatial resolution and sub-picosecond timing. Many exciting new fields are opened up for study by these developments. Ultrafast, single shot diffraction with intense pulses of X-rays supplemented by phase retrieval techniques may eventually offer a challenging alternative and purely photon-based route to dynamic imaging at high spatial resolution.

Relative merits and limiting factors for x-ray and electron microscopy of thick, hydrated organic materials.

PubMed

Du, Ming; Jacobsen, Chris

2018-01-01

Electron and x-ray microscopes allow one to image the entire, unlabeled structure of hydrated materials at a resolution well beyond what visible light microscopes can achieve. However, both approaches involve ionizing radiation, so that radiation damage must be considered as one of the limits to imaging. Drawing upon earlier work, we describe here a unified approach to estimating the image contrast (and thus the required exposure and corresponding radiation dose) in both x-ray and electron microscopy. This approach accounts for factors such as plural and inelastic scattering, and (in electron microscopy) the use of energy filters to obtain so-called "zero loss" images. As expected, it shows that electron microscopy offers lower dose for specimens thinner than about 1 µm (such as for studies of macromolecules, viruses, bacteria and archaebacteria, and thin sectioned material), while x-ray microscopy offers superior characteristics for imaging thicker specimen such as whole eukaryotic cells, thick-sectioned tissues, and organs. The required radiation dose scales strongly as a function of the desired spatial resolution, allowing one to understand the limits of live and frozen hydrated specimen imaging. Finally, we consider the factors limiting x-ray microscopy of thicker materials, suggesting that specimens as thick as a whole mouse brain can be imaged with x-ray microscopes without significant image degradation should appropriate image reconstruction methods be identified. Copyright © 2017 Elsevier B.V. All rights reserved.

An overview of state-of-the-art image restoration in electron microscopy.

PubMed

Roels, J; Aelterman, J; Luong, H Q; Lippens, S; Pižurica, A; Saeys, Y; Philips, W

2018-06-08

In Life Science research, electron microscopy (EM) is an essential tool for morphological analysis at the subcellular level as it allows for visualization at nanometer resolution. However, electron micrographs contain image degradations such as noise and blur caused by electromagnetic interference, electron counting errors, magnetic lens imperfections, electron diffraction, etc. These imperfections in raw image quality are inevitable and hamper subsequent image analysis and visualization. In an effort to mitigate these artefacts, many electron microscopy image restoration algorithms have been proposed in the last years. Most of these methods rely on generic assumptions on the image or degradations and are therefore outperformed by advanced methods that are based on more accurate models. Ideally, a method will accurately model the specific degradations that fit the physical acquisition settings. In this overview paper, we discuss different electron microscopy image degradation solutions and demonstrate that dedicated artefact regularisation results in higher quality restoration and is applicable through recently developed probabilistic methods. © 2018 The Authors Journal of Microscopy © 2018 Royal Microscopical Society.

Detection, mapping, and quantification of single walled carbon nanotubes in histological specimens with photoacoustic microscopy.

PubMed

Avti, Pramod K; Hu, Song; Favazza, Christopher; Mikos, Antonios G; Jansen, John A; Shroyer, Kenneth R; Wang, Lihong V; Sitharaman, Balaji

2012-01-01

In the present study, the efficacy of multi-scale photoacoustic microscopy (PAM) was investigated to detect, map, and quantify trace amounts [nanograms (ng) to micrograms (µg)] of SWCNTs in a variety of histological tissue specimens consisting of cancer and benign tissue biopsies (histological specimens from implanted tissue engineering scaffolds). Optical-resolution (OR) and acoustic-resolution (AR)--Photoacoustic microscopy (PAM) was employed to detect, map and quantify the SWCNTs in a variety of tissue histological specimens and compared with other optical techniques (bright-field optical microscopy, Raman microscopy, near infrared (NIR) fluorescence microscopy). Both optical-resolution and acoustic-resolution PAM, allow the detection and quantification of SWCNTs in histological specimens with scalable spatial resolution and depth penetration. The noise-equivalent detection sensitivity to SWCNTs in the specimens was calculated to be as low as ∼7 pg. Image processing analysis further allowed the mapping, distribution, and quantification of the SWCNTs in the histological sections. The results demonstrate the potential of PAM as a promising imaging technique to detect, map, and quantify SWCNTs in histological specimens, and could complement the capabilities of current optical and electron microscopy techniques in the analysis of histological specimens containing SWCNTs.

Structured Illumination Microscopy for the Investigation of Synaptic Structure and Function.

PubMed

Hong, Soyon; Wilton, Daniel K; Stevens, Beth; Richardson, Douglas S

2017-01-01

The neuronal synapse is a primary building block of the nervous system to which alterations in structure or function can result in numerous pathologies. Studying its formation and elimination is the key to understanding how brains are wired during development, maintained throughout adulthood plasticity, and disrupted during disease. However, due to its diffraction-limited size, investigations of the synaptic junction at the structural level have primarily relied on labor-intensive electron microscopy or ultra-thin section array tomography. Recent advances in the field of super-resolution light microscopy now allow researchers to image synapses and associated molecules with high-spatial resolution, while taking advantage of the key characteristics of light microscopy, such as easy sample preparation and the ability to detect multiple targets with molecular specificity. One such super-resolution technique, Structured Illumination Microscopy (SIM), has emerged as an attractive method to examine synapse structure and function. SIM requires little change in standard light microscopy sample preparation steps, but results in a twofold improvement in both lateral and axial resolutions compared to widefield microscopy. The following protocol outlines a method for imaging synaptic structures at resolutions capable of resolving the intricacies of these neuronal connections.

The Application of High-Resolution Electron Microscopy to Problems in Solid State Chemistry: The Exploits of a Peeping TEM.

ERIC Educational Resources Information Center

Eyring, LeRoy

1980-01-01

Describes methods for using the high-resolution electron microscope in conjunction with other tools to reveal the identity and environment of atoms. Problems discussed include the ultimate structure of real crystalline solids including defect structure and the mechanisms of chemical reactions. (CS)

The impact of recent improvements in cryo-electron microscopy technology on the understanding of bacterial ribosome assembly

PubMed Central

Razi, Aida; Britton, Robert A.

2017-01-01

Abstract Cryo-electron microscopy (cryo-EM) had played a central role in the study of ribosome structure and the process of translation in bacteria since the development of this technique in the mid 1980s. Until recently cryo-EM structures were limited to ∼10 Å in the best cases. However, the recent advent of direct electron detectors has greatly improved the resolution of cryo-EM structures to the point where atomic resolution is now achievable. This improved resolution will allow cryo-EM to make groundbreaking contributions in essential aspects of ribosome biology, including the assembly process. In this review, we summarize important insights that cryo-EM, in combination with chemical and genetic approaches, has already brought to our current understanding of the ribosomal assembly process in bacteria using previous detector technology. More importantly, we discuss how the higher resolution structures now attainable with direct electron detectors can be leveraged to propose precise testable models regarding this process. These structures will provide an effective platform to develop new antibiotics that target this fundamental cellular process. PMID:28180306

Room temperature chemical synthesis of lead selenide thin films with preferred orientation

NASA Astrophysics Data System (ADS)

Kale, R. B.; Sartale, S. D.; Ganesan, V.; Lokhande, C. D.; Lin, Yi-Feng; Lu, Shih-Yuan

2006-11-01

Room temperature chemical synthesis of PbSe thin films was carried out from aqueous ammoniacal solution using Pb(CH3COO)2 as Pb2+ and Na2SeSO3 as Se2- ion sources. The films were characterized by a various techniques including, X-ray diffraction (XRD), energy dispersive X-ray analysis (EDAX), scanning electron microscopy (SEM), transmission electron microscopy (TEM), high resolution transmission electron microscopy (HR-TEM), selected area electron diffraction (SAED), Fast Fourier transform (FFT) and UV-vis-NIR techniques. The study revealed that the PbSe thin film consists of preferentially oriented nanocubes with energy band gap of 0.5 eV.

Imaging of high-angle annular dark-field scanning transmission electron microscopy and observations of GaN-based violet laser diodes.

PubMed

Shiojiri, M; Saijo, H

2006-09-01

The first part of this paper is devoted to physics, to explain high-angle annular dark-field scanning transmission electron microscopy (HAADF-STEM) imaging and to interpret why HAADF-STEM imaging is incoherent, instructing a strict definition of interference and coherence of electron waves. Next, we present our recent investigations of InGaN/GaN multiple quantum wells and AlGaN/GaN strained-layer superlattice claddings in GaN-based violet laser diodes, which have been performed by HAADF-STEM and high-resolution field-emission gun scanning electron microscopy.

Application of He ion microscopy for material analysis

NASA Astrophysics Data System (ADS)

Altmann, F.; Simon, M.; Klengel, R.

2009-05-01

Helium ion beam microscopy (HIM) is a new high resolution imaging technique. The use of Helium ions instead of electrons enables none destructive imaging combined with contrasts quite similar to that from Gallium ion beam imaging. The use of very low probe currents and the comfortable charge compensation using low energy electrons offer imaging of none conductive samples without conductive coating. An ongoing microelectronic sample with Gold/Aluminum interconnects and polymer electronic devices were chosen to evaluate HIM in comparison to scanning electron microscopy (SEM). The aim was to look for key applications of HIM in material analysis. Main focus was on complementary contrast mechanisms and imaging of none conductive samples.

The Probe Profile and Lateral Resolution of Scanning Transmission Electron Microscopy of Thick Specimens

PubMed Central

Demers, Hendrix; Ramachandra, Ranjan; Drouin, Dominique; de Jonge, Niels

2012-01-01

Lateral profiles of the electron probe of scanning transmission electron microscopy (STEM) were simulated at different vertical positions in a micrometers-thick carbon sample. The simulations were carried out using the Monte Carlo method in the CASINO software. A model was developed to fit the probe profiles. The model consisted of the sum of a Gaussian function describing the central peak of the profile, and two exponential decay functions describing the tail of the profile. Calculations were performed to investigate the fraction of unscattered electrons as function of the vertical position of the probe in the sample. Line scans were also simulated over gold nanoparticles at the bottom of a carbon film to calculate the achievable resolution as function of the sample thickness and the number of electrons. The resolution was shown to be noise limited for film thicknesses less than 1 μm. Probe broadening limited the resolution for thicker films. The validity of the simulation method was verified by comparing simulated data with experimental data. The simulation method can be used as quantitative method to predict STEM performance or to interpret STEM images of thick specimens. PMID:22564444

High Resolution Higher Energy X-ray Microscope for Mesoscopic Materials

NASA Astrophysics Data System (ADS)

Snigireva, I.; Snigirev, A.

2013-10-01

We developed a novel X-ray microscopy technique to study mesoscopically structured materials, employing compound refractive lenses. The easily seen advantage of lens-based methodology is the possibility to retrieve high resolution diffraction pattern and real-space images in the same experimental setup. Methodologically the proposed approach is similar to the studies of crystals by high resolution transmission electron microscopy. The proposed microscope was applied for studying of mesoscopic materials such as natural and synthetic opals, inverted photonic crystals.

Angular reconstitution-based 3D reconstructions of nanomolecular structures from superresolution light-microscopy images

PubMed Central

Salas, Desirée; Le Gall, Antoine; Fiche, Jean-Bernard; Valeri, Alessandro; Ke, Yonggang; Bron, Patrick; Bellot, Gaetan

2017-01-01

Superresolution light microscopy allows the imaging of labeled supramolecular assemblies at a resolution surpassing the classical diffraction limit. A serious limitation of the superresolution approach is sample heterogeneity and the stochastic character of the labeling procedure. To increase the reproducibility and the resolution of the superresolution results, we apply multivariate statistical analysis methods and 3D reconstruction approaches originally developed for cryogenic electron microscopy of single particles. These methods allow for the reference-free 3D reconstruction of nanomolecular structures from two-dimensional superresolution projection images. Since these 2D projection images all show the structure in high-resolution directions of the optical microscope, the resulting 3D reconstructions have the best possible isotropic resolution in all directions. PMID:28811371

RBS/C, HRTEM and HRXRD study of damage accumulation in irradiated SrTiO3

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jagielski, Jacek; Jozwik, Przemyslaw A.; Jozwik Biala, Iwona

2013-05-14

Damage accumulation in argon-irradiated SrTiO3 single crystals has been studied by using combination of Rutherford Backscattering/Channeling (RBS/C), High Resolution Transmission Electron Microscopy (HRTEM) and High Resolution X-Ray Diffraction (HRXRD) techniques. The RBS/C spectra were fitted using McChasy, a Monte Carlo simulation code allowing the quantitative analysis of amorphous-like and dislocation-like types of defects. The results were interpreted by using a Multi-Step Damage Accumulation model which assumes, that the damage accumulation occurs in a series of structural transformations, the defect transformations are triggered by a stress caused by formation of a free volume in the irradiated crystal. This assumption has beenmore » confirmed by High Resolution Transmission Electron Microscopy and High Resolution X-Ray Diffraction analysis.« less

An atomic model of brome mosaic virus using direct electron detection and real-space optimization.

PubMed

Wang, Zhao; Hryc, Corey F; Bammes, Benjamin; Afonine, Pavel V; Jakana, Joanita; Chen, Dong-Hua; Liu, Xiangan; Baker, Matthew L; Kao, Cheng; Ludtke, Steven J; Schmid, Michael F; Adams, Paul D; Chiu, Wah

2014-09-04

Advances in electron cryo-microscopy have enabled structure determination of macromolecules at near-atomic resolution. However, structure determination, even using de novo methods, remains susceptible to model bias and overfitting. Here we describe a complete workflow for data acquisition, image processing, all-atom modelling and validation of brome mosaic virus, an RNA virus. Data were collected with a direct electron detector in integrating mode and an exposure beyond the traditional radiation damage limit. The final density map has a resolution of 3.8 Å as assessed by two independent data sets and maps. We used the map to derive an all-atom model with a newly implemented real-space optimization protocol. The validity of the model was verified by its match with the density map and a previous model from X-ray crystallography, as well as the internal consistency of models from independent maps. This study demonstrates a practical approach to obtain a rigorously validated atomic resolution electron cryo-microscopy structure.

Tracking the ultrafast motion of a single molecule by femtosecond orbital imaging

NASA Astrophysics Data System (ADS)

Cocker, Tyler L.; Peller, Dominik; Yu, Ping; Repp, Jascha; Huber, Rupert

2016-11-01

Watching a single molecule move on its intrinsic timescale has been one of the central goals of modern nanoscience, and calls for measurements that combine ultrafast temporal resolution with atomic spatial resolution. Steady-state experiments access the requisite spatial scales, as illustrated by direct imaging of individual molecular orbitals using scanning tunnelling microscopy or the acquisition of tip-enhanced Raman and luminescence spectra with sub-molecular resolution. But tracking the intrinsic dynamics of a single molecule directly in the time domain faces the challenge that interactions with the molecule must be confined to a femtosecond time window. For individual nanoparticles, such ultrafast temporal confinement has been demonstrated by combining scanning tunnelling microscopy with so-called lightwave electronics, which uses the oscillating carrier wave of tailored light pulses to directly manipulate electronic motion on timescales faster even than a single cycle of light. Here we build on ultrafast terahertz scanning tunnelling microscopy to access a state-selective tunnelling regime, where the peak of a terahertz electric-field waveform transiently opens an otherwise forbidden tunnelling channel through a single molecular state. It thereby removes a single electron from an individual pentacene molecule’s highest occupied molecular orbital within a time window shorter than one oscillation cycle of the terahertz wave. We exploit this effect to record approximately 100-femtosecond snapshot images of the orbital structure with sub-ångström spatial resolution, and to reveal, through pump/probe measurements, coherent molecular vibrations at terahertz frequencies directly in the time domain. We anticipate that the combination of lightwave electronics and the atomic resolution of our approach will open the door to visualizing ultrafast photochemistry and the operation of molecular electronics on the single-orbital scale.

In Situ Observation of Carbonaceous Material in the Matrices of CV and CM Carbonaceous Chondrites: Preliminary Results from Energy Filtered Transmission Electron Microscopy

NASA Technical Reports Server (NTRS)

Brearley, A. J.; Abreu, N. M.

2001-01-01

Energy filtered transmission electron microscopy shows that organic matter can be detected in situ in the matrices of carbonaceous chondrites at a spatial resolution of at least 1 nm. In CM chondrites, carbon is often associated with sulfide particles. Additional information is contained in the original extended abstract.

Transmission electron microscopy analysis of skin lesions from sporotrichosis epidemic in Rio de Janeiro, Brazil.

PubMed

Ferreira, Cassio Porto; Oliveira de Almeida, Ana Cristina; Corte-Real, Suzana

2015-02-01

Transmission electron microscopy can yield useful information in a range of scientific fields; it is capable of imaging at a significantly higher resolution than light microscopes and has been a very useful tool in the identification of morphological changes of the dermis as well as assessment of changes in the extracellular matrix. Our aim is to characterize by electron microscopy the cellular profile of lesions caused by Sporothrix schenckii from the sporotrichosis epidemic in its zoonotic form that occurs in Rio de Janeiro, Brazil. © The American Society of Tropical Medicine and Hygiene.

On the state of crystallography at the dawn of the electron microscopy revolution.

PubMed

Higgins, Matthew K; Lea, Susan M

2017-10-01

While protein crystallography has, for many years, been the most used method for structural analysis of macromolecular complexes, remarkable recent advances in high-resolution electron cryo-microscopy led to suggestions that 'the revolution will not be crystallised'. Here we highlight the current success rate, speed and ease of modern crystallographic structure determination and some recent triumphs of both 'classical' crystallography and the use of X-ray free electron lasers. We also outline fundamental differences between structure determination using X-ray crystallography and electron microscopy. We suggest that crystallography will continue to co-exist with electron microscopy as part of an integrated array of methods, allowing structural biologists to focus on fundamental biological questions rather than being constrained by the methods available. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Environmental transmission electron microscopy for catalyst materials using a spherical aberration corrector.

PubMed

Takeda, Seiji; Kuwauchi, Yasufumi; Yoshida, Hideto

2015-04-01

Atomic resolution has been obtained using environmental transmission electron microscopy (ETEM) by installing a spherical aberration corrector (Cs-corrector) on the objective lens. Simultaneously, the technology for controlling the environment around a specimen in ETEM has advanced significantly in the past decade. Quantification methodology has recently been established for deriving relevant experimental data in catalyst materials from substantial and systematic ETEM observation at the atomic scale. With this background, this paper summarizes aspects of the evolutional microscopy technique: necessary conditions for atomic resolution in ETEM; reduction of the scattering of electrons by the medium surrounding a specimen; and an environmental cell for structural imaging of a crystalline specimen. The high spatial resolution of a Cs-corrected ETEM is demonstrated for different observation conditions. After statistical analysis combined with numerical image analysis of ETEM data is briefly described, the recent applications of the Cs-corrected ETEM to catalyst materials are reviewed. For gold nanoparticulate catalysts, the structural information on the reaction sites and adsorption sites are deduced. For Pt nanoparticulate catalysts, ETEM studies elucidate the correlation between the catalytic activity and the morphology of the nanoparticles. These studies also reveal oxidation and reduction on the topmost Pt surface layer at the atomic scale. Finally, current issues and the future perspectives of Cs-corrected ETEM are summarized, including the reproducibility of ETEM observation data, the control of environments, the critical evaluation of electron irradiation effects, the full implementation of transmission electron microscopy technology in ETEM, and the safety issues for an ETEM laboratory. Copyright © 2014 Elsevier B.V. All rights reserved.

Multi-modal Registration for Correlative Microscopy using Image Analogies

PubMed Central

Cao, Tian; Zach, Christopher; Modla, Shannon; Powell, Debbie; Czymmek, Kirk; Niethammer, Marc

2014-01-01

Correlative microscopy is a methodology combining the functionality of light microscopy with the high resolution of electron microscopy and other microscopy technologies for the same biological specimen. In this paper, we propose an image registration method for correlative microscopy, which is challenging due to the distinct appearance of biological structures when imaged with different modalities. Our method is based on image analogies and allows to transform images of a given modality into the appearance-space of another modality. Hence, the registration between two different types of microscopy images can be transformed to a mono-modality image registration. We use a sparse representation model to obtain image analogies. The method makes use of corresponding image training patches of two different imaging modalities to learn a dictionary capturing appearance relations. We test our approach on backscattered electron (BSE) scanning electron microscopy (SEM)/confocal and transmission electron microscopy (TEM)/confocal images. We perform rigid, affine, and deformable registration via B-splines and show improvements over direct registration using both mutual information and sum of squared differences similarity measures to account for differences in image appearance. PMID:24387943

Electron Microscopy of Living Cells During in Situ Fluorescence Microscopy

PubMed Central

Liv, Nalan; van Oosten Slingeland, Daan S. B.; Baudoin, Jean-Pierre; Kruit, Pieter; Piston, David W.; Hoogenboom, Jacob P.

2016-01-01

We present an approach toward dynamic nanoimaging: live fluorescence of cells encapsulated in a bionanoreactor is complemented with in situ scanning electron microscopy (SEM) on an integrated microscope. This allows us to take SEM snapshots on-demand, that is, at a specific location in time, at a desired region of interest, guided by the dynamic fluorescence imaging. We show that this approach enables direct visualization, with EM resolution, of the distribution of bioconjugated quantum dots on cellular extensions during uptake and internalization. PMID:26580231

Tracking the ultrafast motion of a single molecule by femtosecond orbital imaging

PubMed Central

Yu, Ping; Repp, Jascha; Huber, Rupert

2017-01-01

Watching a single molecule move on its intrinsic time scale—one of the central goals of modern nanoscience—calls for measurements that combine ultrafast temporal resolution1–8 with atomic spatial resolution9–30. Steady-state experiments achieve the requisite spatial resolution, as illustrated by direct imaging of individual molecular orbitals using scanning tunnelling microscopy9–11 or the acquisition of tip-enhanced Raman and luminescence spectra with sub-molecular resolution27–29. But tracking the dynamics of a single molecule directly in the time domain faces the challenge that single-molecule excitations need to be confined to an ultrashort time window. A first step towards overcoming this challenge has combined scanning tunnelling microscopy with so-called ‘lightwave electronics”1–8, which uses the oscillating carrier wave of tailored light pulses to directly manipulate electronic motion on time scales faster even than that of a single cycle of light. Here we use such ultrafast terahertz scanning tunnelling microscopy to access a state-selective tunnelling regime, where the peak of a terahertz electric-field waveform transiently opens an otherwise forbidden tunnelling channel through a single molecular state and thereby removes a single electron from an individual pentacene molecule’s highest occupied molecular orbital within a time window shorter than one oscillation cycle of the terahertz wave. We exploit this effect to record ~100 fs snapshot images of the structure of the orbital involved, and to reveal through pump-probe measurements coherent molecular vibrations at terahertz frequencies directly in the time domain and with sub-angstrom spatial resolution. We anticipate that the combination of lightwave electronics1–8 and atomic resolution of our approach will open the door to controlling electronic motion inside individual molecules at optical clock rates. PMID:27830788

Microscopy techniques in flavivirus research.

PubMed

Chong, Mun Keat; Chua, Anthony Jin Shun; Tan, Terence Tze Tong; Tan, Suat Hoon; Ng, Mah Lee

2014-04-01

The Flavivirus genus is composed of many medically important viruses that cause high morbidity and mortality, which include Dengue and West Nile viruses. Various molecular and biochemical techniques have been developed in the endeavour to study flaviviruses. However, microscopy techniques still have irreplaceable roles in the identification of novel virus pathogens and characterization of morphological changes in virus-infected cells. Fluorescence microscopy contributes greatly in understanding the fundamental viral protein localizations and virus-host protein interactions during infection. Electron microscopy remains the gold standard for visualizing ultra-structural features of virus particles and infected cells. New imaging techniques and combinatory applications are continuously being developed to push the limit of resolution and extract more quantitative data. Currently, correlative live cell imaging and high resolution three-dimensional imaging have already been achieved through the tandem use of optical and electron microscopy in analyzing biological specimens. Microscopy techniques are also used to measure protein binding affinities and determine the mobility pattern of proteins in cells. This chapter will consolidate on the applications of various well-established microscopy techniques in flavivirus research, and discuss how recently developed microscopy techniques can potentially help advance our understanding in these membrane viruses. Copyright © 2013 Elsevier Ltd. All rights reserved.

Aberration corrected STEM by means of diffraction gratings

DOE Office of Scientific and Technical Information (OSTI.GOV)

Linck, Martin; Ercius, Peter A.; Pierce, Jordan S.

In the past 15 years, the advent of aberration correction technology in electron microscopy has enabled materials analysis on the atomic scale. This is made possible by precise arrangements of multipole electrodes and magnetic solenoids to compensate the aberrations inherent to any focusing element of an electron microscope. In this paper, we describe an alternative method to correct for the spherical aberration of the objective lens in scanning transmission electron microscopy (STEM) using a passive, nanofabricated diffractive optical element. This holographic device is installed in the probe forming aperture of a conventional electron microscope and can be designed to removemore » arbitrarily complex aberrations from the electron's wave front. In this work, we show a proof-of-principle experiment that demonstrates successful correction of the spherical aberration in STEM by means of such a grating corrector (GCOR). Our GCOR enables us to record aberration-corrected high-resolution high-angle annular dark field (HAADF-) STEM images, although yet without advancement in probe current and resolution. Finally, improvements in this technology could provide an economical solution for aberration-corrected high-resolution STEM in certain use scenarios.« less

Aberration corrected STEM by means of diffraction gratings

DOE PAGES

Linck, Martin; Ercius, Peter A.; Pierce, Jordan S.; ...

2017-06-12

In the past 15 years, the advent of aberration correction technology in electron microscopy has enabled materials analysis on the atomic scale. This is made possible by precise arrangements of multipole electrodes and magnetic solenoids to compensate the aberrations inherent to any focusing element of an electron microscope. In this paper, we describe an alternative method to correct for the spherical aberration of the objective lens in scanning transmission electron microscopy (STEM) using a passive, nanofabricated diffractive optical element. This holographic device is installed in the probe forming aperture of a conventional electron microscope and can be designed to removemore » arbitrarily complex aberrations from the electron's wave front. In this work, we show a proof-of-principle experiment that demonstrates successful correction of the spherical aberration in STEM by means of such a grating corrector (GCOR). Our GCOR enables us to record aberration-corrected high-resolution high-angle annular dark field (HAADF-) STEM images, although yet without advancement in probe current and resolution. Finally, improvements in this technology could provide an economical solution for aberration-corrected high-resolution STEM in certain use scenarios.« less

Correlative 3D imaging of Whole Mammalian Cells with Light and Electron Microscopy

PubMed Central

Murphy, Gavin E.; Narayan, Kedar; Lowekamp, Bradley C.; Hartnell, Lisa M.; Heymann, Jurgen A. W.; Fu, Jing; Subramaniam, Sriram

2011-01-01

We report methodological advances that extend the current capabilities of ion-abrasion scanning electron microscopy (IA–SEM), also known as focused ion beam scanning electron microscopy, a newly emerging technology for high resolution imaging of large biological specimens in 3D. We establish protocols that enable the routine generation of 3D image stacks of entire plastic-embedded mammalian cells by IA-SEM at resolutions of ~10 to 20 nm at high contrast and with minimal artifacts from the focused ion beam. We build on these advances by describing a detailed approach for carrying out correlative live confocal microscopy and IA–SEM on the same cells. Finally, we demonstrate that by combining correlative imaging with newly developed tools for automated image processing, small 100 nm-sized entities such as HIV-1 or gold beads can be localized in SEM image stacks of whole mammalian cells. We anticipate that these methods will add to the arsenal of tools available for investigating mechanisms underlying host-pathogen interactions, and more generally, the 3D subcellular architecture of mammalian cells and tissues. PMID:21907806

Scanning transmission electron microscopy: Albert Crewe's vision and beyond.

PubMed

Krivanek, Ondrej L; Chisholm, Matthew F; Murfitt, Matthew F; Dellby, Niklas

2012-12-01

Some four decades were needed to catch up with the vision that Albert Crewe and his group had for the scanning transmission electron microscope (STEM) in the nineteen sixties and seventies: attaining 0.5Å resolution, and identifying single atoms spectroscopically. With these goals now attained, STEM developments are turning toward new directions, such as rapid atomic resolution imaging and exploring atomic bonding and electronic properties of samples at atomic resolution. The accomplishments and the future challenges are reviewed and illustrated with practical examples. Copyright © 2012 Elsevier B.V. All rights reserved.

Characterization of conductive nanobiomaterials derived from viral assemblies by low-voltage STEM imaging and Raman scattering

NASA Astrophysics Data System (ADS)

Plascencia-Villa, Germán; Carreño-Fuentes, Liliana; Bahena, Daniel; José-Yacamán, Miguel; Palomares, Laura A.; Ramírez, Octavio T.

2014-09-01

New technologies require the development of novel nanomaterials that need to be fully characterized to achieve their potential. High-resolution low-voltage scanning transmission electron microscopy (STEM) has proven to be a very powerful technique in nanotechnology, but its use for the characterization of nanobiomaterials has been limited. Rotavirus VP6 self-assembles into nanotubular assemblies that possess an intrinsic affinity for Au ions. This property was exploited to produce hybrid nanobiomaterials by the in situ functionalization of recombinant VP6 nanotubes with gold nanoparticles. In this work, Raman spectroscopy and advanced analytical electron microscopy imaging with spherical aberration-corrected (Cs) STEM and nanodiffraction at low-voltage doses were employed to characterize nanobiomaterials. STEM imaging revealed the precise structure and arrangement of the protein templates, as well as the nanostructure and atomic arrangement of gold nanoparticles with high spatial sub-Angstrom resolution and avoided radiation damage. The imaging was coupled with backscattered electron imaging, ultra-high resolution scanning electron microscopy and x-ray spectroscopy. The hybrid nanobiomaterials that were obtained showed unique properties as bioelectronic conductive devices and showed enhanced Raman scattering by their precise arrangement into superlattices, displaying the utility of viral assemblies as functional integrative self-assembled nanomaterials for novel applications.

Coupling EELS/EFTEM Imaging with Environmental Fluid Cell Microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Unocic, Raymond R; Baggetto, Loic; Veith, Gabriel M

2012-01-01

Insight into dynamically evolving electrochemical reactions and mechanisms encountered in electrical energy storage (EES) and conversion technologies (batteries, fuel cells, and supercapacitors), materials science (corrosion and oxidation), and materials synthesis (electrodeposition) remains limited due to the present lack of in situ high-resolution characterization methodologies. Electrochemical fluid cell microscopy is an emerging in-situ method that allows for the direct, real-time imaging of electrochemical processes within a fluid environment. This technique is facilitated by the use of MEMS-based biasing microchip platforms that serve the purpose of sealing the highly volatile electrolyte between two electron transparent SiNx membranes and interfacing electrodes to anmore » external potentiostat for controlled nanoscale electrochemislly experiments [!]. In order to elucidate both stmctural and chemical changes during such in situ electrochemical experiments, it is impmtant to first improve upon the spatial resolution by utilizing energy-filtered transmission electron microscopy (EFTEM) (to minimize chromatic aben ation), then to detennine the chemical changes via electron energy loss spectroscopy (EELS). This presents a formidable challenge since the overall thickness through which electrons are scattered through the multiple layers of the cell can be on the order of hundreds of nanometers to microns, scattering through which has the deleterious effect of degrading image resolution and decreasing signal-to noise for spectroscopy [2].« less

Advantages of intermediate X-ray energies in Zernike phase contrast X-ray microscopy.

PubMed

Wang, Zhili; Gao, Kun; Chen, Jian; Hong, Youli; Ge, Xin; Wang, Dajiang; Pan, Zhiyun; Zhu, Peiping; Yun, Wenbing; Jacobsen, Chris; Wu, Ziyu

2013-01-01

Understanding the hierarchical organizations of molecules and organelles within the interior of large eukaryotic cells is a challenge of fundamental interest in cell biology. Light microscopy is a powerful tool for observations of the dynamics of live cells, its resolution attainable is limited and insufficient. While electron microscopy can produce images with astonishing resolution and clarity of ultra-thin (<1 μm thick) sections of biological specimens, many questions involve the three-dimensional organization of a cell or the interconnectivity of cells. X-ray microscopy offers superior imaging resolution compared to light microscopy, and unique capability of nondestructive three-dimensional imaging of hydrated unstained biological cells, complementary to existing light and electron microscopy. Until now, X-ray microscopes operating in the "water window" energy range between carbon and oxygen k-shell absorption edges have produced outstanding 3D images of cryo-preserved cells. The relatively low X-ray energy (<540 eV) of the water window imposes two important limitations: limited penetration (<10 μm) not suitable for imaging larger cells or tissues, and small depth of focus (DoF) for high resolution 3D imaging (e.g., ~1 μm DoF for 20 nm resolution). An X-ray microscope operating at intermediate energy around 2.5 keV using Zernike phase contrast can overcome the above limitations and reduces radiation dose to the specimen. Using a hydrated model cell with an average chemical composition reported in literature, we calculated the image contrast and the radiation dose for absorption and Zernike phase contrast, respectively. The results show that an X-ray microscope operating at ~2.5 keV using Zernike phase contrast offers substantial advantages in terms of specimen size, radiation dose and depth-of-focus. Copyright © 2012 Elsevier Inc. All rights reserved.

Oxidation-state sensitive imaging of cerium dioxide by atomic-resolution low-angle annular dark field scanning transmission electron microscopy.

PubMed

Johnston-Peck, Aaron C; Winterstein, Jonathan P; Roberts, Alan D; DuChene, Joseph S; Qian, Kun; Sweeny, Brendan C; Wei, Wei David; Sharma, Renu; Stach, Eric A; Herzing, Andrew A

2016-03-01

Low-angle annular dark field (LAADF) scanning transmission electron microscopy (STEM) imaging is presented as a method that is sensitive to the oxidation state of cerium ions in CeO2 nanoparticles. This relationship was validated through electron energy loss spectroscopy (EELS), in situ measurements, as well as multislice image simulations. Static displacements caused by the increased ionic radius of Ce(3+) influence the electron channeling process and increase electron scattering to low angles while reducing scatter to high angles. This process manifests itself by reducing the high-angle annular dark field (HAADF) signal intensity while increasing the LAADF signal intensity in close proximity to Ce(3+) ions. This technique can supplement STEM-EELS and in so doing, relax the experimental challenges associated with acquiring oxidation state information at high spatial resolutions. Published by Elsevier B.V.

FitEM2EM—Tools for Low Resolution Study of Macromolecular Assembly and Dynamics

PubMed Central

Frankenstein, Ziv; Sperling, Joseph; Sperling, Ruth; Eisenstein, Miriam

2008-01-01

Studies of the structure and dynamics of macromolecular assemblies often involve comparison of low resolution models obtained using different techniques such as electron microscopy or atomic force microscopy. We present new computational tools for comparing (matching) and docking of low resolution structures, based on shape complementarity. The matched or docked objects are represented by three dimensional grids where the value of each grid point depends on its position with regard to the interior, surface or exterior of the object. The grids are correlated using fast Fourier transformations producing either matches of related objects or docking models depending on the details of the grid representations. The procedures incorporate thickening and smoothing of the surfaces of the objects which effectively compensates for differences in the resolution of the matched/docked objects, circumventing the need for resolution modification. The presented matching tool FitEM2EMin successfully fitted electron microscopy structures obtained at different resolutions, different conformers of the same structure and partial structures, ranking correct matches at the top in every case. The differences between the grid representations of the matched objects can be used to study conformation differences or to characterize the size and shape of substructures. The presented low-to-low docking tool FitEM2EMout ranked the expected models at the top. PMID:18974836

In-line three-dimensional holography of nanocrystalline objects at atomic resolution

PubMed Central

Chen, F.-R.; Van Dyck, D.; Kisielowski, C.

2016-01-01

Resolution and sensitivity of the latest generation aberration-corrected transmission electron microscopes allow the vast majority of single atoms to be imaged with sub-Ångstrom resolution and their locations determined in an image plane with a precision that exceeds the 1.9-pm wavelength of 300 kV electrons. Such unprecedented performance allows expansion of electron microscopic investigations with atomic resolution into the third dimension. Here we report a general tomographic method to recover the three-dimensional shape of a crystalline particle from high-resolution images of a single projection without the need for sample rotation. The method is compatible with low dose rate electron microscopy, which improves on signal quality, while minimizing electron beam-induced structure modifications even for small particles or surfaces. We apply it to germanium, gold and magnesium oxide particles, and achieve a depth resolution of 1–2 Å, which is smaller than inter-atomic distances. PMID:26887849

Detection, Mapping, and Quantification of Single Walled Carbon Nanotubes in Histological Specimens with Photoacoustic Microscopy

PubMed Central

Mikos, Antonios G.; Jansen, John A.; Shroyer, Kenneth R.; Wang, Lihong V.; Sitharaman, Balaji

2012-01-01

Aims In the present study, the efficacy of multi-scale photoacoustic microscopy (PAM) was investigated to detect, map, and quantify trace amounts [nanograms (ng) to micrograms (µg)] of SWCNTs in a variety of histological tissue specimens consisting of cancer and benign tissue biopsies (histological specimens from implanted tissue engineering scaffolds). Materials and Methods Optical-resolution (OR) and acoustic-resolution (AR) - Photoacoustic microscopy (PAM) was employed to detect, map and quantify the SWCNTs in a variety of tissue histological specimens and compared with other optical techniques (bright-field optical microscopy, Raman microscopy, near infrared (NIR) fluorescence microscopy). Results Both optical-resolution and acoustic-resolution PAM, allow the detection and quantification of SWCNTs in histological specimens with scalable spatial resolution and depth penetration. The noise-equivalent detection sensitivity to SWCNTs in the specimens was calculated to be as low as ∼7 pg. Image processing analysis further allowed the mapping, distribution, and quantification of the SWCNTs in the histological sections. Conclusions The results demonstrate the potential of PAM as a promising imaging technique to detect, map, and quantify SWCNTs in histological specimens, and could complement the capabilities of current optical and electron microscopy techniques in the analysis of histological specimens containing SWCNTs. PMID:22496892

Detailed Investigation of Core-Shell Precipitates in a Cu-Containing High Entropy Alloy

NASA Astrophysics Data System (ADS)

Alam, T.; Gwalani, B.; Viswanathan, G.; Fraser, H.; Banerjee, R.

2018-05-01

Due to the competing influences of configurational entropy and enthalpy of mixing, in recent years, secondary (including intermetallic) phases have been reported in many high entropy alloy (HEA) systems. These secondary phases offer great potential in terms of strengthening the HEA beyond the solid solution strengthening effects, and as such are of great interest in regards to alloy design for engineering applications. The present research investigates novel nano-scale core-shell precipitates forming within the disordered bcc matrix phase of an Al2CrCuFeNi2 HEA, utilizing complementary high-resolution microscopy techniques of atom probe tomography (APT) and transmission electron microscopy (TEM). The size, morphology, and local chemistry of these core-shell precipitates was measured by APT, and the composition was further corroborated by high-resolution scanning transmission electron microscopy-energy dispersive spectroscopy in an aberration-corrected TEM. Furthermore, high-resolution TEM imaging of the core-shell structure indicates that the Cu-rich core exhibits a bcc crystal structure.

Characterisation of phases in nanostructured, multilayered titanium alloys by analytical and high-resolution electron microscopy.

PubMed

Czyrska-Filemonowicz, A; Buffat, P A

2009-01-01

Surface processing of a Ti-6Al-4V alloy led to a complex multilayered microstructure containing several phases of the Ni-Ti-P-Al-O system, which improves the mechanical and tribological surface properties. The microstructure, chemical and phase compositions of the hard layer formed on the surface were investigated by LM, XRD, SEM as well as analytical/high-resolution TEM, STEM, EDS, electron diffraction and FIB. Phase identification based on electron diffraction, HRTEM and EDS microanalysis revealed the presence of several binary and ternary phases in the system Ti-Ni-P, sometimes with partial substitution of Ti by Al. However some phases, mainly nanoparticles, still remain not identified satisfactorily. Electron microscopy techniques used for identification of phases present in surface multilayers and some practical limits to their routine application are reminded here.

Femtosecond few- to single-electron point-projection microscopy for nanoscale dynamic imaging

PubMed Central

Bainbridge, A. R.; Barlow Myers, C. W.; Bryan, W. A.

2016-01-01

Femtosecond electron microscopy produces real-space images of matter in a series of ultrafast snapshots. Pulses of electrons self-disperse under space-charge broadening, so without compression, the ideal operation mode is a single electron per pulse. Here, we demonstrate femtosecond single-electron point projection microscopy (fs-ePPM) in a laser-pump fs-e-probe configuration. The electrons have an energy of only 150 eV and take tens of picoseconds to propagate to the object under study. Nonetheless, we achieve a temporal resolution with a standard deviation of 114 fs (equivalent to a full-width at half-maximum of 269 ± 40 fs) combined with a spatial resolution of 100 nm, applied to a localized region of charge at the apex of a nanoscale metal tip induced by 30 fs 800 nm laser pulses at 50 kHz. These observations demonstrate real-space imaging of reversible processes, such as tracking charge distributions, is feasible whilst maintaining femtosecond resolution. Our findings could find application as a characterization method, which, depending on geometry, could resolve tens of femtoseconds and tens of nanometres. Dynamically imaging electric and magnetic fields and charge distributions on sub-micron length scales opens new avenues of ultrafast dynamics. Furthermore, through the use of active compression, such pulses are an ideal seed for few-femtosecond to attosecond imaging applications which will access sub-optical cycle processes in nanoplasmonics. PMID:27158637

The Effect of Electron Beam Irradiation in Environmental Scanning Transmission Electron Microscopy of Whole Cells in Liquid.

PubMed

Hermannsdörfer, Justus; Tinnemann, Verena; Peckys, Diana B; de Jonge, Niels

2016-06-01

Whole cells can be studied in their native liquid environment using electron microscopy, and unique information about the locations and stoichiometry of individual membrane proteins can be obtained from many cells thus taking cell heterogeneity into account. Of key importance for the further development of this microscopy technology is knowledge about the effect of electron beam radiation on the samples under investigation. We used environmental scanning electron microscopy (ESEM) with scanning transmission electron microscopy (STEM) detection to examine the effect of radiation for whole fixed COS7 fibroblasts in liquid. The main observation was the localization of nanoparticle labels attached to epidermal growth factor receptors (EGFRs). It was found that the relative distances between the labels remained mostly unchanged (<1.5%) for electron doses ranging from the undamaged native state at 10 e-/Å2 toward 103 e-/Å2. This dose range was sufficient to determine the EGFR locations with nanometer resolution and to distinguish between monomers and dimers. Various different forms of radiation damage became visible at higher doses, including severe dislocation, and the dissolution of labels.

Visualising reacting single atoms under controlled conditions: Advances in atomic resolution in situ Environmental (Scanning) Transmission Electron Microscopy (E(S)TEM)

NASA Astrophysics Data System (ADS)

Boyes, Edward D.; Gai, Pratibha L.

2014-02-01

Advances in atomic resolution Environmental (Scanning) Transmission Electron Microscopy (E(S)TEM) for probing gas-solid catalyst reactions in situ at the atomic level under controlled reaction conditions of gas environment and temperature are described. The recent development of the ESTEM extends the capability of the ETEM by providing the direct visualisation of single atoms and the atomic structure of selected solid state heterogeneous catalysts in their working states in real-time. Atomic resolution E(S)TEM provides a deeper understanding of the dynamic atomic processes at the surface of solids and their mechanisms of operation. The benefits of atomic resolution-E(S)TEM to science and technology include new knowledge leading to improved technological processes with substantial economic benefits, improved healthcare, reductions in energy needs and the management of environmental waste generation. xml:lang="fr"

Computational methods for constructing protein structure models from 3D electron microscopy maps.

PubMed

Esquivel-Rodríguez, Juan; Kihara, Daisuke

2013-10-01

Protein structure determination by cryo-electron microscopy (EM) has made significant progress in the past decades. Resolutions of EM maps have been improving as evidenced by recently reported structures that are solved at high resolutions close to 3Å. Computational methods play a key role in interpreting EM data. Among many computational procedures applied to an EM map to obtain protein structure information, in this article we focus on reviewing computational methods that model protein three-dimensional (3D) structures from a 3D EM density map that is constructed from two-dimensional (2D) maps. The computational methods we discuss range from de novo methods, which identify structural elements in an EM map, to structure fitting methods, where known high resolution structures are fit into a low-resolution EM map. A list of available computational tools is also provided. Copyright © 2013 Elsevier Inc. All rights reserved.

The extraction of gold nanoparticles from oat and wheat biomasses using sodium citrate and cetyltrimethylammonium bromide, studied by x-ray absorption spectroscopy, high-resolution transmission electron microscopy, and UV-visible spectroscopy.

PubMed

Armendariz, Veronica; Parsons, Jason G; Lopez, Martha L; Peralta-Videa, Jose R; Jose-Yacaman, Miguel; Gardea-Torresdey, Jorge L

2009-03-11

Gold (Au) nanoparticles can be produced through the interaction of Au(III) ions with oat and wheat biomasses. This paper describes a procedure to recover gold nanoparticles from oat and wheat biomasses using cetyltrimethylammonium bromide or sodium citrate. Extracts were analyzed using UV-visible spectroscopy, high-resolution transmission electron microscopy (HRTEM), and x-ray absorption spectroscopy. The HRTEM data demonstrated that smaller nanoparticles are extracted first, followed by larger nanoparticles. In the fourth extraction, coating of chelating agents is visible on the extracted nanoparticles.

High-resolution mapping of molecules in an ionic liquid via scanning transmission electron microscopy.

PubMed

Miyata, Tomohiro; Mizoguchi, Teruyasu

2018-03-01

Understanding structures and spatial distributions of molecules in liquid phases is crucial for the control of liquid properties and to develop efficient liquid-phase processes. Here, real-space mapping of molecular distributions in a liquid was performed. Specifically, the ionic liquid 1-Ethyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide (C2mimTFSI) was imaged using atomic-resolution scanning transmission electron microscopy. Simulations revealed network-like bright regions in the images that were attributed to the TFSI- anion, with minimal contributions from the C2mim+ cation. Simple visualization of the TFSI- distribution in the liquid sample was achieved by binarizing the experimental image.

Correlative two-photon and serial block face scanning electron microscopy in neuronal tissue using 3D near-infrared branding maps.

PubMed

Lees, Robert M; Peddie, Christopher J; Collinson, Lucy M; Ashby, Michael C; Verkade, Paul

2017-01-01

Linking cellular structure and function has always been a key goal of microscopy, but obtaining high resolution spatial and temporal information from the same specimen is a fundamental challenge. Two-photon (2P) microscopy allows imaging deep inside intact tissue, bringing great insight into the structural and functional dynamics of cells in their physiological environment. At the nanoscale, the complex ultrastructure of a cell's environment in tissue can be reconstructed in three dimensions (3D) using serial block face scanning electron microscopy (SBF-SEM). This provides a snapshot of high resolution structural information pertaining to the shape, organization, and localization of multiple subcellular structures at the same time. The pairing of these two imaging modalities in the same specimen provides key information to relate cellular dynamics to the ultrastructural environment. Until recently, approaches to relocate a region of interest (ROI) in tissue from 2P microscopy for SBF-SEM have been inefficient or unreliable. However, near-infrared branding (NIRB) overcomes this by using the laser from a multiphoton microscope to create fiducial markers for accurate correlation of 2P and electron microscopy (EM) imaging volumes. The process is quick and can be user defined for each sample. Here, to increase the efficiency of ROI relocation, multiple NIRB marks are used in 3D to target ultramicrotomy. A workflow is described and discussed to obtain a data set for 3D correlated light and electron microscopy, using three different preparations of brain tissue as examples. Copyright © 2017 Elsevier Inc. All rights reserved.

Theory of the spatial resolution of (scanning) transmission electron microscopy in liquid water or ice layers.

PubMed

de Jonge, Niels

2018-04-01

The sample dependent spatial resolution was calculated for transmission electron microscopy (TEM) and scanning TEM (STEM) of objects (e.g., nanoparticles, proteins) embedded in a layer of liquid water or amorphous ice. The theoretical model includes elastic- and inelastic scattering, beam broadening, and chromatic aberration. Different contrast mechanisms were evaluated as function of the electron dose, the detection angle, and the sample configuration. It was found that the spatial resolution scales with the electron dose to the -1/4th power. Gold- and carbon nanoparticles were examined in the middle of water layers ranging from 0.01--10 µm thickness representing relevant classes of experiments in both materials science and biology. The optimal microscope settings differ between experimental configurations. STEM performs the best for gold nanoparticles for all layer thicknesses, while carbon is best imaged with phase-contrast TEM for thin layers but bright field STEM is preferred for thicker layers. The resolution was also calculated for a water layer enclosed between thin membranes. The influence of chromatic aberration correction for TEM was examined as well. The theory is broadly applicable to other types of materials and sample configurations. Copyright © 2018 Elsevier B.V. All rights reserved.

Three-dimensional scanning transmission electron microscopy of biological specimens.

PubMed

de Jonge, Niels; Sougrat, Rachid; Northan, Brian M; Pennycook, Stephen J

2010-02-01

A three-dimensional (3D) reconstruction of the cytoskeleton and a clathrin-coated pit in mammalian cells has been achieved from a focal-series of images recorded in an aberration-corrected scanning transmission electron microscope (STEM). The specimen was a metallic replica of the biological structure comprising Pt nanoparticles 2-3 nm in diameter, with a high stability under electron beam radiation. The 3D dataset was processed by an automated deconvolution procedure. The lateral resolution was 1.1 nm, set by pixel size. Particles differing by only 10 nm in vertical position were identified as separate objects with greater than 20% dip in contrast between them. We refer to this value as the axial resolution of the deconvolution or reconstruction, the ability to recognize two objects, which were unresolved in the original dataset. The resolution of the reconstruction is comparable to that achieved by tilt-series transmission electron microscopy. However, the focal-series method does not require mechanical tilting and is therefore much faster. 3D STEM images were also recorded of the Golgi ribbon in conventional thin sections containing 3T3 cells with a comparable axial resolution in the deconvolved dataset.

Three-Dimensional Scanning Transmission Electron Microscopy of Biological Specimens

PubMed Central

de Jonge, Niels; Sougrat, Rachid; Northan, Brian M.; Pennycook, Stephen J.

2010-01-01

A three-dimensional (3D) reconstruction of the cytoskeleton and a clathrin-coated pit in mammalian cells has been achieved from a focal-series of images recorded in an aberration-corrected scanning transmission electron microscope (STEM). The specimen was a metallic replica of the biological structure comprising Pt nanoparticles 2–3 nm in diameter, with a high stability under electron beam radiation. The 3D dataset was processed by an automated deconvolution procedure. The lateral resolution was 1.1 nm, set by pixel size. Particles differing by only 10 nm in vertical position were identified as separate objects with greater than 20% dip in contrast between them. We refer to this value as the axial resolution of the deconvolution or reconstruction, the ability to recognize two objects, which were unresolved in the original dataset. The resolution of the reconstruction is comparable to that achieved by tilt-series transmission electron microscopy. However, the focal-series method does not require mechanical tilting and is therefore much faster. 3D STEM images were also recorded of the Golgi ribbon in conventional thin sections containing 3T3 cells with a comparable axial resolution in the deconvolved dataset. PMID:20082729

Optimizing low-light microscopy with back-illuminated electron multiplying charge-coupled device: enhanced sensitivity, speed, and resolution.

PubMed

Coates, Colin G; Denvir, Donal J; McHale, Noel G; Thornbury, Keith D; Hollywood, Mark A

2004-01-01

The back-illuminated electron multiplying charge-coupled device (EMCCD) camera is having a profound influence on the field of low-light dynamic cellular microscopy, combining highest possible photon collection efficiency with the ability to virtually eliminate the readout noise detection limit. We report here the use of this camera, in 512 x 512 frame-transfer chip format at 10-MHz pixel readout speed, in optimizing a demanding ultra-low-light intracellular calcium flux microscopy setup. The arrangement employed includes a spinning confocal Nipkow disk, which, while facilitating the need to both generate images at very rapid frame rates and minimize background photons, yields very weak signals. The challenge for the camera lies not just in detecting as many of these scarce photons as possible, but also in operating at a frame rate that meets the temporal resolution requirements of many low-light microscopy approaches, a particular demand of smooth muscle calcium flux microscopy. Results presented illustrate both the significant sensitivity improvement offered by this technology over the previous standard in ultra-low-light CCD detection, the GenIII+intensified charge-coupled device (ICCD), and also portray the advanced temporal and spatial resolution capabilities of the EMCCD. Copyright 2004 Society of Photo-Optical Instrumentation Engineers.

Development of Scanning Ultrafast Electron Microscope Capability.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Collins, Kimberlee Chiyoko; Talin, Albert Alec; Chandler, David W.

Modern semiconductor devices rely on the transport of minority charge carriers. Direct examination of minority carrier lifetimes in real devices with nanometer-scale features requires a measurement method with simultaneously high spatial and temporal resolutions. Achieving nanometer spatial resolutions at sub-nanosecond temporal resolution is possible with pump-probe methods that utilize electrons as probes. Recently, a stroboscopic scanning electron microscope was developed at Caltech, and used to study carrier transport across a Si p-n junction [ 1 , 2 , 3 ] . In this report, we detail our development of a prototype scanning ultrafast electron microscope system at Sandia National Laboratoriesmore » based on the original Caltech design. This effort represents Sandia's first exploration into ultrafast electron microscopy.« less

The role of gas in determining image quality and resolution during in situ scanning transmission electron microscopy experiments

DOE PAGES

Zhu, Yuanyuan; Browning, Nigel D.

2017-05-24

As gas-solid heterogeneous catalytic reactions are molecular in nature, a full mechanistic understanding of the process requires atomic scale characterization under realistic operating conditions. While atomic resolution imaging has become a routine in modern high-vacuum (scanning) transmission electron microscopy ((S)TEM), both image quality and resolution nominally degrade when reaction gases are introduced. In this work, we systematically assess the effects of different gases at various pressures on the quality and resolution of images obtained at room temperature in the annular dark field STEM imaging mode using a differentially pumped (DP) gas cell. This imaging mode is largely free from inelasticmore » scattering effects induced by the presence of gases and retains good imaging properties over a wide range of gas mass/pressures. Furthermore, we demonstrate the application of the ESTEM with atomic resolution images of a complex oxide alkane oxidation catalyst MoVNbTeOx (M1) immersed in light and heavy gas environments.« less

A Global Approach for Quantitative Super Resolution and Electron Microscopy on Cryo and Epoxy Sections Using Self-labeling Protein Tags.

PubMed

Müller, Andreas; Neukam, Martin; Ivanova, Anna; Sönmez, Anke; Münster, Carla; Kretschmar, Susanne; Kalaidzidis, Yannis; Kurth, Thomas; Verbavatz, Jean-Marc; Solimena, Michele

2017-02-02

Correlative light and electron microscopy (CLEM) is a powerful approach to investigate the molecular ultrastructure of labeled cell compartments. However, quantitative CLEM studies are rare, mainly due to small sample sizes and the sensitivity of fluorescent proteins to strong fixatives and contrasting reagents for EM. Here, we show that fusion of a self-labeling protein to insulin allows for the quantification of age-distinct insulin granule pools in pancreatic beta cells by a combination of super resolution and transmission electron microscopy on Tokuyasu cryosections. In contrast to fluorescent proteins like GFP organic dyes covalently bound to self-labeling proteins retain their fluorescence also in epoxy resin following high pressure freezing and freeze substitution, or remarkably even after strong chemical fixation. This enables for the assessment of age-defined granule morphology and degradation. Finally, we demonstrate that this CLEM protocol is highly versatile, being suitable for single and dual fluorescent labeling and detection of different proteins with optimal ultrastructure preservation and contrast.

New modes of electron microscopy for materials science enabled by fast direct electron detectors

NASA Astrophysics Data System (ADS)

Minor, Andrew

There is an ongoing revolution in the development of electron detector technology that has enabled modes of electron microscopy imaging that had only before been theorized. The age of electron microscopy as a tool for imaging is quickly giving way to a new frontier of multidimensional datasets to be mined. These improvements in electron detection have enabled cryo-electron microscopy to resolve the three-dimensional structures of non-crystalized proteins, revolutionizing structural biology. In the physical sciences direct electron detectors has enabled four-dimensional reciprocal space maps of materials at atomic resolution, providing all the structural information about nanoscale materials in one experiment. This talk will highlight the impact of direct electron detectors for materials science, including a new method of scanning nanobeam diffraction. With faster detectors we can take a series of 2D diffraction patterns at each position in a 2D STEM raster scan resulting in a four-dimensional data set. For thin film analysis, direct electron detectors hold the potential to enable strain, polarization, composition and electrical field mapping over relatively large fields of view, all from a single experiment.

The Electron Microscopy Outreach Program: A Web-based resource for research and education.

PubMed

Sosinsky, G E; Baker, T S; Hand, G; Ellisman, M H

1999-01-01

We have developed a centralized World Wide Web (WWW)-based environment that serves as a resource of software tools and expertise for biological electron microscopy. A major focus is molecular electron microscopy, but the site also includes information and links on structural biology at all levels of resolution. This site serves to help integrate or link structural biology techniques in accordance with user needs. The WWW site, called the Electron Microscopy (EM) Outreach Program (URL: http://emoutreach.sdsc.edu), provides scientists with computational and educational tools for their research and edification. In particular, we have set up a centralized resource containing course notes, references, and links to image analysis and three-dimensional reconstruction software for investigators wanting to learn about EM techniques either within or outside of their fields of expertise. Copyright 1999 Academic Press.

Swift heavy ion track formation in Gd2Zr2-xTixO7 pyrochlore: Effect of electronic energy loss

NASA Astrophysics Data System (ADS)

Lang, Maik; Toulemonde, Marcel; Zhang, Jiaming; Zhang, Fuxiang; Tracy, Cameron L.; Lian, Jie; Wang, Zhongwu; Weber, William J.; Severin, Daniel; Bender, Markus; Trautmann, Christina; Ewing, Rodney C.

2014-10-01

The morphology of swift heavy ion tracks in the Gd2Zr2-xTixO7 pyrochlore system has been investigated as a function of the variation in chemical composition and electronic energy loss, dE/dx, over a range of energetic ions: 58Ni, 101Ru, 129Xe, 181Ta, 197Au, 208Pb, and 238U of 11.1 MeV/u specific energy. Bright-field transmission electron microscopy, synchrotron X-ray diffraction, and Raman spectroscopy reveal an increasing degree of amorphization with increasing Ti-content and dE/dx. The size and morphology of individual ion tracks in Gd2Ti2O7 were characterized by high-resolution transmission electron microscopy revealing a core-shell structure with an outer defect-fluorite dominated shell at low dE/dx to predominantly amorphous tracks at high dE/dx. Inelastic thermal-spike calculations have been used together with atomic-scale characterization of ion tracks in Gd2Ti2O7 by high resolution transmission electron microscopy to deduce critical energy densities for the complex core-shell morphologies induced by ions of different dE/dx.

Live Bacterial Physiology Visualized with 5 nm Resolution Using Scanning Transmission Electron Microscopy.

PubMed

Kennedy, Eamonn; Nelson, Edward M; Tanaka, Tetsuya; Damiano, John; Timp, Gregory

2016-02-23

It is now possible to visualize at nanometer resolution the infection of a living biological cell with virus without compromising cell viability using scanning transmission electron microscopy (STEM). To provide contrast while preserving viability, Escherichia coli and P1 bacteriophages were first positively stained with a very low concentration of uranyl acetate in minimal phosphate medium and then imaged with low-dose STEM in a microfluidic liquid flow cell. Under these conditions, it was established that the median lethal dose of electrons required to kill half the tested population was LD50 = 30 e(-)/nm(2), which coincides with the disruption of a wet biological membrane, according to prior reports. Consistent with the lateral resolution and high-contrast signal-to-noise ratio (SNR) inferred from Monte Carlo simulations, images of the E. coli membrane, flagella, and the bacteriophages were acquired with 5 nm resolution, but the cumulative dose exceeded LD50. On the other hand, with a cumulative dose below LD50 (and lower SNR), it was still possible to visualize the infection of E. coli by P1, showing the insertion of viral DNA within 3 s, with 5 nm resolution.

Surface determination through atomically resolved secondary-electron imaging

PubMed Central

Ciston, J.; Brown, H. G.; D'Alfonso, A. J.; Koirala, P.; Ophus, C.; Lin, Y.; Suzuki, Y.; Inada, H.; Zhu, Y.; Allen, L. J.; Marks, L. D.

2015-01-01

Unique determination of the atomic structure of technologically relevant surfaces is often limited by both a need for homogeneous crystals and ambiguity of registration between the surface and bulk. Atomically resolved secondary-electron imaging is extremely sensitive to this registration and is compatible with faceted nanomaterials, but has not been previously utilized for surface structure determination. Here we report a detailed experimental atomic-resolution secondary-electron microscopy analysis of the c(6 × 2) reconstruction on strontium titanate (001) coupled with careful simulation of secondary-electron images, density functional theory calculations and surface monolayer-sensitive aberration-corrected plan-view high-resolution transmission electron microscopy. Our work reveals several unexpected findings, including an amended registry of the surface on the bulk and strontium atoms with unusual seven-fold coordination within a typically high surface coverage of square pyramidal TiO5 units. Dielectric screening is found to play a critical role in attenuating secondary-electron generation processes from valence orbitals. PMID:26082275

Electron-beam broadening in amorphous carbon films in low-energy scanning transmission electron microscopy.

PubMed

Drees, H; Müller, E; Dries, M; Gerthsen, D

2018-02-01

Resolution in scanning transmission electron microscopy (STEM) is ultimately limited by the diameter of the electron beam. The electron beam diameter is not only determined by the properties of the condenser lens system but also by electron scattering in the specimen which leads to electron-beam broadening and degradation of the resolution with increasing specimen thickness. In this work we introduce a new method to measure electron-beam broadening which is based on STEM imaging with a multi-segmented STEM detector. We focus on STEM at low electron energies between 10 and 30 keV and use an amorphous carbon film with known thickness as test object. The experimental results are compared with calculated beam diameters using different analytical models and Monte-Carlo simulations. We find excellent agreement of the experimental data with the recently published model by Gauvin and Rudinsky [1] for small t/λ el (thickness to elastic mean free path) values which are considered in our study. Copyright © 2017 Elsevier B.V. All rights reserved.

Surface determination through atomically resolved secondary-electron imaging

DOE PAGES

Ciston, J.; Brown, H. G.; D’Alfonso, A. J.; ...

2015-06-17

We report that unique determination of the atomic structure of technologically relevant surfaces is often limited by both a need for homogeneous crystals and ambiguity of registration between the surface and bulk. Atomically resolved secondary-electron imaging is extremely sensitive to this registration and is compatible with faceted nanomaterials, but has not been previously utilized for surface structure determination. Here we show a detailed experimental atomic-resolution secondary-electron microscopy analysis of the c(6 x 2) reconstruction on strontium titanate (001) coupled with careful simulation of secondary-electron images, density functional theory calculations and surface monolayer-sensitive aberration-corrected plan-view high-resolution transmission electron microscopy. Our workmore » reveals several unexpected findings, including an amended registry of the surface on the bulk and strontium atoms with unusual seven-fold coordination within a typically high surface coverage of square pyramidal TiO 5 units. Lastly, dielectric screening is found to play a critical role in attenuating secondary-electron generation processes from valence orbitals.« less

3D structure of individual nanocrystals in solution by electron microscopy

NASA Astrophysics Data System (ADS)

Park, Jungwon; Elmlund, Hans; Ercius, Peter; Yuk, Jong Min; Limmer, David T.; Chen, Qian; Kim, Kwanpyo; Han, Sang Hoon; Weitz, David A.; Zettl, A.; Alivisatos, A. Paul

2015-07-01

Knowledge about the synthesis, growth mechanisms, and physical properties of colloidal nanoparticles has been limited by technical impediments. We introduce a method for determining three-dimensional (3D) structures of individual nanoparticles in solution. We combine a graphene liquid cell, high-resolution transmission electron microscopy, a direct electron detector, and an algorithm for single-particle 3D reconstruction originally developed for analysis of biological molecules. This method yielded two 3D structures of individual platinum nanocrystals at near-atomic resolution. Because our method derives the 3D structure from images of individual nanoparticles rotating freely in solution, it enables the analysis of heterogeneous populations of potentially unordered nanoparticles that are synthesized in solution, thereby providing a means to understand the structure and stability of defects at the nanoscale.

An electromechanical material testing system for in situ electron microscopy and applications.

PubMed

Zhu, Yong; Espinosa, Horacio D

2005-10-11

We report the development of a material testing system for in situ electron microscopy (EM) mechanical testing of nanostructures. The testing system consists of an actuator and a load sensor fabricated by means of surface micromachining. This previously undescribed nanoscale material testing system makes possible continuous observation of the specimen deformation and failure with subnanometer resolution, while simultaneously measuring the applied load electronically with nanonewton resolution. This achievement was made possible by the integration of electromechanical and thermomechanical components based on microelectromechanical system technology. The system capabilities are demonstrated by the in situ EM testing of free-standing polysilicon films, metallic nanowires, and carbon nanotubes. In particular, a previously undescribed real-time instrumented in situ transmission EM observation of carbon nanotubes failure under tensile load is presented here.

Accurate Nanoscale Crystallography in Real-Space Using Scanning Transmission Electron Microscopy.

PubMed

Dycus, J Houston; Harris, Joshua S; Sang, Xiahan; Fancher, Chris M; Findlay, Scott D; Oni, Adedapo A; Chan, Tsung-Ta E; Koch, Carl C; Jones, Jacob L; Allen, Leslie J; Irving, Douglas L; LeBeau, James M

2015-08-01

Here, we report reproducible and accurate measurement of crystallographic parameters using scanning transmission electron microscopy. This is made possible by removing drift and residual scan distortion. We demonstrate real-space lattice parameter measurements with <0.1% error for complex-layered chalcogenides Bi2Te3, Bi2Se3, and a Bi2Te2.7Se0.3 nanostructured alloy. Pairing the technique with atomic resolution spectroscopy, we connect local structure with chemistry and bonding. Combining these results with density functional theory, we show that the incorporation of Se into Bi2Te3 causes charge redistribution that anomalously increases the van der Waals gap between building blocks of the layered structure. The results show that atomic resolution imaging with electrons can accurately and robustly quantify crystallography at the nanoscale.

Challenges of microtome‐based serial block‐face scanning electron microscopy in neuroscience

PubMed Central

WANNER, A. A.; KIRSCHMANN, M. A.

2015-01-01

Summary Serial block‐face scanning electron microscopy (SBEM) is becoming increasingly popular for a wide range of applications in many disciplines from biology to material sciences. This review focuses on applications for circuit reconstruction in neuroscience, which is one of the major driving forces advancing SBEM. Neuronal circuit reconstruction poses exceptional challenges to volume EM in terms of resolution, field of view, acquisition time and sample preparation. Mapping the connections between neurons in the brain is crucial for understanding information flow and information processing in the brain. However, information on the connectivity between hundreds or even thousands of neurons densely packed in neuronal microcircuits is still largely missing. Volume EM techniques such as serial section TEM, automated tape‐collecting ultramicrotome, focused ion‐beam scanning electron microscopy and SBEM (microtome serial block‐face scanning electron microscopy) are the techniques that provide sufficient resolution to resolve ultrastructural details such as synapses and provides sufficient field of view for dense reconstruction of neuronal circuits. While volume EM techniques are advancing, they are generating large data sets on the terabyte scale that require new image processing workflows and analysis tools. In this review, we present the recent advances in SBEM for circuit reconstruction in neuroscience and an overview of existing image processing and analysis pipelines. PMID:25907464

High-resolution imaging by scanning electron microscopy of semithin sections in correlation with light microscopy.

PubMed

Koga, Daisuke; Kusumi, Satoshi; Shodo, Ryusuke; Dan, Yukari; Ushiki, Tatsuo

2015-12-01

In this study, we introduce scanning electron microscopy (SEM) of semithin resin sections. In this technique, semithin sections were adhered on glass slides, stained with both uranyl acetate and lead citrate, and observed with a backscattered electron detector at a low accelerating voltage. As the specimens are stained in the same manner as conventional transmission electron microscopy (TEM), the contrast of SEM images of semithin sections was similar to TEM images of ultrathin sections. Using this technique, wide areas of semithin sections were also observed by SEM, without the obstruction of grids, which was inevitable for traditional TEM. This study also applied semithin section SEM to correlative light and electron microscopy. Correlative immunofluorescence microscopy and immune-SEM were performed in semithin sections of LR white resin-embedded specimens using a FluoroNanogold-labeled secondary antibody. Because LR white resin is hydrophilic and electron stable, this resin is suitable for immunostaining and SEM observation. Using correlative microscopy, the precise localization of the primary antibody was demonstrated by fluorescence microscopy and SEM. This method has great potential for studies examining the precise localization of molecules, including Golgi- and ER-associated proteins, in correlation with LM and SEM. © The Author 2015. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Scanning Transmission Electron Microscopy at High Resolution

PubMed Central

Wall, J.; Langmore, J.; Isaacson, M.; Crewe, A. V.

1974-01-01

We have shown that a scanning transmission electron microscope with a high brightness field emission source is capable of obtaining better than 3 Å resolution using 30 to 40 keV electrons. Elastic dark field images of single atoms of uranium and mercury are shown which demonstrate this fact as determined by a modified Rayleigh criterion. Point-to-point micrograph resolution between 2.5 and 3.0 Å is found in dark field images of micro-crystallites of uranium and thorium compounds. Furthermore, adequate contrast is available to observe single atoms as light as silver. Images PMID:4521050

Inducing fluorescence of uranyl acetate as a dual-purpose contrast agent for correlative light-electron microscopy with nanometre precision.

PubMed

Tuijtel, Maarten W; Mulder, Aat A; Posthuma, Clara C; van der Hoeven, Barbara; Koster, Abraham J; Bárcena, Montserrat; Faas, Frank G A; Sharp, Thomas H

2017-09-05

Correlative light-electron microscopy (CLEM) combines the high spatial resolution of transmission electron microscopy (TEM) with the capability of fluorescence light microscopy (FLM) to locate rare or transient cellular events within a large field of view. CLEM is therefore a powerful technique to study cellular processes. Aligning images derived from both imaging modalities is a prerequisite to correlate the two microscopy data sets, and poor alignment can limit interpretability of the data. Here, we describe how uranyl acetate, a commonly-used contrast agent for TEM, can be induced to fluoresce brightly at cryogenic temperatures (-195 °C) and imaged by cryoFLM using standard filter sets. This dual-purpose contrast agent can be used as a general tool for CLEM, whereby the equivalent staining allows direct correlation between fluorescence and TEM images. We demonstrate the potential of this approach by performing multi-colour CLEM of cells containing equine arteritis virus proteins tagged with either green- or red-fluorescent protein, and achieve high-precision localization of virus-induced intracellular membrane modifications. Using uranyl acetate as a dual-purpose contrast agent, we achieve an image alignment precision of ~30 nm, twice as accurate as when using fiducial beads, which will be essential for combining TEM with the evolving field of super-resolution light microscopy.

Microscopy with slow electrons: from LEEM to XPEEM

ScienceCinema

Bauer, Ernst [Arizona State University, Phoenix, Arizona, United States

2017-12-09

The short penetration and escape depth of electrons with energies below 1 keV make them ideally suited for the study of surfaces and ultrathin films. The combination of the low energy electrons and the high lateral resolution of a microscope produces a powerful method for the characterization of nanostructures on bulk samples, in particular if the microscope is equipped with an imaging energy filter and connected to a synchrotron radiation source. Comprehensive characterization by imaging, diffraction, and spectroscope of the structural, chemical, and magnetic properties is then possible. The Talk will describe the various imaging techniques in using reflected and emitted electrons in low-energy electron microscopy (LEEM) and x-ray photoemission electron microscopy (XPEEM), with an emphasis on magnetic materials with spin-polarized LEEM and x-ray magnetic circular dichroism PEEM. The talk with end with an outlook on future possibilities.

Direct observation of anti-phase boundaries in heteroepitaxy of GaSb thin films grown on Si(001) by transmission electron microscopy

NASA Astrophysics Data System (ADS)

Woo, S. Y.; Hosseini Vajargah, S.; Ghanad-Tavakoli, S.; Kleiman, R. N.; Botton, G. A.

2012-10-01

Unambiguous identification of anti-phase boundaries (APBs) in heteroepitaxial films of GaSb grown on Si has been so far elusive. In this work, we present conventional transmission electron microscopy (TEM) diffraction contrast imaging using superlattice reflections, in conjunction with convergent beam electron diffraction analysis, to determine a change in polarity across APBs in order to confirm the presence of anti-phase disorder. In-depth analysis of anti-phase disorder is further supported with atomic resolution high-angle annular dark-field scanning transmission electron microscopy. The nature of APBs in GaSb is further elucidated by a comparison to previous results for GaAs epilayers grown on Si.

Serial Section Scanning Electron Microscopy (S3EM) on Silicon Wafers for Ultra-Structural Volume Imaging of Cells and Tissues

PubMed Central

Horstmann, Heinz; Körber, Christoph; Sätzler, Kurt; Aydin, Daniel; Kuner, Thomas

2012-01-01

High resolution, three-dimensional (3D) representations of cellular ultrastructure are essential for structure function studies in all areas of cell biology. While limited subcellular volumes have been routinely examined using serial section transmission electron microscopy (ssTEM), complete ultrastructural reconstructions of large volumes, entire cells or even tissue are difficult to achieve using ssTEM. Here, we introduce a novel approach combining serial sectioning of tissue with scanning electron microscopy (SEM) using a conductive silicon wafer as a support. Ribbons containing hundreds of 35 nm thick sections can be generated and imaged on the wafer at a lateral pixel resolution of 3.7 nm by recording the backscattered electrons with the in-lens detector of the SEM. The resulting electron micrographs are qualitatively comparable to those obtained by conventional TEM. S3EM images of the same region of interest in consecutive sections can be used for 3D reconstructions of large structures. We demonstrate the potential of this approach by reconstructing a 31.7 µm3 volume of a calyx of Held presynaptic terminal. The approach introduced here, Serial Section SEM (S3EM), for the first time provides the possibility to obtain 3D ultrastructure of large volumes with high resolution and to selectively and repetitively home in on structures of interest. S3EM accelerates process duration, is amenable to full automation and can be implemented with standard instrumentation. PMID:22523574

Serial section scanning electron microscopy (S3EM) on silicon wafers for ultra-structural volume imaging of cells and tissues.

PubMed

Horstmann, Heinz; Körber, Christoph; Sätzler, Kurt; Aydin, Daniel; Kuner, Thomas

2012-01-01

High resolution, three-dimensional (3D) representations of cellular ultrastructure are essential for structure function studies in all areas of cell biology. While limited subcellular volumes have been routinely examined using serial section transmission electron microscopy (ssTEM), complete ultrastructural reconstructions of large volumes, entire cells or even tissue are difficult to achieve using ssTEM. Here, we introduce a novel approach combining serial sectioning of tissue with scanning electron microscopy (SEM) using a conductive silicon wafer as a support. Ribbons containing hundreds of 35 nm thick sections can be generated and imaged on the wafer at a lateral pixel resolution of 3.7 nm by recording the backscattered electrons with the in-lens detector of the SEM. The resulting electron micrographs are qualitatively comparable to those obtained by conventional TEM. S(3)EM images of the same region of interest in consecutive sections can be used for 3D reconstructions of large structures. We demonstrate the potential of this approach by reconstructing a 31.7 µm(3) volume of a calyx of Held presynaptic terminal. The approach introduced here, Serial Section SEM (S(3)EM), for the first time provides the possibility to obtain 3D ultrastructure of large volumes with high resolution and to selectively and repetitively home in on structures of interest. S(3)EM accelerates process duration, is amenable to full automation and can be implemented with standard instrumentation.

Differential localization of SAP102 and PSD-95 is revealed in hippocampal spines using super-resolution light microscopy.

PubMed

Zheng, Chan-Ying; Wang, Ya-Xia; Kachar, Bechara; Petralia, Ronald S

2011-01-01

Synapse-associated protein 102 (SAP102) and postsynaptic density 95 (PSD-95) are two major cytoskeleton proteins in the postsynaptic density (PSD). Both of them belong to the membrane-associated guanylate kinase (MAGUK) family, which clusters and anchors glutamate receptors and other proteins at synapses. In our previous study, we found that SAP102 and PSD-95 have different distributions, using combined light/electron microscopy (LM/EM) methods.1 Here, we double labeled endogenous SAP102 and PSD-95 in mature hippocampal neurons, and then took images by two different kinds of super resolution microscopy-Stimulated Emission Depletion microscopy (STED) and DeltaVision OMX 3D super resolution microscopy. We found that our 2D and 3D super resolution data were consistent with our previous LM/EM data, showing significant differences in the localization of SAP102 and PSD-95 in spines: SAP102 is distributed in both the PSD and cytoplasm of spines, while PSD-95 is concentrated only in the PSD area. These results indicate functional differences between SAP102 and PSD-95 in synaptic organization and plasticity.

Characterization of fast photoelectron packets in weak and strong laser fields in ultrafast electron microscopy.

PubMed

Plemmons, Dayne A; Tae Park, Sang; Zewail, Ahmed H; Flannigan, David J

2014-11-01

The development of ultrafast electron microscopy (UEM) and variants thereof (e.g., photon-induced near-field electron microscopy, PINEM) has made it possible to image atomic-scale dynamics on the femtosecond timescale. Accessing the femtosecond regime with UEM currently relies on the generation of photoelectrons with an ultrafast laser pulse and operation in a stroboscopic pump-probe fashion. With this approach, temporal resolution is limited mainly by the durations of the pump laser pulse and probe electron packet. The ability to accurately determine the duration of the electron packets, and thus the instrument response function, is critically important for interpretation of dynamics occurring near the temporal resolution limit, in addition to quantifying the effects of the imaging mode. Here, we describe a technique for in situ characterization of ultrashort electron packets that makes use of coupling with photons in the evanescent near-field of the specimen. We show that within the weakly-interacting (i.e., low laser fluence) regime, the zero-loss peak temporal cross-section is precisely the convolution of electron packet and photon pulse profiles. Beyond this regime, we outline the effects of non-linear processes and show that temporal cross-sections of high-order peaks explicitly reveal the electron packet profile, while use of the zero-loss peak becomes increasingly unreliable. Copyright © 2014 Elsevier B.V. All rights reserved.

Software electron counting for low-dose scanning transmission electron microscopy.

PubMed

Mittelberger, Andreas; Kramberger, Christian; Meyer, Jannik C

2018-05-01

The performance of the detector is of key importance for low-dose imaging in transmission electron microscopy, and counting every single electron can be considered as the ultimate goal. In scanning transmission electron microscopy, low-dose imaging can be realized by very fast scanning, however, this also introduces artifacts and a loss of resolution in the scan direction. We have developed a software approach to correct for artifacts introduced by fast scans, making use of a scintillator and photomultiplier response that extends over several pixels. The parameters for this correction can be directly extracted from the raw image. Finally, the images can be converted into electron counts. This approach enables low-dose imaging in the scanning transmission electron microscope via high scan speeds while retaining the image quality of artifact-free slower scans. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Achromatic elemental mapping beyond the nanoscale in the transmission electron microscope.

PubMed

Urban, K W; Mayer, J; Jinschek, J R; Neish, M J; Lugg, N R; Allen, L J

2013-05-03

Newly developed achromatic electron optics allows the use of wide energy windows and makes feasible energy-filtered transmission electron microscopy (EFTEM) at atomic resolution. In this Letter we present EFTEM images formed using electrons that have undergone a silicon L(2,3) core-shell energy loss, exhibiting a resolution in EFTEM of 1.35 Å. This permits elemental mapping beyond the nanoscale provided that quantum mechanical calculations from first principles are done in tandem with the experiment to understand the physical information encoded in the images.

Composition measurement in substitutionally disordered materials by atomic resolution energy dispersive X-ray spectroscopy in scanning transmission electron microscopy.

PubMed

Chen, Z; Taplin, D J; Weyland, M; Allen, L J; Findlay, S D

2017-05-01

The increasing use of energy dispersive X-ray spectroscopy in atomic resolution scanning transmission electron microscopy invites the question of whether its success in precision composition determination at lower magnifications can be replicated in the atomic resolution regime. In this paper, we explore, through simulation, the prospects for composition measurement via the model system of Al x Ga 1-x As, discussing the approximations used in the modelling, the variability in the signal due to changes in configuration at constant composition, and the ability to distinguish between different compositions. Results are presented in such a way that the number of X-ray counts, and thus the expected variation due to counting statistics, can be gauged for a range of operating conditions. Copyright © 2016 Elsevier B.V. All rights reserved.

High-resolution electron microscopy and its applications.

PubMed

Li, F H

1987-12-01

A review of research on high-resolution electron microscopy (HREM) carried out at the Institute of Physics, the Chinese Academy of Sciences, is presented. Apart from the direct observation of crystal and quasicrystal defects for some alloys, oxides, minerals, etc., and the structure determination for some minute crystals, an approximate image-contrast theory named pseudo-weak-phase object approximation (PWPOA), which shows the image contrast change with crystal thickness, is described. Within the framework of PWPOA, the image contrast of lithium ions in the crystal of R-Li2Ti3O7 has been observed. The usefulness of diffraction analysis techniques such as the direct method and Patterson method in HREM is discussed. Image deconvolution and resolution enhancement for weak-phase objects by use of the direct method are illustrated. In addition, preliminary results of image restoration for thick crystals are given.

Four-Dimensional Ultrafast Electron Microscopy: Insights into an Emerging Technique.

PubMed

Adhikari, Aniruddha; Eliason, Jeffrey K; Sun, Jingya; Bose, Riya; Flannigan, David J; Mohammed, Omar F

2017-01-11

Four-dimensional ultrafast electron microscopy (4D-UEM) is a novel analytical technique that aims to fulfill the long-held dream of researchers to investigate materials at extremely short spatial and temporal resolutions by integrating the excellent spatial resolution of electron microscopes with the temporal resolution of ultrafast femtosecond laser-based spectroscopy. The ingenious use of pulsed photoelectrons to probe surfaces and volumes of materials enables time-resolved snapshots of the dynamics to be captured in a way hitherto impossible by other conventional techniques. The flexibility of 4D-UEM lies in the fact that it can be used in both the scanning (S-UEM) and transmission (UEM) modes depending upon the type of electron microscope involved. While UEM can be employed to monitor elementary structural changes and phase transitions in samples using real-space mapping, diffraction, electron energy-loss spectroscopy, and tomography, S-UEM is well suited to map ultrafast dynamical events on materials surfaces in space and time. This review provides an overview of the unique features that distinguish these techniques and also illustrates the applications of both S-UEM and UEM to a multitude of problems relevant to materials science and chemistry.

Immunogold scanning electron microscopy can reveal the polysaccharide architecture of xylem cell walls

PubMed Central

Sun, Yuliang; Juzenas, Kevin

2017-01-01

Abstract Immunofluorescence microscopy (IFM) and immunogold transmission electron microscopy (TEM) are the two main techniques commonly used to detect polysaccharides in plant cell walls. Both are important in localizing cell wall polysaccharides, but both have major limitations, such as low resolution in IFM and restricted sample size for immunogold TEM. In this study, we have developed a robust technique that combines immunocytochemistry with scanning electron microscopy (SEM) to study cell wall polysaccharide architecture in xylem cells at high resolution over large areas of sample. Using multiple cell wall monoclonal antibodies (mAbs), this immunogold SEM technique reliably localized groups of hemicellulosic and pectic polysaccharides in the cell walls of five different xylem structures (vessel elements, fibers, axial and ray parenchyma cells, and tyloses). This demonstrates its important advantages over the other two methods for studying cell wall polysaccharide composition and distribution in these structures. In addition, it can show the three-dimensional distribution of a polysaccharide group in the vessel lateral wall and the polysaccharide components in the cell wall of developing tyloses. This technique, therefore, should be valuable for understanding the cell wall polysaccharide composition, architecture and functions of diverse cell types. PMID:28398585

Serial block face-scanning electron microscopy: a tool for studying embryonic development at the cell-matrix interface.

PubMed

Starborg, Tobias; Kadler, Karl E

2015-03-01

Studies of gene regulation, signaling pathways, and stem cell biology are contributing greatly to our understanding of early embryonic vertebrate development. However, much less is known about the events during the latter half of embryonic development, when tissues comprising mostly extracellular matrix (ECM) are formed. The matrix extends far beyond the boundaries of individual cells and is refractory to study by conventional biochemical and molecular techniques; thus major gaps exist in our knowledge of the formation and three-dimensional (3D) organization of the dense tissues that form the bulk of adult vertebrates. Serial block face-scanning electron microscopy (SBF-SEM) has the ability to image volumes of tissue containing numerous cells at a resolution sufficient to study the organization of the ECM. Furthermore, whereas light microscopy was once relatively straightforward and electron microscopy was performed in specialist laboratories, the tables are turned; SBF-SEM is relatively straightforward and is becoming routine in high-end resolution studies of embryonic structures in vivo. In this review, we discuss the emergence of SBF-SEM as a tool for studying embryonic vertebrate development. © 2015 Wiley Periodicals, Inc.

Cryo-electron microscopy of membrane proteins.

PubMed

Goldie, Kenneth N; Abeyrathne, Priyanka; Kebbel, Fabian; Chami, Mohamed; Ringler, Philippe; Stahlberg, Henning

2014-01-01

Electron crystallography is used to study membrane proteins in the form of planar, two-dimensional (2D) crystals, or other crystalline arrays such as tubular crystals. This method has been used to determine the atomic resolution structures of bacteriorhodopsin, tubulin, aquaporins, and several other membrane proteins. In addition, a large number of membrane protein structures were studied at a slightly lower resolution, whereby at least secondary structure motifs could be identified.In order to conserve the structural details of delicate crystalline arrays, cryo-electron microscopy (cryo-EM) allows imaging and/or electron diffraction of membrane proteins in their close-to-native state within a lipid bilayer membrane.To achieve ultimate high-resolution structural information of 2D crystals, meticulous sample preparation for electron crystallography is of outmost importance. Beam-induced specimen drift and lack of specimen flatness can severely affect the attainable resolution of images for tilted samples. Sample preparations that sandwich the 2D crystals between symmetrical carbon films reduce the beam-induced specimen drift, and the flatness of the preparations can be optimized by the choice of the grid material and the preparation protocol.Data collection in the cryo-electron microscope using either the imaging or the electron diffraction mode has to be performed applying low-dose procedures. Spot-scanning further reduces the effects of beam-induced drift. Data collection using automated acquisition schemes, along with improved and user-friendlier data processing software, is increasingly being used and is likely to bring the technique to a wider user base.

Correction of image drift and distortion in a scanning electron microscopy.

PubMed

Jin, P; Li, X

2015-12-01

Continuous research on small-scale mechanical structures and systems has attracted strong demand for ultrafine deformation and strain measurements. Conventional optical microscope cannot meet such requirements owing to its lower spatial resolution. Therefore, high-resolution scanning electron microscope has become the preferred system for high spatial resolution imaging and measurements. However, scanning electron microscope usually is contaminated by distortion and drift aberrations which cause serious errors to precise imaging and measurements of tiny structures. This paper develops a new method to correct drift and distortion aberrations of scanning electron microscope images, and evaluates the effect of correction by comparing corrected images with scanning electron microscope image of a standard sample. The drift correction is based on the interpolation scheme, where a series of images are captured at one location of the sample and perform image correlation between the first image and the consequent images to interpolate the drift-time relationship of scanning electron microscope images. The distortion correction employs the axial symmetry model of charged particle imaging theory to two images sharing with the same location of one object under different imaging fields of view. The difference apart from rigid displacement between the mentioned two images will give distortion parameters. Three-order precision is considered in the model and experiment shows that one pixel maximum correction is obtained for the employed high-resolution electron microscopic system. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

Cassette Series Designed for Live-Cell Imaging of Proteins and High Resolution Techniques in Yeast

PubMed Central

Young, Carissa L.; Raden, David L.; Caplan, Jeffrey; Czymmek, Kirk; Robinson, Anne S.

2012-01-01

During the past decade, it has become clear that protein function and regulation are highly dependent upon intracellular localization. Although fluorescent protein variants are ubiquitously used to monitor protein dynamics, localization, and abundance; fluorescent light microscopy techniques often lack the resolution to explore protein heterogeneity and cellular ultrastructure. Several approaches have been developed to identify, characterize, and monitor the spatial localization of proteins and complexes at the sub-organelle level; yet, many of these techniques have not been applied to yeast. Thus, we have constructed a series of cassettes containing codon-optimized epitope tags, fluorescent protein variants that cover the full spectrum of visible light, a TetCys motif used for FlAsH-based localization, and the first evaluation in yeast of a photoswitchable variant – mEos2 – to monitor discrete subpopulations of proteins via confocal microscopy. This series of modules, complete with six different selection markers, provides the optimal flexibility during live-cell imaging and multicolor labeling in vivo. Furthermore, high-resolution imaging techniques include the yeast-enhanced TetCys motif that is compatible with diaminobenzidine photooxidation used for protein localization by electron microscopy and mEos2 that is ideal for super-resolution microscopy. We have examined the utility of our cassettes by analyzing all probes fused to the C-terminus of Sec61, a polytopic membrane protein of the endoplasmic reticulum of moderate protein concentration, in order to directly compare fluorescent probes, their utility and technical applications. Our series of cassettes expand the repertoire of molecular tools available to advance targeted spatiotemporal investigations using multiple live-cell, super-resolution or electron microscopy imaging techniques. PMID:22473760

Secretory glands and microvascular systems imaged in aqueous solution by atmospheric scanning electron microscopy (ASEM).

PubMed

Yamazawa, Toshiko; Nakamura, Naotoshi; Sato, Mari; Sato, Chikara

2016-12-01

Exocrine glands, e.g., salivary and pancreatic glands, play an important role in digestive enzyme secretion, while endocrine glands, e.g., pancreatic islets, secrete hormones that regulate blood glucose levels. The dysfunction of these secretory organs immediately leads to various diseases, such as diabetes or Sjögren's syndrome, by poorly understood mechanisms. Gland-related diseases have been studied by optical microscopy (OM), and at higher resolution by transmission electron microscopy (TEM) of Epon embedded samples, which necessitates hydrophobic sample pretreatment. Here, we report the direct observation of tissue in aqueous solution by atmospheric scanning electron microscopy (ASEM). Salivary glands, lacrimal glands, and pancreas were fixed, sectioned into slabs, stained with phosphotungstic acid (PTA), and inspected in radical scavenger d-glucose solution from below by an inverted scanning electron microscopy (SEM), guided by optical microscopy from above to target the tissue substructures. A 2- to 3-µm specimen thickness was visualized by the SEM. In secretory cells, cytoplasmic vesicles and other organelles were clearly imaged at high resolution, and the former could be classified according to the degree of PTA staining. In islets of Langerhans, the microvascular system used as an outlet by the secretory cells was also clearly observed. Microvascular system is also critically involved in the onset of diabetic complications and was clearly visible in subcutaneous tissue imaged by ASEM. The results suggest the use of in-solution ASEM for histology and to study vesicle secretion systems. Further, the high-throughput of ASEM makes it a potential tool for the diagnosis of exocrine and endocrine-related diseases. © 2016 Wiley Periodicals, Inc.

New advances in scanning microscopy and its application to study parasitic protozoa.

PubMed

de Souza, Wanderley; Attias, Marcia

2018-07-01

Scanning electron microscopy has been used to observe and study parasitic protozoa for at least 40 years. However, field emission electron sources, as well as improvements in lenses and detectors, brought the resolution power of scanning electron microscopes (SEM) to a new level. Parallel to the refinement of instruments, protocols for preservation of the ultrastructure, immunolabeling, exposure of cytoskeleton and inner structures of parasites and host cells were developed. This review is focused on protozoan parasites of medical and veterinary relevance, e.g., Toxoplasma gondii, Tritrichomonas foetus, Giardia intestinalis, and Trypanosoma cruzi, compilating the main achievements in describing the fine ultrastructure of their surface, cytoskeleton and interaction with host cells. Two new resources, namely, Helium Ion Microscopy (HIM) and Slice and View, using either Focused Ion Beam (FIB) abrasion or Microtome Serial Sectioning (MSS) within the microscope chamber, combined to backscattered electron imaging of fixed (chemically or by quick freezing followed by freeze substitution and resin embedded samples is bringing an exponential amount of valuable information. In HIM there is no need of conductive coating and the depth of field is much higher than in any field emission SEM. As for FIB- and MSS-SEM, high resolution 3-D models of areas and volumes larger than any other technique allows can be obtained. The main results achieved with all these technological tools and some protocols for sample preparation are included in this review. In addition, we included some results obtained with environmental/low vacuum scanning microscopy and cryo-scanning electron microscopy, both promising, but not yet largely employed SEM modalities. Copyright © 2018. Published by Elsevier Inc.

Correlated fluorescence microscopy and cryo-electron tomography of virus-infected or transfected mammalian cells

PubMed Central

Hampton, Cheri M; Strauss, Joshua D; Ke, Zunlong; Dillard, Rebecca S; Hammonds, Jason E; Alonas, Eric; Desai, Tanay M; Marin, Mariana; Storms, Rachel E; Leon, Fredrick; Melikyan, Gregory B; Santangelo, Philip J; Spearman, Paul W; Wright, Elizabeth R

2016-01-01

Correlative light and electron microscopy (CLEM) combines spatiotemporal information from fluorescence light microscopy (fLM) with high-resolution structural data from cryo-electron tomography (cryo-ET). These technologies provide opportunities to bridge knowledge gaps between cell and structural biology. Here we describe our protocol for correlated cryo-fLM, cryo-electron microscopy (cryo-EM), and cryo-ET (i.e., cryo-CLEM) of virus-infected or transfected mammalian cells. Mammalian-derived cells are cultured on EM substrates, using optimized conditions that ensure that the cells are spread thinly across the substrate and are not physically disrupted. The cells are then screened by fLM and vitrified before acquisition of cryo-fLM and cryo-ET images, which is followed by data processing. A complete session from grid preparation through data collection and processing takes 5–15 d for an individual experienced in cryo-EM. PMID:27977021

In-line three-dimensional holography of nanocrystalline objects at atomic resolution

DOE PAGES

Chen, F. -R.; Van Dyck, D.; Kisielowski, C.

2016-02-18

We report that resolution and sensitivity of the latest generation aberration-corrected transmission electron microscopes allow the vast majority of single atoms to be imaged with sub-Ångstrom resolution and their locations determined in an image plane with a precision that exceeds the 1.9-pm wavelength of 300 kV electrons. Such unprecedented performance allows expansion of electron microscopic investigations with atomic resolution into the third dimension. Here we show a general tomographic method to recover the three-dimensional shape of a crystalline particle from high-resolution images of a single projection without the need for sample rotation. The method is compatible with low dose ratemore » electron microscopy, which improves on signal quality, while minimizing electron beam-induced structure modifications even for small particles or surfaces. Lastly, we apply it to germanium, gold and magnesium oxide particles, and achieve a depth resolution of 1–2 Å, which is smaller than inter-atomic distances.« less

DOE Office of Scientific and Technical Information (OSTI.GOV)

Juffmann, Thomas; Koppell, Stewart A.; Klopfer, Brannon B.

Feynman once asked physicists to build better electron microscopes to be able to watch biology at work. While electron microscopes can now provide atomic resolution, electron beam induced specimen damage precludes high resolution imaging of sensitive materials, such as single proteins or polymers. Here, we use simulations to show that an electron microscope based on a multi-pass measurement protocol enables imaging of single proteins, without averaging structures over multiple images. While we demonstrate the method for particular imaging targets, the approach is broadly applicable and is expected to improve resolution and sensitivity for a range of electron microscopy imaging modalities,more » including, for example, scanning and spectroscopic techniques. The approach implements a quantum mechanically optimal strategy which under idealized conditions can be considered interaction-free.« less

Attainment of 40.5 pm spatial resolution using 300 kV scanning transmission electron microscope equipped with fifth-order aberration corrector.

PubMed

Morishita, Shigeyuki; Ishikawa, Ryo; Kohno, Yuji; Sawada, Hidetaka; Shibata, Naoya; Ikuhara, Yuichi

2018-02-01

The achievement of a fine electron probe for high-resolution imaging in scanning transmission electron microscopy requires technological developments, especially in electron optics. For this purpose, we developed a microscope with a fifth-order aberration corrector that operates at 300 kV. The contrast flat region in an experimental Ronchigram, which indicates the aberration-free angle, was expanded to 70 mrad. By using a probe with convergence angle of 40 mrad in the scanning transmission electron microscope at 300 kV, we attained the spatial resolution of 40.5 pm, which is the projected interatomic distance between Ga-Ga atomic columns of GaN observed along [212] direction.

Comparison of the morphology, chemical composition and microstructure of cryptocrystalline graphite and carbon black

NASA Astrophysics Data System (ADS)

Quan, Ying; Liu, Qinfu; Zhang, Shilong; Zhang, Shuai

2018-07-01

The structures of cryptocrystalline graphite (CG) and carbon black (CB) have been analyzed using scanning electron microscopy (SEM), transmission electron microscopy (TEM), organic elemental analysis (OEA), X-ray diffraction (XRD), RAMAN and high-resolution transmission electron microscopy (HRTEM). These results indicate that CG has the same elemental composition as CB, with carbon being the major element present. SL sample (CG with low graphitization degree) and CB exhibit similar microcrystalline structures. CG was shown to contain a layered graphitic structure that was significantly different to the primary spherical particles present in CB. It is proposed that these CG sheets may potentially be reduced and delaminated to afford multilayer graphene structures with improved material properties.

Hydrothermal Synthesis of Nanoclusters of ZnS Comprised on Nanowires

PubMed Central

Ibupoto, Zafar Hussain; Khun, Kimleang; Liu, Xianjie; Willander, Magnus

2013-01-01

Cetyltrimethyl ammonium bromide cationic (CTAB) surfactant was used as template for the synthesis of nanoclusters of ZnS composed of nanowires, by hydrothermal method. The structural and morphological studies were performed by using X-ray diffraction (XRD), scanning electron microscopy (SEM) and high resolution transmission electron microscopy (HRTEM) techniques. The synthesized ZnS nanoclusters are composed of nanowires and high yield on the substrate was observed. The ZnS nanocrystalline consists of hexagonal phase and polycrystalline in nature. The chemical composition of ZnS nanoclusters composed of nanowires was studied by X-ray photo electron microscopy (XPS). This investigation has shown that the ZnS nanoclusters are composed of Zn and S atoms. PMID:28348350

Hydrothermal Synthesis of Nanoclusters of ZnS Comprised on Nanowires.

PubMed

Ibupoto, Zafar Hussain; Khun, Kimleang; Liu, Xianjie; Willander, Magnus

2013-09-09

Cetyltrimethyl ammonium bromide cationic (CTAB) surfactant was used as template for the synthesis of nanoclusters of ZnS composed of nanowires, by hydrothermal method. The structural and morphological studies were performed by using X-ray diffraction (XRD), scanning electron microscopy (SEM) and high resolution transmission electron microscopy (HRTEM) techniques. The synthesized ZnS nanoclusters are composed of nanowires and high yield on the substrate was observed. The ZnS nanocrystalline consists of hexagonal phase and polycrystalline in nature. The chemical composition of ZnS nanoclusters composed of nanowires was studied by X-ray photo electron microscopy (XPS). This investigation has shown that the ZnS nanoclusters are composed of Zn and S atoms.

Recent advances in electron tomography: TEM and HAADF-STEM tomography for materials science and semiconductor applications.

PubMed

Kübel, Christian; Voigt, Andreas; Schoenmakers, Remco; Otten, Max; Su, David; Lee, Tan-Chen; Carlsson, Anna; Bradley, John

2005-10-01

Electron tomography is a well-established technique for three-dimensional structure determination of (almost) amorphous specimens in life sciences applications. With the recent advances in nanotechnology and the semiconductor industry, there is also an increasing need for high-resolution three-dimensional (3D) structural information in physical sciences. In this article, we evaluate the capabilities and limitations of transmission electron microscopy (TEM) and high-angle-annular-dark-field scanning transmission electron microscopy (HAADF-STEM) tomography for the 3D structural characterization of partially crystalline to highly crystalline materials. Our analysis of catalysts, a hydrogen storage material, and different semiconductor devices shows that features with a diameter as small as 1-2 nm can be resolved in three dimensions by electron tomography. For partially crystalline materials with small single crystalline domains, bright-field TEM tomography provides reliable 3D structural information. HAADF-STEM tomography is more versatile and can also be used for high-resolution 3D imaging of highly crystalline materials such as semiconductor devices.

Correlative cryo-fluorescence light microscopy and cryo-electron tomography of Streptomyces.

PubMed

Koning, Roman I; Celler, Katherine; Willemse, Joost; Bos, Erik; van Wezel, Gilles P; Koster, Abraham J

2014-01-01

Light microscopy and electron microscopy are complementary techniques that in a correlative approach enable identification and targeting of fluorescently labeled structures in situ for three-dimensional imaging at nanometer resolution. Correlative imaging allows electron microscopic images to be positioned in a broader temporal and spatial context. We employed cryo-correlative light and electron microscopy (cryo-CLEM), combining cryo-fluorescence light microscopy and cryo-electron tomography, on vitrified Streptomyces bacteria to study cell division. Streptomycetes are mycelial bacteria that grow as long hyphae and reproduce via sporulation. On solid media, Streptomyces subsequently form distinct aerial mycelia where cell division leads to the formation of unigenomic spores which separate and disperse to form new colonies. In liquid media, only vegetative hyphae are present divided by noncell separating crosswalls. Their multicellular life style makes them exciting model systems for the study of bacterial development and cell division. Complex intracellular structures have been visualized with transmission electron microscopy. Here, we describe the methods for cryo-CLEM that we applied for studying Streptomyces. These methods include cell growth, fluorescent labeling, cryo-fixation by vitrification, cryo-light microscopy using a Linkam cryo-stage, image overlay and relocation, cryo-electron tomography using a Titan Krios, and tomographic reconstruction. Additionally, methods for segmentation, volume rendering, and visualization of the correlative data are described. © 2014 Elsevier Inc. All rights reserved.

X-ray ptychographic and fluorescence microscopy of frozen-hydrated cells using continuous scanning

DOE PAGES

Deng, Junjing; Vine, David J.; Chen, Si; ...

2017-03-27

X-ray microscopy can be used to image whole, unsectioned cells in their native hydrated state. It complements the higher resolution of electron microscopy for submicrometer thick specimens, and the molecule-specific imaging capabilites of fluorescence light microscopy. We describe here the first use of fast, continuous x-ray scanning of frozen hydrated cells for simultaneous sub-20 nm resolution ptychographic transmission imaging with high contrast, and sub-100 nm resolution deconvolved x-ray fluorescence imaging of diffusible and bound ions at native concentrations, without the need to add specific labels. Here, by working with cells that have been rapidly frozen without the use of chemicalmore » fixatives, and imaging them under cryogenic conditions, we are able to obtain images with well preserved structural and chemical composition, and sufficient stability against radiation damage to allow for multiple images to be obtained with no observable change.« less

High-resolution electron microscopy and electron energy-loss spectroscopy of giant palladium clusters

NASA Astrophysics Data System (ADS)

Oleshko, V.; Volkov, V.; Gijbels, R.; Jacob, W.; Vargaftik, M.; Moiseev, I.; van Tendeloo, G.

1995-12-01

Combined structural and chemical characterization of cationic polynuclear palladium coordination compounds Pd561L60(OAc)180, where L=1,10-phenantroline or 2,2'-bipyridine has been carried out by high-resolution electron microscopy (HREM) and analytical electron microscopy methods including electron energy-loss spectroscopy (EELS), zero-loss electron spectroscopic imaging, and energy-dispersive X-ray spectroscopy (EDX). The cell structure of the cluster matter with almost completely uniform metal core size distributions centered around 2.3 ±0.5 nm was observed. Zero-loss energy filtering allowed to improve the image contrast and resolution. HREM images showed that most of the palladium clusters had a cubo-octahedral shape. Some of them had a distorted icosahedron structure exhibiting multiple twinning. The selected-area electron diffraction patterns confirmed the face centered cubic structure with lattice parameter close to that of metallic palladium. The energy-loss spectra of the populations of clusters contained several bands, which could be assigned to the delayed Pd M4, 5-edge at 362 eV, the Pd M3-edge at 533 eV and the Pd M2-edge at 561 eV, the NK-edge at about 400 eV, the O K-edge at 532 eV overlapping with the Pd M3-edge and the carbon C K-edge at 284 eV. Background subtraction was applied to reveal the exact positions and fine structure of low intensity elemental peaks. EELS evaluations have been confirmed by EDX. The recorded series of the Pd M-edges and the N K-edge in the spectra of the giant palladium clusters obviously were related to Pd-Pd- and Pd-ligand bonding.

The quantitative analysis of silicon carbide surface smoothing by Ar and Xe cluster ions

NASA Astrophysics Data System (ADS)

Ieshkin, A. E.; Kireev, D. S.; Ermakov, Yu. A.; Trifonov, A. S.; Presnov, D. E.; Garshev, A. V.; Anufriev, Yu. V.; Prokhorova, I. G.; Krupenin, V. A.; Chernysh, V. S.

2018-04-01

The gas cluster ion beam technique was used for the silicon carbide crystal surface smoothing. The effect of processing by two inert cluster ions, argon and xenon, was quantitatively compared. While argon is a standard element for GCIB, results for xenon clusters were not reported yet. Scanning probe microscopy and high resolution transmission electron microscopy techniques were used for the analysis of the surface roughness and surface crystal layer quality. The gas cluster ion beam processing results in surface relief smoothing down to average roughness about 1 nm for both elements. It was shown that xenon as the working gas is more effective: sputtering rate for xenon clusters is 2.5 times higher than for argon at the same beam energy. High resolution transmission electron microscopy analysis of the surface defect layer gives values of 7 ± 2 nm and 8 ± 2 nm for treatment with argon and xenon clusters.

Energy-Filtered High-Resolution Electron Microscopy for Quantitative Solid State Structure Determination

DTIC Science & Technology

1997-01-01

is given by [19]: vP = e Ïne /(m0««0) , where e is the elementary charge, m0 is the electron rest mass, ne is the density of free electrons, and...and energy filtering systems. Fig. 8. EELS spectra acquired from a Ni/Ti multilayer specimen when the elctron beam is positioned at (a) Ti and (b) Ni...332 (1991). [11] T. Tanji, K. Urata, K. Ishizuka, Q. Ru, and A. Tonomura, Ultramicroscopy 49, 259–264 (1993). [12] D. E . Newbury, Microscopy: The Key

Nanoscale live cell optical imaging of the dynamics of intracellular microvesicles in neural cells.

PubMed

Lee, Sohee; Heo, Chaejeong; Suh, Minah; Lee, Young Hee

2013-11-01

Recent advances in biotechnology and imaging technology have provided great opportunities to investigate cellular dynamics. Conventional imaging methods such as transmission electron microscopy, scanning electron microscopy, and atomic force microscopy are powerful techniques for cellular imaging, even at the nanoscale level. However, these techniques have limitations applications in live cell imaging because of the experimental preparation required, namely cell fixation, and the innately small field of view. In this study, we developed a nanoscale optical imaging (NOI) system that combines a conventional optical microscope with a high resolution dark-field condenser (Cytoviva, Inc.) and halogen illuminator. The NOI system's maximum resolution for live cell imaging is around 100 nm. We utilized NOI to investigate the dynamics of intracellular microvesicles of neural cells without immunocytological analysis. In particular, we studied direct, active random, and moderate random dynamic motions of intracellular microvesicles and visualized lysosomal vesicle changes after treatment of cells with a lysosomal inhibitor (NH4Cl). Our results indicate that the NOI system is a feasible, high-resolution optical imaging system for live small organelles that does not require complicated optics or immunocytological staining processes.

3D structure of individual nanocrystals in solution by electron microscopy

DOE PAGES

Park, Jungwok; Elmlund, Hans; Ercius, Peter; ...

2015-07-17

Here, knowledge about the synthesis, growth mechanisms, and physical properties of colloidal nanoparticles has been limited by technical impediments. We introduce a method for determining three-dimensional (3D) structures of individual nanoparticles in solution. We combine a graphene liquid cell, high-resolution transmission electron microscopy, a direct electron detector, and an algorithm for single-particle 3D reconstruction originally developed for analysis of biological molecules. This method yielded two 3D structures of individual platinum nanocrystals at near-atomic resolution. Because our method derives the 3D structure from images of individual nanoparticles rotating freely in solution, it enables the analysis of heterogeneous populations of potentially unorderedmore » nanoparticles that are synthesized in solution, thereby providing a means to understand the structure and stability of defects at the nanoscale.« less

Nanoparticle imaging. 3D structure of individual nanocrystals in solution by electron microscopy.

PubMed

Park, Jungwon; Elmlund, Hans; Ercius, Peter; Yuk, Jong Min; Limmer, David T; Chen, Qian; Kim, Kwanpyo; Han, Sang Hoon; Weitz, David A; Zettl, A; Alivisatos, A Paul

2015-07-17

Knowledge about the synthesis, growth mechanisms, and physical properties of colloidal nanoparticles has been limited by technical impediments. We introduce a method for determining three-dimensional (3D) structures of individual nanoparticles in solution. We combine a graphene liquid cell, high-resolution transmission electron microscopy, a direct electron detector, and an algorithm for single-particle 3D reconstruction originally developed for analysis of biological molecules. This method yielded two 3D structures of individual platinum nanocrystals at near-atomic resolution. Because our method derives the 3D structure from images of individual nanoparticles rotating freely in solution, it enables the analysis of heterogeneous populations of potentially unordered nanoparticles that are synthesized in solution, thereby providing a means to understand the structure and stability of defects at the nanoscale. Copyright © 2015, American Association for the Advancement of Science.

A history of scanning electron microscopy developments: towards "wet-STEM" imaging.

PubMed

Bogner, A; Jouneau, P-H; Thollet, G; Basset, D; Gauthier, C

2007-01-01

A recently developed imaging mode called "wet-STEM" and new developments in environmental scanning electron microscopy (ESEM) allows the observation of nano-objects suspended in a liquid phase, with a few manometers resolution and a good signal to noise ratio. The idea behind this technique is simply to perform STEM-in-SEM, that is SEM in transmission mode, in an environmental SEM. The purpose of the present contribution is to highlight the main advances that contributed to development of the wet-STEM technique. Although simple in principle, the wet-STEM imaging mode would have been limited before high brightness electron sources became available, and needed some progresses and improvements in ESEM. This new technique extends the scope of SEM as a high-resolution microscope, relatively cheap and widely available imaging tool, for a wider variety of samples.

Unusual inhomogeneous microstructures in charge glass state of PbCrO3

NASA Astrophysics Data System (ADS)

Kurushima, Kosuke; Tsukasaki, Hirofumi; Ogata, Takahiro; Sakai, Yuki; Azuma, Masaki; Ishii, Yui; Mori, Shigeo

2018-05-01

We investigated the microstructures and local structures of perovskite PbCrO3, which shows a metal-to-insulator transition and a 9.8% volume collapse, by electron diffraction, high-resolution transmission electron microscopy (TEM), and high-angle annular dark-field scanning transmission electron microscopy (HAADF-STEM). It is revealed that the charge glass state is characterized by the unique coexistence of the crystalline state with a cubic symmetry on average and the noncrystalline state. HAADF-STEM observation at atomic resolution revealed that Pb ions were displaced from the ideal A site position of the cubic perovskite structure, which gives rise to characteristic diffuse scatterings around the fundamental Bragg reflections. These structural inhomogeneities are crucial to the understanding of the unique physical properties in the charge glass state of PbCrO3.

3D structure of individual nanocrystals in solution by electron microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Jungwok; Elmlund, Hans; Ercius, Peter

Here, knowledge about the synthesis, growth mechanisms, and physical properties of colloidal nanoparticles has been limited by technical impediments. We introduce a method for determining three-dimensional (3D) structures of individual nanoparticles in solution. We combine a graphene liquid cell, high-resolution transmission electron microscopy, a direct electron detector, and an algorithm for single-particle 3D reconstruction originally developed for analysis of biological molecules. This method yielded two 3D structures of individual platinum nanocrystals at near-atomic resolution. Because our method derives the 3D structure from images of individual nanoparticles rotating freely in solution, it enables the analysis of heterogeneous populations of potentially unorderedmore » nanoparticles that are synthesized in solution, thereby providing a means to understand the structure and stability of defects at the nanoscale.« less

TEM characterization of nanodiamond thin films.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Qin, L.-C.; Zhou, D.; Krauss, A. R.

The microstructure of thin films grown by microwave plasma-enhanced chemical vapor deposition (MPCVD) from fullerene C{sub 60} precursors has been characterized by scanning electron microscopy (SEM), selected-area electron diffraction (SAED), bright-field electron microscopy, high-resolution electron microscopy (HREM), and parallel electron energy loss spectroscopy (PEELS). The films are composed of nanosize crystallites of diamond, and no graphitic or amorphous phases were observed. The diamond crystallite size measured from lattice images shows that most grains range between 3-5 nm, reflecting a gamma distribution. SAED gave no evidence of either sp2-bonded glassy carbon or sp3-bonded diamondlike amorphous carbon. The sp2-bonded configuration found inmore » PEELS was attributed to grain boundary carbon atoms, which constitute 5-10% of the total. Occasionally observed larger diamond grains tend to be highly faulted.« less

Nanoparticle discrimination based on wavelength and lifetime-multiplexed cathodoluminescence microscopy.

PubMed

Garming, Mathijs W H; Weppelman, I Gerward C; de Boer, Pascal; Martínez, Felipe Perona; Schirhagl, Romana; Hoogenboom, Jacob P; Moerland, Robert J

2017-08-31

Nanomaterials can be identified in high-resolution electron microscopy images using spectrally-selective cathodoluminescence. Capabilities for multiplex detection can however be limited, e.g., due to spectral overlap or availability of filters. Also, the available photon flux may be limited due to degradation under electron irradiation. Here, we demonstrate single-pass cathodoluminescence-lifetime based discrimination of different nanoparticles, using a pulsed electron beam. We also show that cathodoluminescence lifetime is a robust parameter even when the nanoparticle cathodoluminescence intensity decays over an order of magnitude. We create lifetime maps, where the lifetime of the cathodoluminescence emission is correlated with the emission intensity and secondary-electron images. The consistency of lifetime-based discrimination is verified by also correlating the emission wavelength and the lifetime of nanoparticles. Our results show how cathodoluminescence lifetime provides an additional channel of information in electron microscopy.

Characterization of Ultra-fine Grained and Nanocrystalline Materials Using Transmission Kikuchi Diffraction

PubMed Central

Proust, Gwénaëlle; Trimby, Patrick; Piazolo, Sandra; Retraint, Delphine

2017-01-01

One of the challenges in microstructure analysis nowadays resides in the reliable and accurate characterization of ultra-fine grained (UFG) and nanocrystalline materials. The traditional techniques associated with scanning electron microscopy (SEM), such as electron backscatter diffraction (EBSD), do not possess the required spatial resolution due to the large interaction volume between the electrons from the beam and the atoms of the material. Transmission electron microscopy (TEM) has the required spatial resolution. However, due to a lack of automation in the analysis system, the rate of data acquisition is slow which limits the area of the specimen that can be characterized. This paper presents a new characterization technique, Transmission Kikuchi Diffraction (TKD), which enables the analysis of the microstructure of UFG and nanocrystalline materials using an SEM equipped with a standard EBSD system. The spatial resolution of this technique can reach 2 nm. This technique can be applied to a large range of materials that would be difficult to analyze using traditional EBSD. After presenting the experimental set up and describing the different steps necessary to realize a TKD analysis, examples of its use on metal alloys and minerals are shown to illustrate the resolution of the technique and its flexibility in term of material to be characterized. PMID:28447998

Characterization of Ultra-fine Grained and Nanocrystalline Materials Using Transmission Kikuchi Diffraction.

PubMed

Proust, Gwénaëlle; Trimby, Patrick; Piazolo, Sandra; Retraint, Delphine

2017-04-01

One of the challenges in microstructure analysis nowadays resides in the reliable and accurate characterization of ultra-fine grained (UFG) and nanocrystalline materials. The traditional techniques associated with scanning electron microscopy (SEM), such as electron backscatter diffraction (EBSD), do not possess the required spatial resolution due to the large interaction volume between the electrons from the beam and the atoms of the material. Transmission electron microscopy (TEM) has the required spatial resolution. However, due to a lack of automation in the analysis system, the rate of data acquisition is slow which limits the area of the specimen that can be characterized. This paper presents a new characterization technique, Transmission Kikuchi Diffraction (TKD), which enables the analysis of the microstructure of UFG and nanocrystalline materials using an SEM equipped with a standard EBSD system. The spatial resolution of this technique can reach 2 nm. This technique can be applied to a large range of materials that would be difficult to analyze using traditional EBSD. After presenting the experimental set up and describing the different steps necessary to realize a TKD analysis, examples of its use on metal alloys and minerals are shown to illustrate the resolution of the technique and its flexibility in term of material to be characterized.

The application of scanning electron microscopy to fractography

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brooks, C.R.; McGill, B.L.

1994-10-01

Many failures involve fracture, and determination of the fracture process is a key factor in understanding the failure. This is frequently accomplished by characterizing the topography of the fracture surface. Scanning electron microscopy has a prominent role in fractography due to three features of the scanning electron microscope (SEM): high resolution, great depth of field, and the ability to obtain chemical information via analysis of the X-rays generated by the electrons. A qualitative treatment is presented of the interaction of electrons with a sample and the effect of the SEM operating parameters on image formation, quality, and X-ray analysis. Fractographsmore » are presented to illustrate these features of scanning electron microscopy and to illustrate the limitations and precautions in obtaining fractographs and x-ray analyses. The review is concluded with examples of fracture surface features of metallic, ceramic, and polymeric materials.« less

The spatial coherence function in scanning transmission electron microscopy and spectroscopy.

PubMed

Nguyen, D T; Findlay, S D; Etheridge, J

2014-11-01

We investigate the implications of the form of the spatial coherence function, also referred to as the effective source distribution, for quantitative analysis in scanning transmission electron microscopy, and in particular for interpreting the spatial origin of imaging and spectroscopy signals. These questions are explored using three different source distribution models applied to a GaAs crystal case study. The shape of the effective source distribution was found to have a strong influence not only on the scanning transmission electron microscopy (STEM) image contrast, but also on the distribution of the scattered electron wavefield and hence on the spatial origin of the detected electron intensities. The implications this has for measuring structure, composition and bonding at atomic resolution via annular dark field, X-ray and electron energy loss STEM imaging are discussed. Copyright © 2014 Elsevier B.V. All rights reserved.

X-ray structure determination using low-resolution electron microscopy maps for molecular replacement

DOE PAGES

Jackson, Ryan N.; McCoy, Airlie J.; Terwilliger, Thomas C.; ...

2015-07-30

Structures of multi-subunit macromolecular machines are primarily determined by either electron microscopy (EM) or X-ray crystallography. In many cases, a structure for a complex can be obtained at low resolution (at a coarse level of detail) with EM and at higher resolution (with finer detail) by X-ray crystallography. The integration of these two structural techniques is becoming increasingly important for generating atomic models of macromolecular complexes. A low-resolution EM image can be a powerful tool for obtaining the "phase" information that is missing from an X-ray crystallography experiment, however integration of EM and X-ray diffraction data has been technically challenging.more » Here we show a step-by-step protocol that explains how low-resolution EM maps can be placed in the crystallographic unit cell by molecular replacement, and how initial phases computed from the placed EM density are extended to high resolution by averaging maps over non-crystallographic symmetry. As the resolution gap between EM and Xray crystallography continues to narrow, the use of EM maps to help with X-ray crystal structure determination, as described in this protocol, will become increasingly effective.« less

Super-Resolution Image Reconstruction by Nonlocal Means Applied to High-Angle Annular Darkfield Scanning Transmission Electron Microscopy (HAADF-STEM)

DTIC Science & Technology

2009-10-06

When talking about superresolution we always mean to recover the level of resolution set by the microscope, but by using a time series of low...on low resolution possibly very noisy data, is not feasible. Thus, standard superresolution concepts as described above that are based on registration

Environmental scanning electron microscopy in cell biology.

PubMed

McGregor, J E; Staniewicz, L T L; Guthrie Neé Kirk, S E; Donald, A M

2013-01-01

Environmental scanning electron microscopy (ESEM) (1) is an imaging technique which allows hydrated, insulating samples to be imaged under an electron beam. The resolution afforded by this technique is higher than conventional optical microscopy but lower than conventional scanning electron microscopy (CSEM). The major advantage of the technique is the minimal sample preparation needed, making ESEM quick to use and the images less susceptible to the artifacts that the extensive sample preparation usually required for CSEM may introduce. Careful manipulation of both the humidity in the microscope chamber and the beam energy are nevertheless essential to prevent dehydration and beam damage artifacts. In some circumstances it is possible to image live cells in the ESEM (2).In the following sections we introduce the fundamental principles of ESEM imaging before presenting imaging protocols for plant epidermis, mammalian cells, and bacteria. In the first two cases samples are imaged using the secondary electron (topographic) signal, whereas a transmission technique is employed to image bacteria.

High resolution low dose transmission electron microscopy real-time imaging and manipulation of nano-scale objects in the electron beam

DOEpatents

Brown, Jr., R. Malcolm; Barnes, Zack [Austin, TX; Sawatari, Chie [Shizuoka, JP; Kondo, Tetsuo [Kukuoka, JP

2008-02-26

The present invention includes a method, apparatus and system for nanofabrication in which one or more target molecules are identified for manipulation with an electron beam and the one or more target molecules are manipulated with the electron beam to produce new useful materials.

High-resolution quantitative determination of dielectric function by using scattering scanning near-field optical microscopy

PubMed Central

Tranca, D. E.; Stanciu, S. G.; Hristu, R.; Stoichita, C.; Tofail, S. A. M.; Stanciu, G. A.

2015-01-01

A new method for high-resolution quantitative measurement of the dielectric function by using scattering scanning near-field optical microscopy (s-SNOM) is presented. The method is based on a calibration procedure that uses the s-SNOM oscillating dipole model of the probe-sample interaction and quantitative s-SNOM measurements. The nanoscale capabilities of the method have the potential to enable novel applications in various fields such as nano-electronics, nano-photonics, biology or medicine. PMID:26138665

Molecular dynamics-based refinement and validation for sub-5 Å cryo-electron microscopy maps.

PubMed

Singharoy, Abhishek; Teo, Ivan; McGreevy, Ryan; Stone, John E; Zhao, Jianhua; Schulten, Klaus

2016-07-07

Two structure determination methods, based on the molecular dynamics flexible fitting (MDFF) paradigm, are presented that resolve sub-5 Å cryo-electron microscopy (EM) maps with either single structures or ensembles of such structures. The methods, denoted cascade MDFF and resolution exchange MDFF, sequentially re-refine a search model against a series of maps of progressively higher resolutions, which ends with the original experimental resolution. Application of sequential re-refinement enables MDFF to achieve a radius of convergence of ~25 Å demonstrated with the accurate modeling of β-galactosidase and TRPV1 proteins at 3.2 Å and 3.4 Å resolution, respectively. The MDFF refinements uniquely offer map-model validation and B-factor determination criteria based on the inherent dynamics of the macromolecules studied, captured by means of local root mean square fluctuations. The MDFF tools described are available to researchers through an easy-to-use and cost-effective cloud computing resource on Amazon Web Services.

Preparation of isolated nuclei from K 562 haemopoietic cell line for high resolution scanning electron microscopy.

PubMed

Reipert, S; Reipert, B M; Allen, T D

1994-09-01

The aim of the work is to visualise nuclear pore complexes (NPCs) in mammalian cells by high resolution scanning electron microscopy. A detergent-free isolation protocol was employed to obtain clean nuclei from the haemopoietic cell line K 562. Nuclear isolation was performed by mechanical homogenisation under hypotonic conditions followed by purification of the nuclear fraction. The isolated nuclei were attached to silicon chips, fixed, critical point dried, and sputter coated with a thin film (3-4 nm) of tantalum. Analysis of the nuclear surface by scanning electron microscopy (SEM) revealed a strong sensitivity of the outer nuclear membrane (ONM) to disruption during the isolation procedure. A significant reduction of the characteristic pattern of damage to the ONM was achieved by means of an isopicnic centrifugation on an isoosmolar balanced Percoll gradient. Analysis of the population of isolated nuclei by flow cytometry showed no signs of cell cycle specific losses of nuclei during isolation. The SEM investigations of the morphology of the nuclear envelope (NE) and of substructural details of NPCs and polyribosomes were performed using an in-lens field emission scanning electron microscope.

An introduction to optical super-resolution microscopy for the adventurous biologist

NASA Astrophysics Data System (ADS)

Vangindertael, J.; Camacho, R.; Sempels, W.; Mizuno, H.; Dedecker, P.; Janssen, K. P. F.

2018-04-01

Ever since the inception of light microscopy, the laws of physics have seemingly thwarted every attempt to visualize the processes of life at its most fundamental, sub-cellular, level. The diffraction limit has restricted our view to length scales well above 250 nm and in doing so, severely compromised our ability to gain true insights into many biological systems. Fortunately, continuous advancements in optics, electronics and mathematics have since provided the means to once again make physics work to our advantage. Even though some of the fundamental concepts enabling super-resolution light microscopy have been known for quite some time, practically feasible implementations have long remained elusive. It should therefore not come as a surprise that the 2014 Nobel Prize in Chemistry was awarded to the scientists who, each in their own way, contributed to transforming super-resolution microscopy from a technological tour de force to a staple of the biologist’s toolkit. By overcoming the diffraction barrier, light microscopy could once again be established as an indispensable tool in an age where the importance of understanding life at the molecular level cannot be overstated. This review strives to provide the aspiring life science researcher with an introduction to optical microscopy, starting from the fundamental concepts governing compound and fluorescent confocal microscopy to the current state-of-the-art of super-resolution microscopy techniques and their applications.

Utility of fluorescence microscopy in embryonic/fetal topographical analysis.

PubMed

Zucker, R M; Elstein, K H; Shuey, D L; Ebron-McCoy, M; Rogers, J M

1995-06-01

For topographical analysis of developing embryos, investigators typically rely on scanning electron microscopy (SEM) to provide the surface detail not attainable with light microscopy. SEM is an expensive and time-consuming technique, however, and the preparation procedure may alter morphology and leave the specimen friable. We report that by using a high-resolution compound epifluorescence microscope with inexpensive low-power objectives and the fluorochrome acridine orange, we were able to obtain surface images of fixed or fresh whole rat embryos and fetal palates of considerably greater topographical detail than those obtained using routine light microscopy. Indeed the resulting high-resolution images afford not only superior qualitative documentation of morphological observations, but the capability for detailed morphometry via digitization and computer-assisted image analysis.

Pump-probe Kelvin-probe force microscopy: Principle of operation and resolution limits

DOE Office of Scientific and Technical Information (OSTI.GOV)

Murawski, J.; Graupner, T.; Milde, P., E-mail: peter.milde@tu-dresden.de

Knowledge on surface potential dynamics is crucial for understanding the performance of modern-type nanoscale devices. We describe an electrical pump-probe approach in Kelvin-probe force microscopy that enables a quantitative measurement of dynamic surface potentials at nanosecond-time and nanometer-length scales. Also, we investigate the performance of pump-probe Kelvin-probe force microscopy with respect to the relevant experimental parameters. We exemplify a measurement on an organic field effect transistor that verifies the undisturbed functionality of our pump-probe approach in terms of simultaneous and quantitative mapping of topographic and electronic information at a high lateral and temporal resolution.

Determination of atomic-scale chemical composition at semiconductor heteroepitaxial interfaces by high-resolution transmission electron microscopy.

PubMed

Wen, C; Ma, Y J

2018-03-01

The determination of atomic structures and further quantitative information such as chemical compositions at atomic scale for semiconductor defects or heteroepitaxial interfaces can provide direct evidence to understand their formation, modification, and/or effects on the properties of semiconductor films. The commonly used method, high-resolution transmission electron microscopy (HRTEM), suffers from difficulty in acquiring images that correctly show the crystal structure at atomic resolution, because of the limitation in microscope resolution or deviation from the Scherzer-defocus conditions. In this study, an image processing method, image deconvolution, was used to achieve atomic-resolution (∼1.0 Å) structure images of small lattice-mismatch (∼1.0%) AlN/6H-SiC (0001) and large lattice-mismatch (∼8.5%) AlSb/GaAs (001) heteroepitaxial interfaces using simulated HRTEM images of a conventional 300-kV field-emission-gun transmission electron microscope under non-Scherzer-defocus conditions. Then, atomic-scale chemical compositions at the interface were determined for the atomic intermixing and Lomer dislocation with an atomic step by analyzing the deconvoluted image contrast. Furthermore, the effect of dynamical scattering on contrast analysis was also evaluated for differently weighted atomic columns in the compositions. Copyright © 2018 Elsevier Ltd. All rights reserved.

ATOMIC RESOLUTION CRYO ELECTRON MICROSCOPY OF MACROMOLECULAR COMPLEXES

PubMed Central

ZHOU, Z. HONG

2013-01-01

Single-particle cryo electron microscopy (cryoEM) is a technique for determining three-dimensional (3D) structures from projection images of molecular complexes preserved in their “native,” noncrystalline state. Recently, atomic or near-atomic resolution structures of several viruses and protein assemblies have been determined by single-particle cryoEM, allowing ab initio atomic model building by following the amino acid side chains or nucleic acid bases identifiable in their cryoEM density maps. In particular, these cryoEM structures have revealed extended arms contributing to molecular interactions that are otherwise not resolved by the conventional structural method of X-ray crystallography at similar resolutions. High-resolution cryoEM requires careful consideration of a number of factors, including proper sample preparation to ensure structural homogeneity, optimal configuration of electron imaging conditions to record high-resolution cryoEM images, accurate determination of image parameters to correct image distortions, efficient refinement and computation to reconstruct a 3D density map, and finally appropriate choice of modeling tools to construct atomic models for functional interpretation. This progress illustrates the power of cryoEM and ushers it into the arsenal of structural biology, alongside conventional techniques of X-ray crystallography and NMR, as a major tool (and sometimes the preferred one) for the studies of molecular interactions in supramolecular assemblies or machines. PMID:21501817

Far-field optical imaging with subdiffraction resolution enabled by nonlinear saturation absorption

NASA Astrophysics Data System (ADS)

Ding, Chenliang; Wei, Jingsong

2016-01-01

The resolution of far-field optical imaging is required to improve beyond the Abbe limit to the subdiffraction or even the nanoscale. In this work, inspired by scanning electronic microscopy (SEM) imaging, in which carbon (or Au) thin films are usually required to be coated on the sample surface before imaging to remove the charging effect while imaging by electrons. We propose a saturation-absorption-induced far-field super-resolution optical imaging method (SAI-SRIM). In the SAI-SRIM, the carbon (or Au) layers in SEM imaging are replaced by nonlinear-saturation-absorption (NSA) thin films, which are directly coated onto the sample surfaces using advanced thin film deposition techniques. The surface fluctuant morphologies are replicated to the NSA thin films, accordingly. The coated sample surfaces are then imaged using conventional laser scanning microscopy. Consequently, the imaging resolution is greatly improved, and subdiffraction-resolved optical images are obtained theoretically and experimentally. The SAI-SRIM provides an effective and easy way to achieve far-field super-resolution optical imaging for sample surfaces with geometric fluctuant morphology characteristics.

Optimized Deconvolution for Maximum Axial Resolution in Three-Dimensional Aberration-Corrected Scanning Transmission Electron Microscopy

PubMed Central

Ramachandra, Ranjan; de Jonge, Niels

2012-01-01

Three-dimensional (3D) data sets were recorded of gold nanoparticles placed on both sides of silicon nitride membranes using focal series aberration-corrected scanning transmission electron microscopy (STEM). The deconvolution of the 3D datasets was optimized to obtain the highest possible axial resolution. The deconvolution involved two different point spread function (PSF)s, each calculated iteratively via blind deconvolution.. Supporting membranes of different thicknesses were tested to study the effect of beam broadening on the deconvolution. It was found that several iterations of deconvolution was efficient in reducing the imaging noise. With an increasing number of iterations, the axial resolution was increased, and most of the structural information was preserved. Additional iterations improved the axial resolution by maximal a factor of 4 to 6, depending on the particular dataset, and up to 8 nm maximal, but at the cost of a reduction of the lateral size of the nanoparticles in the image. Thus, the deconvolution procedure optimized for highest axial resolution is best suited for applications where one is interested in the 3D locations of nanoparticles only. PMID:22152090

Integration of a high-NA light microscope in a scanning electron microscope.

PubMed

Zonnevylle, A C; Van Tol, R F C; Liv, N; Narvaez, A C; Effting, A P J; Kruit, P; Hoogenboom, J P

2013-10-01

We present an integrated light-electron microscope in which an inverted high-NA objective lens is positioned inside a scanning electron microscope (SEM). The SEM objective lens and the light objective lens have a common axis and focal plane, allowing high-resolution optical microscopy and scanning electron microscopy on the same area of a sample simultaneously. Components for light illumination and detection can be mounted outside the vacuum, enabling flexibility in the construction of the light microscope. The light objective lens can be positioned underneath the SEM objective lens during operation for sub-10 μm alignment of the fields of view of the light and electron microscopes. We demonstrate in situ epifluorescence microscopy in the SEM with a numerical aperture of 1.4 using vacuum-compatible immersion oil. For a 40-nm-diameter fluorescent polymer nanoparticle, an intensity profile with a FWHM of 380 nm is measured whereas the SEM performance is uncompromised. The integrated instrument may offer new possibilities for correlative light and electron microscopy in the life sciences as well as in physics and chemistry. © 2013 The Authors Journal of Microscopy © 2013 Royal Microscopical Society.

Shedding light on cell compartmentation in the candidate phylum Poribacteria by high resolution visualisation and transcriptional profiling

NASA Astrophysics Data System (ADS)

Jahn, Martin T.; Markert, Sebastian M.; Ryu, Taewoo; Ravasi, Timothy; Stigloher, Christian; Hentschel, Ute; Moitinho-Silva, Lucas

2016-10-01

Assigning functions to uncultivated environmental microorganisms continues to be a challenging endeavour. Here, we present a new microscopy protocol for fluorescence in situ hybridisation-correlative light and electron microscopy (FISH-CLEM) that enabled, to our knowledge for the first time, the identification of single cells within their complex microenvironment at electron microscopy resolution. Members of the candidate phylum Poribacteria, common and uncultivated symbionts of marine sponges, were used towards this goal. Cellular 3D reconstructions revealed bipolar, spherical granules of low electron density, which likely represent carbon reserves. Poribacterial activity profiles were retrieved from prokaryotic enriched sponge metatranscriptomes using simulation-based optimised mapping. We observed high transcriptional activity for proteins related to bacterial microcompartments (BMC) and we resolved their subcellular localisation by combining FISH-CLEM with immunohistochemistry (IHC) on ultra-thin sponge tissue sections. In terms of functional relevance, we propose that the BMC-A region may be involved in 1,2-propanediol degradation. The FISH-IHC-CLEM approach was proven an effective toolkit to combine -omics approaches with functional studies and it should be widely applicable in environmental microbiology.

Genetically targeted 3D visualisation of Drosophila neurons under Electron Microscopy and X-Ray Microscopy using miniSOG

PubMed Central

Ng, Julian; Browning, Alyssa; Lechner, Lorenz; Terada, Masako; Howard, Gillian; Jefferis, Gregory S. X. E.

2016-01-01

Large dimension, high-resolution imaging is important for neural circuit visualisation as neurons have both long- and short-range patterns: from axons and dendrites to the numerous synapses at terminal endings. Electron Microscopy (EM) is the favoured approach for synaptic resolution imaging but how such structures can be segmented from high-density images within large volume datasets remains challenging. Fluorescent probes are widely used to localise synapses, identify cell-types and in tracing studies. The equivalent EM approach would benefit visualising such labelled structures from within sub-cellular, cellular, tissue and neuroanatomical contexts. Here we developed genetically-encoded, electron-dense markers using miniSOG. We demonstrate their ability in 1) labelling cellular sub-compartments of genetically-targeted neurons, 2) generating contrast under different EM modalities, and 3) segmenting labelled structures from EM volumes using computer-assisted strategies. We also tested non-destructive X-ray imaging on whole Drosophila brains to evaluate contrast staining. This enabled us to target specific regions for EM volume acquisition. PMID:27958322

Correlating Fluorescence and High-Resolution Scanning Electron Microscopy (HRSEM) for the study of GABAA receptor clustering induced by inhibitory synaptic plasticity.

PubMed

Orlando, Marta; Ravasenga, Tiziana; Petrini, Enrica Maria; Falqui, Andrea; Marotta, Roberto; Barberis, Andrea

2017-10-23

Both excitatory and inhibitory synaptic contacts display activity dependent dynamic changes in their efficacy that are globally termed synaptic plasticity. Although the molecular mechanisms underlying glutamatergic synaptic plasticity have been extensively investigated and described, those responsible for inhibitory synaptic plasticity are only beginning to be unveiled. In this framework, the ultrastructural changes of the inhibitory synapses during plasticity have been poorly investigated. Here we combined confocal fluorescence microscopy (CFM) with high resolution scanning electron microscopy (HRSEM) to characterize the fine structural rearrangements of post-synaptic GABA A Receptors (GABA A Rs) at the nanometric scale during the induction of inhibitory long-term potentiation (iLTP). Additional electron tomography (ET) experiments on immunolabelled hippocampal neurons allowed the visualization of synaptic contacts and confirmed the reorganization of post-synaptic GABA A R clusters in response to chemical iLTP inducing protocol. Altogether, these approaches revealed that, following the induction of inhibitory synaptic potentiation, GABA A R clusters increase in size and number at the post-synaptic membrane with no other major structural changes of the pre- and post-synaptic elements.

The surface topography of the choroid plexus. Environmental, low and high vacuum scanning electron microscopy.

PubMed

Mestres, Pedro; Pütz, Norbert; Garcia Gómez de Las Heras, Soledad; García Poblete, Eduardo; Morguet, Andrea; Laue, Michael

2011-05-01

Environmental scanning electron microscopy (ESEM) allows the examination of hydrated and dried specimens without a conductive metal coating which could be advantageous in the imaging of biological and medical objects. The aim of this study was to assess the performance and benefits of wet-mode and low vacuum ESEM in comparison to high vacuum scanning electron microscopy (SEM) using the choroid plexus of chicken embryos as a model, an organ of the brain involved in the formation of cerebrospinal fluid in vertebrates. Specimens were fixed with or without heavy metals and examined directly or after critical point drying with or without metal coating. For wet mode ESEM freshly excised specimens without any pre-treatment were also examined. Conventional high vacuum SEM revealed the characteristic morphology of the choroid plexus cells at a high resolution and served as reference. With low vacuum ESEM of dried but uncoated samples the structure appeared well preserved but charging was a problem. It could be reduced by a short beam dwell time and averaging of images or by using the backscattered electron detector instead of the gaseous secondary electron detector. However, resolution was lower than with conventional SEM. Wet mode imaging was only possible with tissue that had been stabilized by fixation. Not all surface details (e.g. microvilli) could be visualized and other structures, like the cilia, were deformed. In summary, ESEM is an additional option for the imaging of bio-medical samples but it is problematic with regard to resolution and sample stability during imaging. Copyright © 2011 Elsevier GmbH. All rights reserved.

Creating a Structurally Realistic Finite Element Geometric Model of a Cardiomyocyte to Study the Role of Cellular Architecture in Cardiomyocyte Systems Biology.

PubMed

Rajagopal, Vijay; Bass, Gregory; Ghosh, Shouryadipta; Hunt, Hilary; Walker, Cameron; Hanssen, Eric; Crampin, Edmund; Soeller, Christian

2018-04-18

With the advent of three-dimensional (3D) imaging technologies such as electron tomography, serial-block-face scanning electron microscopy and confocal microscopy, the scientific community has unprecedented access to large datasets at sub-micrometer resolution that characterize the architectural remodeling that accompanies changes in cardiomyocyte function in health and disease. However, these datasets have been under-utilized for investigating the role of cellular architecture remodeling in cardiomyocyte function. The purpose of this protocol is to outline how to create an accurate finite element model of a cardiomyocyte using high resolution electron microscopy and confocal microscopy images. A detailed and accurate model of cellular architecture has significant potential to provide new insights into cardiomyocyte biology, more than experiments alone can garner. The power of this method lies in its ability to computationally fuse information from two disparate imaging modalities of cardiomyocyte ultrastructure to develop one unified and detailed model of the cardiomyocyte. This protocol outlines steps to integrate electron tomography and confocal microscopy images of adult male Wistar (name for a specific breed of albino rat) rat cardiomyocytes to develop a half-sarcomere finite element model of the cardiomyocyte. The procedure generates a 3D finite element model that contains an accurate, high-resolution depiction (on the order of ~35 nm) of the distribution of mitochondria, myofibrils and ryanodine receptor clusters that release the necessary calcium for cardiomyocyte contraction from the sarcoplasmic reticular network (SR) into the myofibril and cytosolic compartment. The model generated here as an illustration does not incorporate details of the transverse-tubule architecture or the sarcoplasmic reticular network and is therefore a minimal model of the cardiomyocyte. Nevertheless, the model can already be applied in simulation-based investigations into the role of cell structure in calcium signaling and mitochondrial bioenergetics, which is illustrated and discussed using two case studies that are presented following the detailed protocol.

Precise and economic FIB/SEM for CLEM: with 2 nm voxels through mitosis.

PubMed

Luckner, Manja; Wanner, Gerhard

2018-05-23

A portfolio is presented documenting economic, high-resolution correlative focused ion beam scanning electron microscopy (FIB/SEM) in routine, comprising: (i) the use of custom-labeled slides and coverslips, (ii) embedding of cells in thin, or ultra-thin resin layers for correlative light and electron microscopy (CLEM) and (iii) the claim to reach the highest resolution possible with FIB/SEM in xyz. Regions of interest (ROIs) defined in light microscope (LM), can be relocated quickly and precisely in SEM. As proof of principle, HeLa cells were investigated in 3D context at all stages of the cell cycle, documenting ultrastructural changes during mitosis: nuclear envelope breakdown and reassembly, Golgi degradation and reconstitution and the formation of the midzone and midbody.

Limiting factors in atomic resolution cryo electron microscopy: No simple tricks

PubMed Central

Zhang, Xing; Zhou, Z. Hong

2013-01-01

To bring cryo electron microscopy (cryoEM) of large biological complexes to atomic resolution, several factors – in both cryoEM image acquisition and 3D reconstruction – that may be neglected at low resolution become significantly limiting. Here we present thorough analyses of four limiting factors: (a) electron-beam tilt, (b) inaccurate determination of defocus values, (c) focus gradient through particles, and (d) particularly for large particles, dynamic (multiple) scattering of electrons. We also propose strategies to cope with these factors: (a) the divergence and direction tilt components of electron-beam tilt could be reduced by maintaining parallel illumination and by using a coma-free alignment procedure, respectively. Moreover, the effect of all beam tilt components, including spiral tilt, could be eliminated by use of a spherical aberration corrector. (b) More accurate measurement of defocus value could be obtained by imaging areas adjacent to the target area at high electron dose and by measuring the image shift induced by tilting the electron beam. (c) Each known Fourier coefficient in the Fourier transform of a cryoEM image is the sum of two Fourier coefficients of the 3D structure, one on each of two curved ‘characteristic surfaces’ in 3D Fourier space. We describe a simple model-based iterative method that could recover these two Fourier coefficients on the two characteristic surfaces. (d) The effect of dynamic scattering could be corrected by deconvolution of a transfer function. These analyses and our proposed strategies offer useful guidance for future experimental designs targeting atomic resolution cryoEM reconstruction. PMID:21627992

From 2D slices to 3D volumes: image based reconstruction and morphological characterization of hippocampal cells on charged and uncharged surfaces using FIB/SEM serial sectioning.

PubMed

Schmidt, Franziska; Kühbacher, Markus; Gross, Ulrich; Kyriakopoulos, Antonius; Schubert, Helmut; Zehbe, Rolf

2011-03-01

3D imaging at a subcellular resolution is a powerful tool in the life sciences to investigate cells and their interactions with native tissues or artificial objects. While a tomographic experimental setup achieving a sufficient structural resolution can be established with either X-rays or electrons, the use of electrons is usually limited to very thin samples in transmission electron microscopy due to the poor penetration depths of electrons. The combination of a serial sectioning approach and scanning electron microscopy in state of the art dual beam experimental setups therefore offers a means to image highly resolved spatial details using a focused ion beam for slicing and an electron beam for imaging. The advantage of this technique over X-ray μCT or X-ray microscopy attributes to the fact that absorption is not a limiting factor in imaging and therefore even strong absorbing structures can be spatially reconstructed with a much higher possible resolution. This approach was used in this study to elucidate the effect of an electric potential on the morphology of cells from a hippocampal cell line (HT22) deposited on gold microelectrodes. While cells cultivated on two different controls (gold and polymer substrates) did show the expected stretched morphology, cells on both the anode and the cathode differed significantly. Cells deposited on the anode part of the electrode exhibited the most extreme deviation, being almost spherical and showed signs of chromatin condensation possibly indicating cell death. Furthermore, EDX was used as supplemental methodology for combined chemical and structural analyses. Copyright © 2010 Elsevier B.V. All rights reserved.

Electron microscopy of whole cells in liquid with nanometer resolution

PubMed Central

de Jonge, N.; Peckys, D. B.; Kremers, G. J.; Piston, D. W.

2009-01-01

Single gold-tagged epidermal growth factor (EGF) molecules bound to cellular EGF receptors of fixed fibroblast cells were imaged in liquid with a scanning transmission electron microscope (STEM). The cells were placed in buffer solution in a microfluidic device with electron transparent windows inside the vacuum of the electron microscope. A spatial resolution of 4 nm and a pixel dwell time of 20 μs were obtained. The liquid layer was sufficiently thick to contain the cells with a thickness of 7 ± 1 μm. The experimental findings are consistent with a theoretical calculation. Liquid STEM is a unique approach for imaging single molecules in whole cells with significantly improved resolution and imaging speed over existing methods. PMID:19164524

Insights into radiation damage from atomic resolution scanning transmission electron microscopy imaging of mono-layer CuPcCl16 films on graphene.

PubMed

Mittelberger, Andreas; Kramberger, Christian; Meyer, Jannik C

2018-03-19

Atomically resolved images of monolayer organic crystals have only been obtained with scanning probe methods so far. On the one hand, they are usually prepared on surfaces of bulk materials, which are not accessible by (scanning) transmission electron microscopy. On the other hand, the critical electron dose of a monolayer organic crystal is orders of magnitudes lower than the one for bulk crystals, making (scanning) transmission electron microscopy characterization very challenging. In this work we present an atomically resolved study on the dynamics of a monolayer CuPcCl 16 crystal under the electron beam as well as an image of the undamaged molecules obtained by low-dose electron microscopy. The results show the dynamics and the radiation damage mechanisms in the 2D layer of this material, complementing what has been found for bulk crystals in earlier studies. Furthermore, being able to image the undamaged molecular crystal allows the characterization of new composites consisting of 2D materials and organic molecules.

Correlative Microscopy Combining Secondary Ion Mass Spectrometry and Electron Microscopy: Comparison of Intensity-Hue-Saturation and Laplacian Pyramid Methods for Image Fusion.

PubMed

Vollnhals, Florian; Audinot, Jean-Nicolas; Wirtz, Tom; Mercier-Bonin, Muriel; Fourquaux, Isabelle; Schroeppel, Birgit; Kraushaar, Udo; Lev-Ram, Varda; Ellisman, Mark H; Eswara, Santhana

2017-10-17

Correlative microscopy combining various imaging modalities offers powerful insights into obtaining a comprehensive understanding of physical, chemical, and biological phenomena. In this article, we investigate two approaches for image fusion in the context of combining the inherently lower-resolution chemical images obtained using secondary ion mass spectrometry (SIMS) with the high-resolution ultrastructural images obtained using electron microscopy (EM). We evaluate the image fusion methods with three different case studies selected to broadly represent the typical samples in life science research: (i) histology (unlabeled tissue), (ii) nanotoxicology, and (iii) metabolism (isotopically labeled tissue). We show that the intensity-hue-saturation fusion method often applied for EM-sharpening can result in serious image artifacts, especially in cases where different contrast mechanisms interplay. Here, we introduce and demonstrate Laplacian pyramid fusion as a powerful and more robust alternative method for image fusion. Both physical and technical aspects of correlative image overlay and image fusion specific to SIMS-based correlative microscopy are discussed in detail alongside the advantages, limitations, and the potential artifacts. Quantitative metrics to evaluate the results of image fusion are also discussed.

Three-dimensional nanoscale imaging by plasmonic Brownian microscopy

NASA Astrophysics Data System (ADS)

Labno, Anna; Gladden, Christopher; Kim, Jeongmin; Lu, Dylan; Yin, Xiaobo; Wang, Yuan; Liu, Zhaowei; Zhang, Xiang

2017-12-01

Three-dimensional (3D) imaging at the nanoscale is a key to understanding of nanomaterials and complex systems. While scanning probe microscopy (SPM) has been the workhorse of nanoscale metrology, its slow scanning speed by a single probe tip can limit the application of SPM to wide-field imaging of 3D complex nanostructures. Both electron microscopy and optical tomography allow 3D imaging, but are limited to the use in vacuum environment due to electron scattering and to optical resolution in micron scales, respectively. Here we demonstrate plasmonic Brownian microscopy (PBM) as a way to improve the imaging speed of SPM. Unlike photonic force microscopy where a single trapped particle is used for a serial scanning, PBM utilizes a massive number of plasmonic nanoparticles (NPs) under Brownian diffusion in solution to scan in parallel around the unlabeled sample object. The motion of NPs under an evanescent field is three-dimensionally localized to reconstruct the super-resolution topology of 3D dielectric objects. Our method allows high throughput imaging of complex 3D structures over a large field of view, even with internal structures such as cavities that cannot be accessed by conventional mechanical tips in SPM.

Nanometres-resolution Kikuchi patterns from materials science specimens with transmission electron forward scatter diffraction in the scanning electron microscope.

PubMed

Brodusch, N; Demers, H; Gauvin, R

2013-04-01

A charge-coupled device camera of an electron backscattered diffraction system in a scanning electron microscope was positioned below a thin specimen and transmission Kikuchi patterns were collected. Contrary to electron backscattered diffraction, transmission electron forward scatter diffraction provides phase identification and orientation mapping at the nanoscale. The minimum Pd particle size for which a Kikuchi diffraction pattern was detected and indexed reliably was 5.6 nm. An orientation mapping resolution of 5 nm was measured at 30 kV. The resolution obtained with transmission electron forward scatter diffraction was of the same order of magnitude than that reported in electron nanodiffraction in the transmission electron microscope. An energy dispersive spectrometer X-ray map and a transmission electron forward scatter diffraction orientation map were acquired simultaneously. The high-resolution chemical, phase and orientation maps provided at once information on the chemical form, orientation and coherency of precipitates in an aluminium-lithium 2099 alloy. © 2013 The Authors Journal of Microscopy © 2013 Royal Microscopical Society.

Probing the electronic transport on the reconstructed Au/Ge(001) surface

PubMed Central

Krok, Franciszek; Kaspers, Mark R; Bernhart, Alexander M; Nikiel, Marek; Jany, Benedykt R; Indyka, Paulina; Wojtaszek, Mateusz; Möller, Rolf

2014-01-01

Summary By using scanning tunnelling potentiometry we characterized the lateral variation of the electrochemical potential µec on the gold-induced Ge(001)-c(8 × 2)-Au surface reconstruction while a lateral current flows through the sample. On the reconstruction and across domain boundaries we find that µec shows a constant gradient as a function of the position between the contacts. In addition, nanoscale Au clusters on the surface do not show an electronic coupling to the gold-induced surface reconstruction. In combination with high resolution scanning electron microscopy and transmission electron microscopy, we conclude that an additional transport channel buried about 2 nm underneath the surface represents a major transport channel for electrons. PMID:25247129

High-resolution scanning electron microscopy of frozen-hydrated cells.

PubMed

Walther, P; Chen, Y; Pech, L L; Pawley, J B

1992-11-01

Cryo-fixed yeast Paramecia and sea urchin embryos were investigated with an in-lens type field-emission SEM using a cold stage. The goal was to further develop and investigate the processing of frozen samples for the low-temperature scanning electron microscope (LTSEM). Uncoated frozen-hydrated samples were imaged with the low-voltage backscattered electron signal (BSE). Resolution and contrast were sufficient to visualize cross-fractured membranes, nuclear pores and small vesicles in the cytoplasm. It is assumed that the resolution of this approach is limited by the extraction depth of the BSE which depends upon the accelerating voltage of the primary beam (V0). In this study, the lowest possible V0 was 2.6 kV because below this value the sensitivity of the BSE detector is insufficient. It is concluded that the resolution of the uncoated specimen could be improved if equipment were available for high-resolution BSE imaging at 0.5-2 kV. Higher resolution was obtained with platinum cryo-coated samples, on which intramembranous particles were easily imaged. These images even show the ring-like appearance of the hexagonally arranged intramembranous particles known from high-resolution replica studies. On fully hydrated samples at high magnification, the observation time for a particular area is limited by mass loss caused by electron irradiation. Other potential sources of artefacts are the deposition of water vapour contamination and shrinkage caused by the sublimation of ice. Imaging of partially dehydrated (partially freeze-dried) samples, e.g. high-pressure frozen Paramecium and sea urchin embryos, will probably become the main application in cell biology. In spite of possible shrinkage problems, this approach has a number of advantages compared with any other electron microscopy preparation method: no chemical fixation is necessary, eliminating this source of artefacts; due to partial removal of the water additional structures in the cytoplasm can be investigated; and finally, the mass loss due to electron beam irradiation is greatly reduced compared to fully frozen-hydrated specimens.

High-resolution transmission electron microscopy of hexagonal and rhombohedral molybdenum disulfide crystals.

PubMed

Isshiki, T; Nishio, K; Saijo, H; Shiojiri, M; Yabuuchi, Y; Takahashi, N

1993-07-01

Natural (molybdenite) and synthesized molybdenum disulfide crystals have been studied by high-resolution transmission electron microscopy. The image simulation demonstrates that the [0001] and [0110] HRTEM images of hexagonal and rhombohedral MoS2 crystals hardly disclose their stacking sequences, and that the [2110] images can distinguish the Mo and S columns along the incident electron beam and enable one to determine not only the crystal structure but also the fault structure. Observed [0001] images of cleaved molybdenite and synthesized MoS2 crystals, however, reveal the strain field around partial dislocations limiting an extended dislocation. A cross-sectional image of a single molecular (S-Mo-S) layer cleaved from molybdenite has been observed. Synthesized MoS2 flakes which were prepared by grinding have been found to be rhombohedral crystals containing many stacking faults caused by glides between S/S layers.

3D imaging by serial block face scanning electron microscopy for materials science using ultramicrotomy.

PubMed

Hashimoto, Teruo; Thompson, George E; Zhou, Xiaorong; Withers, Philip J

2016-04-01

Mechanical serial block face scanning electron microscopy (SBFSEM) has emerged as a means of obtaining three dimensional (3D) electron images over volumes much larger than possible by focused ion beam (FIB) serial sectioning and at higher spatial resolution than achievable with conventional X-ray computed tomography (CT). Such high resolution 3D electron images can be employed for precisely determining the shape, volume fraction, distribution and connectivity of important microstructural features. While soft (fixed or frozen) biological samples are particularly well suited for nanoscale sectioning using an ultramicrotome, the technique can also produce excellent 3D images at electron microscope resolution in a time and resource-efficient manner for engineering materials. Currently, a lack of appreciation of the capabilities of ultramicrotomy and the operational challenges associated with minimising artefacts for different materials is limiting its wider application to engineering materials. Consequently, this paper outlines the current state of the art for SBFSEM examining in detail how damage is introduced during slicing and highlighting strategies for minimising such damage. A particular focus of the study is the acquisition of 3D images for a variety of metallic and coated systems. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Atomic scale imaging of magnetic circular dichroism by achromatic electron microscopy.

PubMed

Wang, Zechao; Tavabi, Amir H; Jin, Lei; Rusz, Ján; Tyutyunnikov, Dmitry; Jiang, Hanbo; Moritomo, Yutaka; Mayer, Joachim; Dunin-Borkowski, Rafal E; Yu, Rong; Zhu, Jing; Zhong, Xiaoyan

2018-03-01

In order to obtain a fundamental understanding of the interplay between charge, spin, orbital and lattice degrees of freedom in magnetic materials and to predict and control their physical properties 1-3 , experimental techniques are required that are capable of accessing local magnetic information with atomic-scale spatial resolution. Here, we show that a combination of electron energy-loss magnetic chiral dichroism 4 and chromatic-aberration-corrected transmission electron microscopy, which reduces the focal spread of inelastically scattered electrons by orders of magnitude when compared with the use of spherical aberration correction alone, can achieve atomic-scale imaging of magnetic circular dichroism and provide element-selective orbital and spin magnetic moments atomic plane by atomic plane. This unique capability, which we demonstrate for Sr 2 FeMoO 6 , opens the door to local atomic-level studies of spin configurations in a multitude of materials that exhibit different types of magnetic coupling, thereby contributing to a detailed understanding of the physical origins of magnetic properties of materials at the highest spatial resolution.

Low cost, high performance processing of single particle cryo-electron microscopy data in the cloud.

PubMed

Cianfrocco, Michael A; Leschziner, Andres E

2015-05-08

The advent of a new generation of electron microscopes and direct electron detectors has realized the potential of single particle cryo-electron microscopy (cryo-EM) as a technique to generate high-resolution structures. Calculating these structures requires high performance computing clusters, a resource that may be limiting to many likely cryo-EM users. To address this limitation and facilitate the spread of cryo-EM, we developed a publicly available 'off-the-shelf' computing environment on Amazon's elastic cloud computing infrastructure. This environment provides users with single particle cryo-EM software packages and the ability to create computing clusters with 16-480+ CPUs. We tested our computing environment using a publicly available 80S yeast ribosome dataset and estimate that laboratories could determine high-resolution cryo-EM structures for $50 to $1500 per structure within a timeframe comparable to local clusters. Our analysis shows that Amazon's cloud computing environment may offer a viable computing environment for cryo-EM.

Application of two-dimensional crystallography and image processing to atomic resolution Z-contrast images.

PubMed

Morgan, David G; Ramasse, Quentin M; Browning, Nigel D

2009-06-01

Zone axis images recorded using high-angle annular dark-field scanning transmission electron microscopy (HAADF-STEM or Z-contrast imaging) reveal the atomic structure with a resolution that is defined by the probe size of the microscope. In most cases, the full images contain many sub-images of the crystal unit cell and/or interface structure. Thanks to the repetitive nature of these images, it is possible to apply standard image processing techniques that have been developed for the electron crystallography of biological macromolecules and have been used widely in other fields of electron microscopy for both organic and inorganic materials. These methods can be used to enhance the signal-to-noise present in the original images, to remove distortions in the images that arise from either the instrumentation or the specimen itself and to quantify properties of the material in ways that are difficult without such data processing. In this paper, we describe briefly the theory behind these image processing techniques and demonstrate them for aberration-corrected, high-resolution HAADF-STEM images of Si(46) clathrates developed for hydrogen storage.

Evidence for anisotropic dielectric properties of monoclinic hafnia using valence electron energy-loss spectroscopy in high-resolution transmission electron microscopy and ab initio time-dependent density-functional theory

DOE Office of Scientific and Technical Information (OSTI.GOV)

Guedj, C.; CEA, LETI, MINATEC Campus, F-38054 Grenoble; Hung, L.

2014-12-01

The effect of nanocrystal orientation on the energy loss spectra of monoclinic hafnia (m-HfO{sub 2}) is measured by high resolution transmission electron microscopy (HRTEM) and valence energy loss spectroscopy (VEELS) on high quality samples. For the same momentum-transfer directions, the dielectric properties are also calculated ab initio by time-dependent density-functional theory (TDDFT). Experiments and simulations evidence anisotropy in the dielectric properties of m-HfO{sub 2}, most notably with the direction-dependent oscillator strength of the main bulk plasmon. The anisotropic nature of m-HfO{sub 2} may contribute to the differences among VEELS spectra reported in literature. The good agreement between the complex dielectricmore » permittivity extracted from VEELS with nanometer spatial resolution, TDDFT modeling, and past literature demonstrates that the present HRTEM-VEELS device-oriented methodology is a possible solution to the difficult nanocharacterization challenges given in the International Technology Roadmap for Semiconductors.« less

Optical gating and streaking of free electrons with sub-optical cycle precision

PubMed Central

Kozák, M.; McNeur, J.; Leedle, K. J.; Deng, H.; Schönenberger, N.; Ruehl, A.; Hartl, I.; Harris, J. S.; Byer, R. L.; Hommelhoff, P.

2017-01-01

The temporal resolution of ultrafast electron diffraction and microscopy experiments is currently limited by the available experimental techniques for the generation and characterization of electron bunches with single femtosecond or attosecond durations. Here, we present proof of principle experiments of an optical gating concept for free electrons via direct time-domain visualization of the sub-optical cycle energy and transverse momentum structure imprinted on the electron beam. We demonstrate a temporal resolution of 1.2±0.3 fs. The scheme is based on the synchronous interaction between electrons and the near-field mode of a dielectric nano-grating excited by a femtosecond laser pulse with an optical period duration of 6.5 fs. The sub-optical cycle resolution demonstrated here is promising for use in laser-driven streak cameras for attosecond temporal characterization of bunched particle beams as well as time-resolved experiments with free-electron beams. PMID:28120930

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kendall, Amy; Bian, Wen; Maris, Alexander

We have used fiber diffraction, cryo-electron microscopy, and scanning transmission electron microscopy to confirm the symmetry of three potexviruses, potato virus X, papaya mosaic virus, and narcissus mosaic virus, and to determine their low-resolution structures. All three viruses have slightly less than nine subunits per turn of the viral helix. Our data strongly support the view that all potexviruses have approximately the same symmetry. The structures are dominated by a large domain at high radius in the virion, with a smaller domain, which includes the putative RNA-binding site, extending to low radius.

Epitaxy of GaN in high aspect ratio nanoscale holes over silicon substrate

NASA Astrophysics Data System (ADS)

Wang, Kejia; Wang, Anqi; Ji, Qingbin; Hu, Xiaodong; Xie, Yahong; Sun, Ying; Cheng, Zhiyuan

2017-12-01

Dislocation filtering in gallium nitride (GaN) by epitaxial growth through patterned nanoscale holes is studied. GaN grown from extremely high aspect ratio holes by metalorganic chemical vapor deposition is examined by transmission electron microscopy and high-resolution transmission electron microscopy. This selective area epitaxial growth method with a reduced epitaxy area and an increased depth to width ratio of holes leads to effective filtering of dislocations within the hole and improves the quality of GaN significantly.

Electrical conductivity and morphology of electrochemical synthesized polyaniline/CuO nano composites

NASA Astrophysics Data System (ADS)

Ashokkumar, S. P.; Yesappa, L.; Vijeth, H.; Niranjana, M.; Devendrappa, H.

2018-05-01

Polyaniline (PANI) and Polyaniline/CuO nanocomposite have been synthesized by using electrochemical deposition method. The composite was characterized using Fourier transform infra-red spectroscopy (FT-IR) to confirm the chemical interaction changes, micro structural morphology was done by Field Emission Scanning Electronic Microscopy (FESEM) and High Resolution Transmission Electron Microscopy (HRTEM). The dielectric constant and AC conductivity are found to increases with increase in temperature range (303 to 393K), these results shows enhancement in electrical conductivity due to effect of nanocomposite.

Novel Crystalline SiO2 Nanoparticles via Annelids Bioprocessing of Agro-Industrial Wastes

PubMed Central

2010-01-01

The synthesis of nanoparticles silica oxide from rice husk, sugar cane bagasse and coffee husk, by employing vermicompost with annelids (Eisenia foetida) is reported. The product (humus) is calcinated and extracted to recover the crystalline nanoparticles. X-ray diffraction (XRD), transmission electron microscopy (TEM), high-resolution transmission electron microscopy (HRTEM) and dynamic light scattering (DLS) show that the biotransformation allows creating specific crystalline phases, since equivalent particles synthesized without biotransformation are bigger and with different crystalline structure. PMID:20802789

Analysis of Multilayer Devices for Superconducting Electronics by High-Resolution Scanning Transmission Electron Microscopy and Energy Dispersive Spectroscopy

DOE PAGES

Missert, Nancy; Kotula, Paul G.; Rye, Michael; ...

2017-02-15

We used a focused ion beam to obtain cross-sectional specimens from both magnetic multilayer and Nb/Al-AlOx/Nb Josephson junction devices for characterization by scanning transmission electron microscopy (STEM) and energy dispersive X-ray spectroscopy (EDX). An automated multivariate statistical analysis of the EDX spectral images produced chemically unique component images of individual layers within the multilayer structures. STEM imaging elucidated distinct variations in film morphology, interface quality, and/or etch artifacts that could be correlated to magnetic and/or electrical properties measured on the same devices.

Cryo-Scanning Electron Microscopy (SEM) and Scanning Transmission Electron Microscopy (STEM)-in-SEM for Bio- and Organo-Mineral Interface Characterization in the Environment.

PubMed

Wille, Guillaume; Hellal, Jennifer; Ollivier, Patrick; Richard, Annie; Burel, Agnes; Jolly, Louis; Crampon, Marc; Michel, Caroline

2017-12-01

Understanding biofilm interactions with surrounding substratum and pollutants/particles can benefit from the application of existing microscopy tools. Using the example of biofilm interactions with zero-valent iron nanoparticles (nZVI), this study aims to apply various approaches in biofilm preparation and labeling for fluorescent or electron microscopy and energy dispersive X-ray spectrometry (EDS) microanalysis for accurate observations. According to the targeted microscopy method, biofilms were sampled as flocs or attached biofilm, submitted to labeling using 4',6-diamidino-2-phenylindol, lectins PNA and ConA coupled to fluorescent dye or gold nanoparticles, and prepared for observation (fixation, cross-section, freezing, ultramicrotomy). Fluorescent microscopy revealed that nZVI were embedded in the biofilm structure as aggregates but the resolution was insufficient to observe individual nZVI. Cryo-scanning electron microscopy (SEM) observations showed nZVI aggregates close to bacteria, but it was not possible to confirm direct interactions between nZVI and cell membranes. Scanning transmission electron microscopy in the SEM (STEM-in-SEM) showed that nZVI aggregates could enter the biofilm to a depth of 7-11 µm. Bacteria were surrounded by a ring of extracellular polymeric substances (EPS) preventing direct nZVI/membrane interactions. STEM/EDS mapping revealed a co-localization of nZVI aggregates with lectins suggesting a potential role of EPS in nZVI embedding. Thus, the combination of divergent microscopy approaches is a good approach to better understand and characterize biofilm/metal interactions.

Whole-cell imaging of the budding yeast Saccharomyces cerevisiae by high-voltage scanning transmission electron tomography.

PubMed

Murata, Kazuyoshi; Esaki, Masatoshi; Ogura, Teru; Arai, Shigeo; Yamamoto, Yuta; Tanaka, Nobuo

2014-11-01

Electron tomography using a high-voltage electron microscope (HVEM) provides three-dimensional information about cellular components in sections thicker than 1 μm, although in bright-field mode image degradation caused by multiple inelastic scattering of transmitted electrons limit the attainable resolution. Scanning transmission electron microscopy (STEM) is believed to give enhanced contrast and resolution compared to conventional transmission electron microscopy (CTEM). Samples up to 1 μm in thickness have been analyzed with an intermediate-voltage electron microscope because inelastic scattering is not a critical limitation, and probe broadening can be minimized. Here, we employed STEM at 1 MeV high-voltage to extend the useful specimen thickness for electron tomography, which we demonstrate by a seamless tomographic reconstruction of a whole, budding Saccharomyces cerevisiae yeast cell, which is ~3 μm in thickness. High-voltage STEM tomography, especially in the bright-field mode, demonstrated sufficiently enhanced contrast and intensity, compared to CTEM tomography, to permit segmentation of major organelles in the whole cell. STEM imaging also reduced specimen shrinkage during tilt-series acquisition. The fidelity of structural preservation was limited by cytoplasmic extraction, and the spatial resolution was limited by the relatively large convergence angle of the scanning probe. However, the new technique has potential to solve longstanding problems of image blurring in biological specimens beyond 1 μm in thickness, and may facilitate new research in cellular structural biology. Copyright © 2014 Elsevier B.V. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Deng, Junjing; Vine, David J.; Chen, Si

X-ray microscopy can be used to image whole, unsectioned cells in their native hydrated state. It complements the higher resolution of electron microscopy for submicrometer thick specimens, and the molecule-specific imaging capabilites of fluorescence light microscopy. We describe here the first use of fast, continuous x-ray scanning of frozen hydrated cells for simultaneous sub-20 nm resolution ptychographic transmission imaging with high contrast, and sub-100 nm resolution deconvolved x-ray fluorescence imaging of diffusible and bound ions at native concentrations, without the need to add specific labels. Here, by working with cells that have been rapidly frozen without the use of chemicalmore » fixatives, and imaging them under cryogenic conditions, we are able to obtain images with well preserved structural and chemical composition, and sufficient stability against radiation damage to allow for multiple images to be obtained with no observable change.« less

Crystallographic features related to a van der Waals coupling in the layered chalcogenide FePS{sub 3}

DOE Office of Scientific and Technical Information (OSTI.GOV)

Murayama, Chisato; Okabe, Momoko; Fukuda, Koichiro

We investigated the crystallographic structure of FePS{sub 3} with a layered structure using transmission electron microscopy and powder X-ray diffraction. We found that FePS{sub 3} forms a rotational twin structure with the common axis along the c*-axis. The high-resolution transmission electron microscopy images revealed that the twin boundaries were positioned at the van der Waals gaps between the layers. The narrow bands of dark contrast were observed in the bright-field transmission electron microscopy images below the antiferromagnetic transition temperature, T{sub N} ≈ 120 K. Low-temperature X-ray diffraction showed a lattice distortion; the a- and b-axes shortened and lengthened, respectively, as the temperature decreasedmore » below T{sub N.} We propose that the narrow bands of dark contrast observed in the bright-field transmission electron microscopy images are caused by the directional lattice distortion with respect to each micro-twin variant in the antiferromagnetic phase.« less

High-resolution high-sensitivity elemental imaging by secondary ion mass spectrometry: from traditional 2D and 3D imaging to correlative microscopy

NASA Astrophysics Data System (ADS)

Wirtz, T.; Philipp, P.; Audinot, J.-N.; Dowsett, D.; Eswara, S.

2015-10-01

Secondary ion mass spectrometry (SIMS) constitutes an extremely sensitive technique for imaging surfaces in 2D and 3D. Apart from its excellent sensitivity and high lateral resolution (50 nm on state-of-the-art SIMS instruments), advantages of SIMS include high dynamic range and the ability to differentiate between isotopes. This paper first reviews the underlying principles of SIMS as well as the performance and applications of 2D and 3D SIMS elemental imaging. The prospects for further improving the capabilities of SIMS imaging are discussed. The lateral resolution in SIMS imaging when using the microprobe mode is limited by (i) the ion probe size, which is dependent on the brightness of the primary ion source, the quality of the optics of the primary ion column and the electric fields in the near sample region used to extract secondary ions; (ii) the sensitivity of the analysis as a reasonable secondary ion signal, which must be detected from very tiny voxel sizes and thus from a very limited number of sputtered atoms; and (iii) the physical dimensions of the collision cascade determining the origin of the sputtered ions with respect to the impact site of the incident primary ion probe. One interesting prospect is the use of SIMS-based correlative microscopy. In this approach SIMS is combined with various high-resolution microscopy techniques, so that elemental/chemical information at the highest sensitivity can be obtained with SIMS, while excellent spatial resolution is provided by overlaying the SIMS images with high-resolution images obtained by these microscopy techniques. Examples of this approach are given by presenting in situ combinations of SIMS with transmission electron microscopy (TEM), helium ion microscopy (HIM) and scanning probe microscopy (SPM).

Nanopore fabrication and characterization by helium ion microscopy

NASA Astrophysics Data System (ADS)

Emmrich, D.; Beyer, A.; Nadzeyka, A.; Bauerdick, S.; Meyer, J. C.; Kotakoski, J.; Gölzhäuser, A.

2016-04-01

The Helium Ion Microscope (HIM) has the capability to image small features with a resolution down to 0.35 nm due to its highly focused gas field ionization source and its small beam-sample interaction volume. In this work, the focused helium ion beam of a HIM is utilized to create nanopores with diameters down to 1.3 nm. It will be demonstrated that nanopores can be milled into silicon nitride, carbon nanomembranes, and graphene with well-defined aspect ratio. To image and characterize the produced nanopores, helium ion microscopy and high resolution scanning transmission electron microscopy were used. The analysis of the nanopores' growth behavior allows inferring on the profile of the helium ion beam.

An Unroofing Method to Observe the Cytoskeleton Directly at Molecular Resolution Using Atomic Force Microscopy

PubMed Central

Usukura, Eiji; Narita, Akihiro; Yagi, Akira; Ito, Shuichi; Usukura, Jiro

2016-01-01

An improved unroofing method enabled the cantilever of an atomic force microscope (AFM) to reach directly into a cell to visualize the intracellular cytoskeletal actin filaments, microtubules, clathrin coats, and caveolae in phosphate-buffered saline (PBS) at a higher resolution than conventional electron microscopy. All of the actin filaments clearly exhibited a short periodicity of approximately 5–6 nm, which was derived from globular actins linked to each other to form filaments, as well as a long helical periodicity. The polarity of the actin filaments appeared to be determined by the shape of the periodic striations. Microtubules were identified based on their thickness. Clathrin coats and caveolae were observed on the cytoplasmic surface of cell membranes. The area containing clathrin molecules and their terminal domains was directly visualized. Characteristic ridge structures located at the surface of the caveolae were observed at high resolution, similar to those observed with electron microscopy (EM). Overall, unroofing allowed intracellular AFM imaging in a liquid environment with a level of quality equivalent or superior to that of EM. Thus, AFMs are anticipated to provide cutting-edge findings in cell biology and histology. PMID:27273367

High-resolution, high-throughput imaging with a multibeam scanning electron microscope.

PubMed

Eberle, A L; Mikula, S; Schalek, R; Lichtman, J; Knothe Tate, M L; Zeidler, D

2015-08-01

Electron-electron interactions and detector bandwidth limit the maximal imaging speed of single-beam scanning electron microscopes. We use multiple electron beams in a single column and detect secondary electrons in parallel to increase the imaging speed by close to two orders of magnitude and demonstrate imaging for a variety of samples ranging from biological brain tissue to semiconductor wafers. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

Measurement of replication structures at the nanometer scale using super-resolution light microscopy

PubMed Central

Baddeley, D.; Chagin, V. O.; Schermelleh, L.; Martin, S.; Pombo, A.; Carlton, P. M.; Gahl, A.; Domaing, P.; Birk, U.; Leonhardt, H.; Cremer, C.; Cardoso, M. C.

2010-01-01

DNA replication, similar to other cellular processes, occurs within dynamic macromolecular structures. Any comprehensive understanding ultimately requires quantitative data to establish and test models of genome duplication. We used two different super-resolution light microscopy techniques to directly measure and compare the size and numbers of replication foci in mammalian cells. This analysis showed that replication foci vary in size from 210 nm down to 40 nm. Remarkably, spatially modulated illumination (SMI) and 3D-structured illumination microscopy (3D-SIM) both showed an average size of 125 nm that was conserved throughout S-phase and independent of the labeling method, suggesting a basic unit of genome duplication. Interestingly, the improved optical 3D resolution identified 3- to 5-fold more distinct replication foci than previously reported. These results show that optical nanoscopy techniques enable accurate measurements of cellular structures at a level previously achieved only by electron microscopy and highlight the possibility of high-throughput, multispectral 3D analyses. PMID:19864256

Super-resolution photoacoustic microscopy using a localized near-field of a plasmonic nanoaperture: a three-dimensional simulation study

NASA Astrophysics Data System (ADS)

Park, Byullee; Lee, Hongki; Upputuri, Paul Kumar; Pramanik, Manojit; Kim, Donghyun; Kim, Chulhong

2018-02-01

Super-resolution microscopy has been increasingly important to delineate nanoscale biological structures or nanoparticles. With these increasing demands, several imaging modalities, including super-resolution fluorescence microscope (SRFM) and electron microscope (EM), have been developed and commercialized. These modalities achieve nanoscale resolution, however, SRFM cannot image without fluorescence, and sample preparation of EM is not suitable for biological specimens. To overcome those disadvantages, we have numerically studied the possibility of superresolution photoacoustic microscopy (SR-PAM) based on near-field localization of light. Photoacoustic (PA) signal is generally acquired based on optical absorption contrast; thus it requires no agents or pre-processing for the samples. The lateral resolution of the conventional photoacoustic microscopy is limited to 200 nm by diffraction limit, therefore reducing the lateral resolution is a major research impetus. Our approach to breaking resolution limit is to use laser pulses of extremely small spot size as a light source. In this research, we simulated the PA signal by constructing the three dimensional SR-PAM system environment using the k-Wave toolbox. As the light source, we simulated ultrashort light pulses using geometrical nanoaperture with near-field localization of surface plasmons. Through the PA simulation, we have successfully distinguish cuboids spaced 3 nm apart. In the near future, we will develop the SR-PAM and it will contribute to biomedical and material sciences.

ultraLM and miniLM: Locator tools for smart tracking of fluorescent cells in correlative light and electron microscopy.

PubMed

Brama, Elisabeth; Peddie, Christopher J; Wilkes, Gary; Gu, Yan; Collinson, Lucy M; Jones, Martin L

2016-12-13

In-resin fluorescence (IRF) protocols preserve fluorescent proteins in resin-embedded cells and tissues for correlative light and electron microscopy, aiding interpretation of macromolecular function within the complex cellular landscape. Dual-contrast IRF samples can be imaged in separate fluorescence and electron microscopes, or in dual-modality integrated microscopes for high resolution correlation of fluorophore to organelle. IRF samples also offer a unique opportunity to automate correlative imaging workflows. Here we present two new locator tools for finding and following fluorescent cells in IRF blocks, enabling future automation of correlative imaging. The ultraLM is a fluorescence microscope that integrates with an ultramicrotome, which enables 'smart collection' of ultrathin sections containing fluorescent cells or tissues for subsequent transmission electron microscopy or array tomography. The miniLM is a fluorescence microscope that integrates with serial block face scanning electron microscopes, which enables 'smart tracking' of fluorescent structures during automated serial electron image acquisition from large cell and tissue volumes.

Fitting Multimeric Protein Complexes into Electron Microscopy Maps Using 3D Zernike Descriptors

PubMed Central

Esquivel-Rodríguez, Juan; Kihara, Daisuke

2012-01-01

A novel computational method for fitting high-resolution structures of multiple proteins into a cryoelectron microscopy map is presented. The method named EMLZerD generates a pool of candidate multiple protein docking conformations of component proteins, which are later compared with a provided electron microscopy (EM) density map to select the ones that fit well into the EM map. The comparison of docking conformations and the EM map is performed using the 3D Zernike descriptor (3DZD), a mathematical series expansion of three-dimensional functions. The 3DZD provides a unified representation of the surface shape of multimeric protein complex models and EM maps, which allows a convenient, fast quantitative comparison of the three dimensional structural data. Out of 19 multimeric complexes tested, near native complex structures with a root mean square deviation of less than 2.5 Å were obtained for 14 cases while medium range resolution structures with correct topology were computed for the additional 5 cases. PMID:22417139

Fitting multimeric protein complexes into electron microscopy maps using 3D Zernike descriptors.

PubMed

Esquivel-Rodríguez, Juan; Kihara, Daisuke

2012-06-14

A novel computational method for fitting high-resolution structures of multiple proteins into a cryoelectron microscopy map is presented. The method named EMLZerD generates a pool of candidate multiple protein docking conformations of component proteins, which are later compared with a provided electron microscopy (EM) density map to select the ones that fit well into the EM map. The comparison of docking conformations and the EM map is performed using the 3D Zernike descriptor (3DZD), a mathematical series expansion of three-dimensional functions. The 3DZD provides a unified representation of the surface shape of multimeric protein complex models and EM maps, which allows a convenient, fast quantitative comparison of the three-dimensional structural data. Out of 19 multimeric complexes tested, near native complex structures with a root-mean-square deviation of less than 2.5 Å were obtained for 14 cases while medium range resolution structures with correct topology were computed for the additional 5 cases.

Direct Investigation of Mg Intercalation into the Orthorhombic V 2O 5 Cathode Using Atomic-Resolution Transmission Electron Microscopy [Direct Investigation of Mg intercalation into orthorhombic V 2O 5 cathode using Atomic Resolution Electron Microscopy Methods

DOE PAGES

Mukherjee, Arijita; Sa, Niya; Phillips, Patrick J.; ...

2017-02-13

Batteries based on Mg metal anode can promise much higher specific volumetric capacity and energy density compared to Li-ion systems and are, at the same time, safer and more cost-effective. While previous experimental reports have claimed reversible Mg intercalation into beyond Chevrel phase cathodes, they provide limited evidence of true Mg intercalation other than electrochemical data. Transmission electron microscopy techniques provide unique capabilities to directly image Mg intercalation and quantify the redox reaction within the cathode material. Here, we present a systematic study of Mg insertion into orthorhombic V 2O 5, combining aberration-corrected scanning transmission electron microscopy (STEM) imaging, electronmore » energy-loss spectroscopy (EELS), and energy-dispersive X-ray spectroscopy (EDX) analysis. We compare the results from an electrochemically cycled V 2O 5 cathode in a prospective full cell with Mg metal anode with a chemically synthesized MgV 2O 5 sample. Results suggest that the electrochemically cycled orthorhombic V 2O 5 cathode shows a local formation of the theoretically predicted ϵ-Mg0.5V2O5 phase; however, the intercalation levels of Mg are lower than predicted. Lastly, this phase is different from the chemically synthesized sample, which is found to represent the δ-MgV 2O 5 phase.« less

TEM studies of plasma nitrided austenitic stainless steel.

PubMed

Stróz, D; Psoda, M

2010-03-01

Cross-sectional transmission electron microscopy and X-ray phase analysis were used to study the structure of a layer formed during nitriding the AISI 316L stainless steel at temperature 440 degrees C. It was found that the applied treatment led to the formation of 6-microm-thick layer of the S-phase. There is no evidence of CrN precipitation. The X-ray diffraction experiments proved that the occurred austenite lattice expansion - due to nitrogen atoms - depended on the crystallographic direction. The cross-sectional transmission electron microscopy studies showed that the layer consisted of a single cubic phase that contained a lot of defects such as dislocations, stacking faults, slip bands and twins. The high-resolution electron microscopy observations were applied to study the defect formation due to the nitriding process. It was shown that the presence of great number of stacking faults leads to formation of nanotwins. Weak, forbidden {100} reflections were still another characteristic feature of the S-phase. These were not detected in the X-ray spectra of the phase. Basing on the high-resolution electron microscopy studies it can be suggested that the short-range ordering of the nitrogen atoms in the octahedral sites inside the f.c.c. matrix lattice takes place and gives rise to appearance of these spots. It is suggested that the cubic lattice undergoes not only expansion but also slight rombohedral distortion that explains differences in the lattice expansion for different crystallographic directions.

Contrast enhancement of biological nanoporous materials with zinc oxide infiltration for electron and X-ray nanoscale microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ocola, L. E.; Sampathkumar, V.; Kasthuri, N.

Here, we show that using infiltration of ZnO metal oxide can be useful for high resolution imaging of biological samples in electron and X-ray microscopy. This method is compatible with standard fixation techniques that leave the sample dry, such as finishing with super critical CO 2 drying, or simple vacuum drying at 95°C. We demonstrate this technique can be applied on tooth and brain tissue samples. We also show that high resolution X-ray tomography can be performed on biological systems using Zn K edge (1s) absorption to enhance internal structures, and obtained the first nanoscale 10 KeV X-ray absorption imagesmore » of the interior regions of a tooth.« less

Tools for Model Building and Optimization into Near-Atomic Resolution Electron Cryo-Microscopy Density Maps.

PubMed

DiMaio, F; Chiu, W

2016-01-01

Electron cryo-microscopy (cryoEM) has advanced dramatically to become a viable tool for high-resolution structural biology research. The ultimate outcome of a cryoEM study is an atomic model of a macromolecule or its complex with interacting partners. This chapter describes a variety of algorithms and software to build a de novo model based on the cryoEM 3D density map, to optimize the model with the best stereochemistry restraints and finally to validate the model with proper protocols. The full process of atomic structure determination from a cryoEM map is described. The tools outlined in this chapter should prove extremely valuable in revealing atomic interactions guided by cryoEM data. © 2016 Elsevier Inc. All rights reserved.

Contrast enhancement of biological nanoporous materials with zinc oxide infiltration for electron and X-ray nanoscale microscopy

DOE PAGES

Ocola, L. E.; Sampathkumar, V.; Kasthuri, N.; ...

2017-07-19

Here, we show that using infiltration of ZnO metal oxide can be useful for high resolution imaging of biological samples in electron and X-ray microscopy. This method is compatible with standard fixation techniques that leave the sample dry, such as finishing with super critical CO 2 drying, or simple vacuum drying at 95°C. We demonstrate this technique can be applied on tooth and brain tissue samples. We also show that high resolution X-ray tomography can be performed on biological systems using Zn K edge (1s) absorption to enhance internal structures, and obtained the first nanoscale 10 KeV X-ray absorption imagesmore » of the interior regions of a tooth.« less

Thermal Characterization of Carbon Nanotubes by Photothermal Techniques

NASA Astrophysics Data System (ADS)

Leahu, G.; Li Voti, R.; Larciprete, M. C.; Sibilia, C.; Bertolotti, M.; Nefedov, I.; Anoshkin, I. V.

2015-06-01

Carbon nanotubes (CNTs) are multifunctional materials commonly used in a large number of applications in electronics, sensors, nanocomposites, thermal management, actuators, energy storage and conversion, and drug delivery. Despite recent important advances in the development of CNT purity assessment tools and atomic resolution imaging of individual nanotubes by scanning tunnelling microscopy and high-resolution transmission electron microscopy, the macroscale assessment of the overall surface qualities of commercial CNT materials remains a great challenge. The lack of quantitative measurement technology to characterize and compare the surface qualities of bulk manufactured and engineered CNT materials has negative impacts on the reliable and consistent nanomanufacturing of CNT products. In this paper it is shown how photoacoustic spectroscopy and photothermal radiometry represent useful non-destructive tools to study the optothermal properties of carbon nanotube thin films.

A Microstructural Comparison of the Initial Growth of AIN and GaN Layers on Basal Plane Sapphire and SiC Substrates by Low Pressure Metalorganic Chemical Vapor Depositon

NASA Technical Reports Server (NTRS)

George, T.; Pike, W. T.; Khan, M. A.; Kuznia, J. N.; Chang-Chien, P.

1994-01-01

The initial growth by low pressure metalorganic chemical vapor deposition and subsequent thermal annealing of AIN and GaN epitaxial layers on SiC and sapphire substrates is examined using high resolution transmission electron microscopy and atomic force microscopy.

Multi-Resolution Imaging of Electron Dynamics in Nanostructure Interfaces

DTIC Science & Technology

2010-07-27

metallic carbon nanotubes from semiconducting ones. In pentacene transistors, we used scanning photocurrent microscopy to study spatially resolved...photoelectric response of pentacene thin films, which showed that point contacts formed near the hole injection points limit the overall performance of the...photothermal current microscopy, carbon nanotube transistor, pentacene transistor, contact resistance, hole injection 16. SECURITY CLASSIFICATION OF

High-resolution ab initio three-dimensional x-ray diffraction microscopy

DOE PAGES

Chapman, Henry N.; Barty, Anton; Marchesini, Stefano; ...

2006-01-01

Coherent x-ray diffraction microscopy is a method of imaging nonperiodic isolated objects at resolutions limited, in principle, by only the wavelength and largest scattering angles recorded. We demonstrate x-ray diffraction imaging with high resolution in all three dimensions, as determined by a quantitative analysis of the reconstructed volume images. These images are retrieved from the three-dimensional diffraction data using no a priori knowledge about the shape or composition of the object, which has never before been demonstrated on a nonperiodic object. We also construct two-dimensional images of thick objects with greatly increased depth of focus (without loss of transverse spatialmore » resolution). These methods can be used to image biological and materials science samples at high resolution with x-ray undulator radiation and establishes the techniques to be used in atomic-resolution ultrafast imaging at x-ray free-electron laser sources.« less

Atmospheric scanning electron microscope observes cells and tissues in open medium through silicon nitride film.

PubMed

Nishiyama, Hidetoshi; Suga, Mitsuo; Ogura, Toshihiko; Maruyama, Yuusuke; Koizumi, Mitsuru; Mio, Kazuhiro; Kitamura, Shinichi; Sato, Chikara

2010-03-01

Direct observation of subcellular structures and their characterization is essential for understanding their physiological functions. To observe them in open environment, we have developed an inverted scanning electron microscope with a detachable, open-culture dish, capable of 8 nm resolution, and combined with a fluorescence microscope quasi-simultaneously observing the same area from the top. For scanning electron microscopy from the bottom, a silicon nitride film window in the base of the dish maintains a vacuum between electron gun and open sample dish while allowing electrons to pass through. Electrons are backscattered from the sample and captured by a detector under the dish. Cells cultured on the open dish can be externally manipulated under optical microscopy, fixed, and observed using scanning electron microscopy. Once fine structures have been revealed by scanning electron microscopy, their component proteins may be identified by comparison with separately prepared fluorescence-labeled optical microscopic images of the candidate proteins, with their heavy-metal-labeled or stained ASEM images. Furthermore, cell nuclei in a tissue block stained with platinum-blue were successfully observed without thin-sectioning, which suggests the applicability of this inverted scanning electron microscope to cancer diagnosis. This microscope visualizes mesoscopic-scale structures, and is also applicable to non-bioscience fields including polymer chemistry. (c) 2010 Elsevier Inc. All rights reserved.

Next-generation endomyocardial biopsy: the potential of confocal and super-resolution microscopy.

PubMed

Crossman, David J; Ruygrok, Peter N; Hou, Yu Feng; Soeller, Christian

2015-03-01

Confocal laser scanning microscopy and super-resolution microscopy provide high-contrast and high-resolution fluorescent imaging, which has great potential to increase the diagnostic yield of endomyocardial biopsy (EMB). EMB is currently the gold standard for identification of cardiac allograft rejection, myocarditis, and infiltrative and storage diseases. However, standard analysis is dominated by low-contrast bright-field light and electron microscopy (EM); this lack of contrast makes quantification of pathological features difficult. For example, assessment of cardiac allograft rejection relies on subjective grading of H&E histology, which may lead to diagnostic variability between pathologists. This issue could be solved by utilising the high contrast provided by fluorescence methods such as confocal to quantitatively assess the degree of lymphocytic infiltrate. For infiltrative diseases such as amyloidosis, the nanometre resolution provided by EM can be diagnostic in identifying disease-causing fibrils. The recent advent of super-resolution imaging, particularly direct stochastic optical reconstruction microscopy (dSTORM), provides high-contrast imaging at resolution approaching that of EM. Moreover, dSTORM utilises conventional fluorescence dyes allowing for the same structures to be routinely imaged at the cellular scale and then at the nanoscale. The key benefit of these technologies is that the high contrast facilitates quantitative digital analysis and thereby provides a means to robustly assess critical pathological features. Ultimately, this technology has the ability to provide greater accuracy and precision to EMB assessment, which could result in better outcomes for patients.

Method to determine thermal profiles of nanoscale circuitry

DOEpatents

Zettl, Alexander K; Begtrup, Gavi E

2013-04-30

A platform that can measure the thermal profiles of devices with nanoscale resolution has been developed. The system measures the local temperature by using an array of nanoscale thermometers. This process can be observed in real time using a high resolution imagining technique such as electron microscopy. The platform can operate at extremely high temperatures.

Molecular dynamics-based refinement and validation for sub-5 Å cryo-electron microscopy maps

PubMed Central

Singharoy, Abhishek; Teo, Ivan; McGreevy, Ryan; Stone, John E; Zhao, Jianhua; Schulten, Klaus

2016-01-01

Two structure determination methods, based on the molecular dynamics flexible fitting (MDFF) paradigm, are presented that resolve sub-5 Å cryo-electron microscopy (EM) maps with either single structures or ensembles of such structures. The methods, denoted cascade MDFF and resolution exchange MDFF, sequentially re-refine a search model against a series of maps of progressively higher resolutions, which ends with the original experimental resolution. Application of sequential re-refinement enables MDFF to achieve a radius of convergence of ~25 Å demonstrated with the accurate modeling of β-galactosidase and TRPV1 proteins at 3.2 Å and 3.4 Å resolution, respectively. The MDFF refinements uniquely offer map-model validation and B-factor determination criteria based on the inherent dynamics of the macromolecules studied, captured by means of local root mean square fluctuations. The MDFF tools described are available to researchers through an easy-to-use and cost-effective cloud computing resource on Amazon Web Services. DOI: http://dx.doi.org/10.7554/eLife.16105.001 PMID:27383269

Refinement procedure for the image alignment in high-resolution electron tomography.

PubMed

Houben, L; Bar Sadan, M

2011-01-01

High-resolution electron tomography from a tilt series of transmission electron microscopy images requires an accurate image alignment procedure in order to maximise the resolution of the tomogram. This is the case in particular for ultra-high resolution where even very small misalignments between individual images can dramatically reduce the fidelity of the resultant reconstruction. A tomographic-reconstruction based and marker-free method is proposed, which uses an iterative optimisation of the tomogram resolution. The method utilises a search algorithm that maximises the contrast in tomogram sub-volumes. Unlike conventional cross-correlation analysis it provides the required correlation over a large tilt angle separation and guarantees a consistent alignment of images for the full range of object tilt angles. An assessment based on experimental reconstructions shows that the marker-free procedure is competitive to the reference of marker-based procedures at lower resolution and yields sub-pixel accuracy even for simulated high-resolution data. Copyright © 2011 Elsevier B.V. All rights reserved.

High-resolution measurements of the multilayer ultra-structure of articular cartilage and their translational potential

PubMed Central

2014-01-01

Current musculoskeletal imaging techniques usually target the macro-morphology of articular cartilage or use histological analysis. These techniques are able to reveal advanced osteoarthritic changes in articular cartilage but fail to give detailed information to distinguish early osteoarthritis from healthy cartilage, and this necessitates high-resolution imaging techniques measuring cells and the extracellular matrix within the multilayer structure of articular cartilage. This review provides a comprehensive exploration of the cellular components and extracellular matrix of articular cartilage as well as high-resolution imaging techniques, including magnetic resonance image, electron microscopy, confocal laser scanning microscopy, second harmonic generation microscopy, and laser scanning confocal arthroscopy, in the measurement of multilayer ultra-structures of articular cartilage. This review also provides an overview for micro-structural analysis of the main components of normal or osteoarthritic cartilage and discusses the potential and challenges associated with developing non-invasive high-resolution imaging techniques for both research and clinical diagnosis of early to late osteoarthritis. PMID:24946278

Correlation of two-photon in vivo imaging and FIB/SEM microscopy

PubMed Central

Blazquez-Llorca, L; Hummel, E; Zimmerman, H; Zou, C; Burgold, S; Rietdorf, J; Herms, J

2015-01-01

Advances in the understanding of brain functions are closely linked to the technical developments in microscopy. In this study, we describe a correlative microscopy technique that offers a possibility of combining two-photon in vivo imaging with focus ion beam/scanning electron microscope (FIB/SEM) techniques. Long-term two-photon in vivo imaging allows the visualization of functional interactions within the brain of a living organism over the time, and therefore, is emerging as a new tool for studying the dynamics of neurodegenerative diseases, such as Alzheimer’s disease. However, light microscopy has important limitations in revealing alterations occurring at the synaptic level and when this is required, electron microscopy is mandatory. FIB/SEM microscopy is a novel tool for three-dimensional high-resolution reconstructions, since it acquires automated serial images at ultrastructural level. Using FIB/SEM imaging, we observed, at 10 nm isotropic resolution, the same dendrites that were imaged in vivo over 9 days. Thus, we analyzed their ultrastructure and monitored the dynamics of the neuropil around them. We found that stable spines (present during the 9 days of imaging) formed typical asymmetric contacts with axons, whereas transient spines (present only during one day of imaging) did not form a synaptic contact. Our data suggest that the morphological classification that was assigned to a dendritic spine according to the in vivo images did not fit with its ultrastructural morphology. The correlative technique described herein is likely to open opportunities for unravelling the earlier unrecognized complexity of the nervous system. Lay Description Neuroscience and the understanding of brain functions are closely linked to the technical advances in microscopy. In this study we performed a correlative microscopy technique that offers the possibility to combine 2 photon in vivo imaging and FIB/SEM microscopy. Long term 2 photon in vivo imaging allows the visualization of functional interactions within the brain of a living organism over the time, and therefore, is emerging as a new tool to study the dynamics of neurodegenerative diseases, such as Alzheimer’s disease. However, light microscopy has important limitations in revealing synapses that are the connections between neurons, and for this purpose, the electron microscopy is necessary. FIB/SEM microscopy is a novel tool for three-dimensional (3D) high resolution reconstructions since it acquires automated serial images at ultrastructural level. This correlative technique will open up new horizons and opportunities for unravelling the complexity of the nervous system. PMID:25786682

Concept for room temperature single-spin tunneling force microscopy with atomic spatial resolution

NASA Astrophysics Data System (ADS)

Payne, Adam

A study of a force detected single-spin magnetic resonance measurement concept with atomic spatial resolution is presented. The method is based upon electrostatic force detection of spin-selection rule controlled single electron tunneling between two electrically isolated paramagnetic states. Single-spin magnetic resonance detection is possible by measuring the force detected tunneling charge noise on and off spin resonance. Simulation results of this charge noise, based upon physical models of the tunneling and spin physics, are directly compared to measured atomic force microscopy (AFM) system noise. The results show that the approach could provide single-spin measurement of electrically isolated defect states with atomic spatial resolution at room temperature.

Atomic-resolution single-spin magnetic resonance detection concept based on tunneling force microscopy

NASA Astrophysics Data System (ADS)

Payne, A.; Ambal, K.; Boehme, C.; Williams, C. C.

2015-05-01

A study of a force detected single-spin magnetic resonance measurement concept with atomic spatial resolution is presented. The method is based upon electrostatic force detection of spin-selection rule controlled single-electron tunneling between two electrically isolated paramagnetic states. Single-spin magnetic resonance detection is possible by measuring the force detected tunneling charge noise on and off spin resonance. Simulation results of this charge noise, based upon physical models of the tunneling and spin physics, are directly compared to measured atomic force microscopy system noise. The results show that the approach could provide single-spin measurement of electrically isolated qubit states with atomic spatial resolution at room temperature.

Light propagation and interaction observed with electrons.

PubMed

Word, Robert C; Fitzgerald, J P S; Könenkamp, R

2016-01-01

We discuss possibilities for a microscopic optical characterization of thin films and surfaces based on photoemission electron microscopy. We show that propagating light with wavelengths across the visible range can readily be visualized, and linear and non-linear materials properties can be evaluated non-invasively with nanometer spatial resolution. While femtosecond temporal resolution can be achieved in pump-probe-type experiments, the interferometric approach presented here has typical image frame times of ~200 fs. Copyright © 2015 Elsevier B.V. All rights reserved.

Nanoscale observation of the natural structure of milk-fat globules and casein micelles in the liquid condition using a scanning electron assisted dielectric microscopy.

PubMed

Ogura, Toshihiko; Okada, Tomoko

2017-09-30

Recently, aqueous nanoparticles have been used in drug-delivery systems for new type medicines. In particular, milk-casein micelles have been used as drug nanocarriers for targeting cancer cells. Therefore, nanostructure observation of particles and micelles in their native liquid condition is indispensable for analysing their function and mechanisms. However, traditional optical and scanning electron microscopy have difficulty observing the nanostructures of aqueous micelles. Recently, we developed a novel imaging technique called scanning electron-assisted dielectric microscopy (SE-ADM) that enables observation of various biological specimens in water with very little radiation damage and high-contrast imaging without staining or fixation at an 8-nm spatial resolution. In this study, for the first time, we show that the SE-ADM system is capable of high-resolution observation of whole-milk specimens in their natural state. Moreover, we successfully observe the casein micelles and milk-fat globules in an intact liquid condition. Our SE-ADM system can be applied to various biological particles and micelles in a native liquid state. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Attomicroscopy: from femtosecond to attosecond electron microscopy

NASA Astrophysics Data System (ADS)

Hassan, Mohammed Th

2018-02-01

In the last decade, the development of ultrafast electron diffraction (UED) and microscopy (UEM) have enabled the imaging of atomic motion in real time and space. These pivotal table-top tools opened the door for a vast range of applications in different areas of science spanning chemistry, physics, materials science, and biology. We first discuss the basic principles and recent advancements, including some of the important applications, of both UED and UEM. Then, we discuss the recent advances in the field that have enhanced the spatial and temporal resolutions, where the latter, is however, still limited to a few hundreds of femtoseconds, preventing the imaging of ultrafast dynamics of matter lasting few tens of femtoseconds. Then, we present our new optical gating approach for generating an isolated 30 fs electron pulse with sufficient intensity to attain a temporal resolution on the same time scale. This achievement allows, for the first time, imaging the electron dynamics of matter. Finally, we demonstrate the feasibility of the optical gating approach to generate an isolated attosecond electron pulse, utilizing our recently demonstrated optical attosecond laser pulse, which paves the way for establishing the field of ‘Attomicroscopy’, ultimately enabling us to image the electron motion in action.

TEM preparation methods and influence of radiation damage on the beam sensitive CaCO3 shell of Emiliania huxleyi.

PubMed

Hoffmann, Ramona; Wochnik, Angela S; Betzler, Sophia B; Matich, Sonja; Griesshaber, Erika; Schmahl, Wolfgang W; Scheu, Christina

2014-07-01

The ultrastructure of biologically formed calcium carbonate crystals like the shell of Emiliania huxleyi depends on the environmental conditions such as pH value, temperature and salinity. Therefore, they can be used as indicator for climate changes. However, for this a detailed understanding of their crystal structure and chemical composition is required. High resolution methods like transmission electron microscopy can provide those information on the nanoscale, given that sufficiently thin samples can be prepared. In our study, we developed sample preparation techniques for cross-section and plan-view investigations and studied the sample stability under electron bombardment. In addition to the biological material (Emiliania huxleyi) we also prepared mineralogical samples (Iceland spar) for comparison. High resolution transmission electron microscopy imaging, electron diffraction and electron energy-loss spectroscopy studies revealed that all prepared samples are relatively stable under electron bombardment at an acceleration voltage of 300 kV when using a parallel illumination. Above an accumulated dose of ∼10(5) e/nm2 the material--independent whether its origin is biological or geological--transformed to poly-crystalline calcium oxide. Copyright © 2014 Elsevier Ltd. All rights reserved.

Seeing tobacco mosaic virus through direct electron detectors

PubMed Central

Fromm, Simon A.; Bharat, Tanmay A.M.; Jakobi, Arjen J.; Hagen, Wim J.H.; Sachse, Carsten

2015-01-01

With the introduction of direct electron detectors (DED) to the field of electron cryo-microscopy, a wave of atomic-resolution structures has become available. As the new detectors still require comparative characterization, we have used tobacco mosaic virus (TMV) as a test specimen to study the quality of 3D image reconstructions from data recorded on the two direct electron detector cameras, K2 Summit and Falcon II. Using DED movie frames, we explored related image-processing aspects and compared the performance of micrograph-based and segment-based motion correction approaches. In addition, we investigated the effect of dose deposition on the atomic-resolution structure of TMV and show that radiation damage affects negative carboxyl chains first in a side-chain specific manner. Finally, using 450,000 asymmetric units and limiting the effects of radiation damage, we determined a high-resolution cryo-EM map at 3.35 Å resolution. Here, we provide a comparative case study of highly ordered TMV recorded on different direct electron detectors to establish recording and processing conditions that enable structure determination up to 3.2 Å in resolution using cryo-EM. PMID:25528571

Analysis of acute brain slices by electron microscopy: a correlative light-electron microscopy workflow based on Tokuyasu cryo-sectioning.

PubMed

Loussert Fonta, Celine; Leis, Andrew; Mathisen, Cliff; Bouvier, David S; Blanchard, Willy; Volterra, Andrea; Lich, Ben; Humbel, Bruno M

2015-01-01

Acute brain slices are slices of brain tissue that are kept vital in vitro for further recordings and analyses. This tool is of major importance in neurobiology and allows the study of brain cells such as microglia, astrocytes, neurons and their inter/intracellular communications via ion channels or transporters. In combination with light/fluorescence microscopies, acute brain slices enable the ex vivo analysis of specific cells or groups of cells inside the slice, e.g. astrocytes. To bridge ex vivo knowledge of a cell with its ultrastructure, we developed a correlative microscopy approach for acute brain slices. The workflow begins with sampling of the tissue and precise trimming of a region of interest, which contains GFP-tagged astrocytes that can be visualised by fluorescence microscopy of ultrathin sections. The astrocytes and their surroundings are then analysed by high resolution scanning transmission electron microscopy (STEM). An important aspect of this workflow is the modification of a commercial cryo-ultramicrotome to observe the fluorescent GFP signal during the trimming process. It ensured that sections contained at least one GFP astrocyte. After cryo-sectioning, a map of the GFP-expressing astrocytes is established and transferred to correlation software installed on a focused ion beam scanning electron microscope equipped with a STEM detector. Next, the areas displaying fluorescence are selected for high resolution STEM imaging. An overview area (e.g. a whole mesh of the grid) is imaged with an automated tiling and stitching process. In the final stitched image, the local organisation of the brain tissue can be surveyed or areas of interest can be magnified to observe fine details, e.g. vesicles or gold labels on specific proteins. The robustness of this workflow is contingent on the quality of sample preparation, based on Tokuyasu's protocol. This method results in a reasonable compromise between preservation of morphology and maintenance of antigenicity. Finally, an important feature of this approach is that the fluorescence of the GFP signal is preserved throughout the entire preparation process until the last step before electron microscopy. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Three-dimensional bright-field scanning transmission electron microscopy elucidate novel nanostructure in microbial biofilms.

PubMed

Hickey, William J; Shetty, Ameesha R; Massey, Randall J; Toso, Daniel B; Austin, Jotham

2017-01-01

Bacterial biofilms play key roles in environmental and biomedical processes, and understanding their activities requires comprehension of their nanoarchitectural characteristics. Electron microscopy (EM) is an essential tool for nanostructural analysis, but conventional EM methods are limited in that they either provide topographical information alone, or are suitable for imaging only relatively thin (<300 nm) sample volumes. For biofilm investigations, these are significant restrictions. Understanding structural relations between cells requires imaging of a sample volume sufficiently large to encompass multiple cells and the capture of both external and internal details of cell structure. An emerging EM technique with such capabilities is bright-field scanning transmission electron microscopy (BF-STEM) and in the present report BF-STEM was coupled with tomography to elucidate nanostructure in biofilms formed by the polycyclic aromatic hydrocarbon-degrading soil bacterium, Delftia acidovorans Cs1-4. Dual-axis BF-STEM enabled high-resolution 3-D tomographic recontructions (6-10 nm) visualization of thick (1250 and 1500 nm) sections. The 3-D data revealed that novel extracellular structures, termed nanopods, were polymorphic and formed complex networks within cell clusters. BF-STEM tomography enabled visualization of conduits formed by nanopods that could enable intercellular movement of outer membrane vesicles, and thereby enable direct communication between cells. This report is the first to document application of dual-axis BF-STEM tomography to obtain high-resolution 3-D images of novel nanostructures in bacterial biofilms. Future work with dual-axis BF-STEM tomography combined with correlative light electron microscopy may provide deeper insights into physiological functions associated with nanopods as well as other nanostructures. © 2016 The Authors Journal of Microscopy © 2016 Royal Microscopical Society.

Morphometric, quantitative, and three-dimensional analysis of the heart muscle fibers of old rats: transmission electron microscopy and high-resolution scanning electron microscopy methods.

PubMed

Cury, Diego Pulzatto; Dias, Fernando José; Sosthenes, Marcia Consentino Kronka; Dos Santos Haemmerle, Carlos Alexandre; Ogawa, Koichi; Da Silva, Marcelo Cavenaghi Pereira; Mardegan Issa, João Paulo; Iyomasa, Mamie Mizusaki; Watanabe, Ii-Sei

2013-02-01

This research investigated the morphological, morphometric, and ultrastructural cardiomyocyte characteristics of male Wistar rats at 18 months of age. The animals were euthanized using an overdose of anesthesia (ketamine and xylazine, 150/10 mg/kg) and perfused transcardially, after which samples were collected for light microscopy, transmission electron microscopy, and high-resolution scanning electron microscopy. The results showed that cardiomyocyte arrangement was disposed parallel between the mitochondria and the A-, I-, and H-bands and their M- and Z-lines from the sarcomere. The sarcomere junction areas had intercalated disks, a specific structure of heart muscle. The ultrastructural analysis revealed several mitochondria of various sizes and shapes intermingled between the blood capillaries and their endothelial cells; some red cells inside vessels are noted. The muscle cell sarcolemma could be observed associated with the described structures. The cardiomyocytes of old rats presented an average sarcomere length of 2.071 ± 0.09 μm, a mitochondrial volume density (Vv) of 0.3383, a mitochondrial average area of 0.537 ± 0.278 μm(2), a mitochondrial average length of 1.024 ± 0.352 μm, an average mitochondrial cristae thickness of 0.038 ± 0.09 μm and a ratio of mitochondrial greater length/lesser length of 1.929 ± 0.965. Of the observed mitochondrial shapes, 23.4% were rounded, 45.3% were elongated, and 31.1% had irregular profiles. In this study, we analyzed the morphology and morphometry of cardiomyocytes in old rats, focusing on mitochondria. These data are important for researchers who focus the changes in cardiac tissue, especially changes owing to pathologies and drug administration that may or may not be correlated with aging. Copyright © 2012 Wiley Periodicals, Inc.

Low-voltage electron microscopy of polymer and organic molecular thin films.

PubMed

Drummy, Lawrence F; Yang, Junyan; Martin, David C

2004-06-01

We have demonstrated the capabilities of a novel low-voltage electron microscope (LVEM) for imaging polymer and organic molecular thin films. The LVEM can operate in transmission electron microscopy, scanning transmission electron microscopy, scanning electron microscopy, and electron diffraction modes. The microscope operates at a nominal accelerating voltage of 5 kV and fits on a tabletop. A detailed discussion of the electron-sample interaction processes is presented, and the mean free path for total electron scattering was calculated to be 15 nm for organic samples at 5 kV. The total end point dose for the destruction of crystallinity at 5 kV was estimated at 5 x 10(-4) and 3.5 x 10(-2) C/cm2 for polyethylene and pentacene, respectively. These values are significantly lower than those measured at voltages greater than 100 kV. A defocus series of colloidal gold particles allowed us to estimate the experimental contrast transfer function of the microscope. Images taken of several organic materials have shown high contrast for low atomic number elements and a resolution of 2.5 nm. The materials studied here include thin films of the organic semiconductor pentacene, triblock copolymer films, single-molecule dendrimers, electrospun polymer fibers and gold nanoparticles. Copyright 2004 Elsevier B.V.

Correlated Optical Spectroscopy and Transmission Electron Microscopy of Individual Hollow Nanoparticles and their Dimers

PubMed Central

Yang, Linglu; Yan, Bo; Reinhard, Björn M.

2009-01-01

The optical spectra of individual Ag-Au alloy hollow particles were correlated with the particles’ structures obtained by transmission electron microscopy (TEM). The TEM provided direct experimental access to the dimension of the cavity, thickness of the metal shell, and the interparticle distance of hollow particle dimers with high spatial resolution. The analysis of correlated spectral and structural information enabled the quantification of the influence of the core-shell structure on the resonance energy, plasmon lifetime, and plasmon coupling efficiency. Electron beam exposure during TEM inspection was observed to affect plasmon wavelength and lifetime, making optical inspection prior to structural characterization mandatory. PMID:19768108

Defects in paramagnetic Co-doped ZnO films studied by transmission electron microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kovacs, Andras; Ney, A.; Duchamp, Martial

2013-12-23

We have studied planar defects in epitaxial Co:ZnO dilute magnetic semiconductor thin films deposited on c-plane sapphire (Al2O3) and the Co:ZnO/Al2O3 interface structure at atomic resolution using aberration-corrected transmission electron microscopy (TEM) and electron energy-loss spectroscopy (EELS). Comparing Co:ZnO samples deposited by pulsed laser deposition and reactive magnetron sputtering, both exhibit extrinsic stacking faults, incoherent interface structures, and compositional variations within the first 3-4 Co:ZnO layers at the interface.. In addition, we have measured the local strain which reveals the lattice distortion around the stacking faults.

Ultrafast core-loss spectroscopy in four-dimensional electron microscopy

PubMed Central

van der Veen, Renske M.; Penfold, Thomas J.; Zewail, Ahmed H.

2015-01-01

We demonstrate ultrafast core-electron energy-loss spectroscopy in four-dimensional electron microscopy as an element-specific probe of nanoscale dynamics. We apply it to the study of photoexcited graphite with femtosecond and nanosecond resolutions. The transient core-loss spectra, in combination with ab initio molecular dynamics simulations, reveal the elongation of the carbon-carbon bonds, even though the overall behavior is a contraction of the crystal lattice. A prompt energy-gap shrinkage is observed on the picosecond time scale, which is caused by local bond length elongation and the direct renormalization of band energies due to temperature-dependent electron–phonon interactions. PMID:26798793

Direct observation of charged domain walls in hybrid improper ferroelectric (Ca,Sr)3Ti2O7

NASA Astrophysics Data System (ADS)

Kurushima, Kousuke; Yoshimoto, Wataru; Ishii, Yui; Cheong, Sang-Wook; Mori, Shigeo

2017-10-01

We investigated ferroelectric (FE) domain wall structures including “charged domain walls” of hybrid improper FE (Ca,Sr)3Ti2O7 at the subatomic resolution by dark-field transmission electron microscopy (TEM) and high-resolution state-of-the-art aberration-corrected high-angle annular-dark-field (HAADF) scanning transmission electron microscopy (STEM). Dark-field TEM and high-resolution HAADF-STEM images obtained in the FE phase of single crystals of Ca2.46Sr0.54Ti2O7 revealed the formation of abundant charged domain walls with the head-to-head and tail-to-tail configurations in the FE domain structure, in addition to the FE 180° domain structure. The charged domain walls with the head-to-head and tail-to-tail FE polarizations exist stably and can be characterized as the unique double arc-type displacement of Ca/Sr ions in a unit cell without charge accumulation.

Challenges in Resolution for IC Failure Analysis

NASA Astrophysics Data System (ADS)

Martinez, Nick

1999-10-01

Resolution is becoming more and more of a challenge in the world of Failure Analysis in integrated circuits. This is a result of the ongoing size reduction in microelectronics. Determining the cause of a failure depends upon being able to find the responsible defect. The time it takes to locate a given defect is extremely important so that proper corrective actions can be taken. The limits of current microscopy tools are being pushed. With sub-micron feature sizes and even smaller killing defects, optical microscopes are becoming obsolete. With scanning electron microscopy (SEM), the resolution is high but the voltage involved can make these small defects transparent due to the large mean-free path of incident electrons. In this presentation, I will give an overview of the use of inspection methods in Failure Analysis and show example studies of my work as an Intern student at Texas Instruments. 1. Work at Texas Instruments, Stafford, TX, was supported by TI. 2. Work at Texas Tech University, was supported by NSF Grant DMR9705498.

Three-Dimensional Unstained Live-Cell Imaging Using Stimulated Parametric Emission Microscopy

NASA Astrophysics Data System (ADS)

Dang, Hieu M.; Kawasumi, Takehito; Omura, Gen; Umano, Toshiyuki; Kajiyama, Shin'ichiro; Ozeki, Yasuyuki; Itoh, Kazuyoshi; Fukui, Kiichi

2009-09-01

The ability to perform high-resolution unstained live imaging is very important to in vivo study of cell structures and functions. Stimulated parametric emission (SPE) microscopy is a nonlinear-optical microscopy based on ultra-fast electronic nonlinear-optical responses. For the first time, we have successfully applied this technique to archive three-dimensional (3D) images of unstained sub-cellular structures, such as, microtubules, nuclei, nucleoli, etc. in live cells. Observation of a complete cell division confirms the ability of SPE microscopy for long time-scale imaging.

The Influence of Beam Broadening on the Spatial Resolution of Annular Dark Field Scanning Transmission Electron Microscopy.

PubMed

de Jonge, Niels; Verch, Andreas; Demers, Hendrix

2018-02-01

The spatial resolution of aberration-corrected annular dark field scanning transmission electron microscopy was studied as function of the vertical position z within a sample. The samples consisted of gold nanoparticles (AuNPs) positioned in different horizontal layers within aluminum matrices of 0.6 and 1.0 µm thickness. The highest resolution was achieved in the top layer, whereas the resolution was reduced by beam broadening for AuNPs deeper in the sample. To examine the influence of the beam broadening, the intensity profiles of line scans over nanoparticles at a certain vertical location were analyzed. The experimental data were compared with Monte Carlo simulations that accurately matched the data. The spatial resolution was also calculated using three different theoretical models of the beam blurring as function of the vertical position within the sample. One model considered beam blurring to occur as a single scattering event but was found to be inaccurate for larger depths of the AuNPs in the sample. Two models were adapted and evaluated that include estimates for multiple scattering, and these described the data with sufficient accuracy to be able to predict the resolution. The beam broadening depended on z 1.5 in all three models.

The Three-Dimensional Point Spread Function of Aberration-Corrected Scanning Transmission Electron Microscopy

PubMed Central

Lupini, A.R.; de Jonge, N.

2012-01-01

Aberration-correction reduces the depth of field in scanning transmission electron microscopy (STEM) and thus allows three-dimensional imaging by depth-sectioning. This imaging mode offers the potential for sub-Ångstrom lateral resolution and nanometer-scale depth sensitivity. For biological samples, which may be many microns across and where high lateral resolution may not always be needed, optimizing the depth resolution even at the expense of lateral resolution may be desired, aiming to image through thick specimens. Although there has been extensive work examining and optimizing the probe formation in two-dimensions, there is less known about the probe shape along the optical axis. Here the probe shape is examined in three-dimensions in an attempt to better understand the depth-resolution in this mode. Examples are presented of how aberrations change the probe shape in three-dimensions, and it is found that off-axial aberrations may need to be considered for focal series of large areas. It is shown that oversized or annular apertures theoretically improve the vertical resolution for 3D imaging of nanoparticles. When imaging nanoparticles of several nanometer size, regular STEM can thereby be optimized such that the vertical full width at half maximum approaches that of the aberration corrected STEM with a standard aperture. PMID:21878149

Study of the structure of a thin aluminum layer on the vicinal surface of a gallium arsenide substrate by high-resolution electron microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lovygin, M. V., E-mail: lemi@miee.ru; Borgardt, N. I.; Seibt, M.

2015-12-15

The results of electron-microscopy studies of a thin epitaxial aluminum layer deposited onto a misoriented gallium-arsenide substrate are reported. It is established that the layer consists of differently oriented grains, whose crystal lattices are coherently conjugated with the substrate with the formation of misfit dislocations, as in the case of a layer on a singular substrate. Atomic steps on the substrate surface are visualized, and their influence on the growth of aluminum crystal grains is discussed.

Carbon Nanostructure Examined by Lattice Fringe Analysis of High Resolution Transmission Electron Microscopy Images

NASA Technical Reports Server (NTRS)

VanderWal, Randy L.; Tomasek, Aaron J.; Street, Kenneth; Thompson, William K.

2002-01-01

The dimensions of graphitic layer planes directly affect the reactivity of soot towards oxidation and growth. Quantification of graphitic structure could be used to develop and test correlations between the soot nanostructure and its reactivity. Based upon transmission electron microscopy images, this paper provides a demonstration of the robustness of a fringe image analysis code for determining the level of graphitic structure within nanoscale carbon, i.e. soot. Results, in the form of histograms of graphitic layer plane lengths, are compared to their determination through Raman analysis.

Carbon Nanostructure Examined by Lattice Fringe Analysis of High Resolution Transmission Electron Microscopy Images

NASA Technical Reports Server (NTRS)

VanderWal, Randy L.; Tomasek, Aaron J.; Street, Kenneth; Thompson, William K.; Hull, David R.

2003-01-01

The dimensions of graphitic layer planes directly affect the reactivity of soot towards oxidation and growth. Quantification of graphitic structure could be used to develop and test correlations between the soot nanostructure and its reactivity. Based upon transmission electron microscopy images, this paper provides a demonstration of the robustness of a fringe image analysis code for determining the level of graphitic structure within nanoscale carbon, i.e., soot. Results, in the form of histograms of graphitic layer plane lengths, are compared to their determination through Raman analysis.

Single particle analysis based on Zernike phase contrast transmission electron microscopy.

PubMed

Danev, Radostin; Nagayama, Kuniaki

2008-02-01

We present the first application of Zernike phase-contrast transmission electron microscopy to single-particle 3D reconstruction of a protein, using GroEL chaperonin as the test specimen. We evaluated the performance of the technique by comparing 3D models derived from Zernike phase contrast imaging, with models from conventional underfocus phase contrast imaging. The same resolution, about 12A, was achieved by both imaging methods. The reconstruction based on Zernike phase contrast data required about 30% fewer particles. The advantages and prospects of each technique are discussed.

Application of high-angle annular dark field scanning transmission electron microscopy, scanning transmission electron microscopy-energy dispersive X-ray spectrometry, and energy-filtered transmission electron microscopy to the characterization of nanoparticles in the environment.

PubMed

Utsunomiya, Satoshi; Ewing, Rodney C

2003-02-15

A major challenge to the development of a fundamental understanding of transport and retardation mechanisms of trace metal contaminants (<10 ppm) is their identification and characterization at the nanoscale. Atomic-scale techniques, such as conventional transmission electron microscopy, although powerful, are limited by the extremely small amounts of material that are examined. However, recent advances in electron microscopy provide a number of new analytical techniques that expand its application in environmental studies, particularly those concerning heavy metals on airborne particulates or water-borne colloids. High-angle annular dark field scanning transmission electron microscopy (HAADF-STEM), STEM-energy-dispersive X-ray spectrometry (EDX), and energy-filtered TEM (EFTEM) can be effectively used to identify and characterize nanoparticles. The image contrast in HAADF-STEM is strongly correlated to the atomic mass: heavier elements contribute to brighter contrast. Gold nanocrystals in pyrite and uranium nanocrystals in atmospheric aerosols have been identified by HAADF-STEM and STEM-EDX mapping and subsequently characterized by high-resolution TEM (HRTEM). EFTEM was used to identify U and Fe nanocrystals embedded in an aluminosilicate. A rare, As-bearing nanophase, westerveldite (FeAs), was identified by STEM-EDX and HRTEM. The combined use of these techniques greatly expands the effective application of electron microscopy in environmental studies, especially when applied to metals of very low concentrations. This paper describes examples of how these electron microbeam techniques can be used in combination to characterize a low concentration of heavy metals (a few ppm) on nanoscale particles.

Scanning capacitance microscopy of ErAs nanoparticles embedded in GaAs pn junctions

NASA Astrophysics Data System (ADS)

Park, K. W.; Nair, H. P.; Crook, A. M.; Bank, S. R.; Yu, E. T.

2011-09-01

Scanning capacitance microscopy is used to characterize the electronic properties of ErAs nanoparticles embedded in GaAs pn junctions grown by molecular beam epitaxy. Voltage-dependent capacitance images reveal localized variations in subsurface electronic structure near buried ErAs nanoparticles at lateral length scales of 20-30 nm. Numerical modeling indicates that these variations arise from inhomogeneities in charge modulation due to Fermi level pinning behavior associated with the embedded ErAs nanoparticles. Statistical analysis of image data yields an average particle radius of 6-8 nm—well below the direct resolution limit in scanning capacitance microscopy but discernible via analysis of patterns in nanoscale capacitance images.

National Cryo-Electron Microscopy Facility

Cancer.gov

Information about the National Cryo-EM Facility at NCI, created to provide researchers access to the latest cryo-EM technology for high resolution imaging. Includes timeline for installation and how to access the facility.

Efficient linear phase contrast in scanning transmission electron microscopy with matched illumination and detector interferometry

DOE PAGES

Ophus, Colin; Ciston, Jim; Pierce, Jordan; ...

2016-02-29

The ability to image light elements in soft matter at atomic resolution enables unprecedented insight into the structure and properties of molecular heterostructures and beam-sensitive nanomaterials. In this study, we introduce a scanning transmission electron microscopy technique combining a pre-specimen phase plate designed to produce a probe with structured phase with a high-speed direct electron detector to generate nearly linear contrast images with high efficiency. We demonstrate this method by using both experiment and simulation to simultaneously image the atomic-scale structure of weakly scattering amorphous carbon and strongly scattering gold nanoparticles. Our method demonstrates strong contrast for both materials, makingmore » it a promising candidate for structural determination of heterogeneous soft/hard matter samples even at low electron doses comparable to traditional phase-contrast transmission electron microscopy. Ultimately, simulated images demonstrate the extension of this technique to the challenging problem of structural determination of biological material at the surface of inorganic crystals.« less

Efficient linear phase contrast in scanning transmission electron microscopy with matched illumination and detector interferometry.

PubMed

Ophus, Colin; Ciston, Jim; Pierce, Jordan; Harvey, Tyler R; Chess, Jordan; McMorran, Benjamin J; Czarnik, Cory; Rose, Harald H; Ercius, Peter

2016-02-29

The ability to image light elements in soft matter at atomic resolution enables unprecedented insight into the structure and properties of molecular heterostructures and beam-sensitive nanomaterials. In this study, we introduce a scanning transmission electron microscopy technique combining a pre-specimen phase plate designed to produce a probe with structured phase with a high-speed direct electron detector to generate nearly linear contrast images with high efficiency. We demonstrate this method by using both experiment and simulation to simultaneously image the atomic-scale structure of weakly scattering amorphous carbon and strongly scattering gold nanoparticles. Our method demonstrates strong contrast for both materials, making it a promising candidate for structural determination of heterogeneous soft/hard matter samples even at low electron doses comparable to traditional phase-contrast transmission electron microscopy. Simulated images demonstrate the extension of this technique to the challenging problem of structural determination of biological material at the surface of inorganic crystals.

Efficient linear phase contrast in scanning transmission electron microscopy with matched illumination and detector interferometry

PubMed Central

Ophus, Colin; Ciston, Jim; Pierce, Jordan; Harvey, Tyler R.; Chess, Jordan; McMorran, Benjamin J.; Czarnik, Cory; Rose, Harald H.; Ercius, Peter

2016-01-01

The ability to image light elements in soft matter at atomic resolution enables unprecedented insight into the structure and properties of molecular heterostructures and beam-sensitive nanomaterials. In this study, we introduce a scanning transmission electron microscopy technique combining a pre-specimen phase plate designed to produce a probe with structured phase with a high-speed direct electron detector to generate nearly linear contrast images with high efficiency. We demonstrate this method by using both experiment and simulation to simultaneously image the atomic-scale structure of weakly scattering amorphous carbon and strongly scattering gold nanoparticles. Our method demonstrates strong contrast for both materials, making it a promising candidate for structural determination of heterogeneous soft/hard matter samples even at low electron doses comparable to traditional phase-contrast transmission electron microscopy. Simulated images demonstrate the extension of this technique to the challenging problem of structural determination of biological material at the surface of inorganic crystals. PMID:26923483

Static and Dynamic Electron Microscopy Investigations at the Atomic and Ultrafast Scales

NASA Astrophysics Data System (ADS)

Suri, Pranav Kumar

Advancements in the electron microscopy capabilities - aberration-corrected imaging, monochromatic spectroscopy, direct-electron detectors - have enabled routine visualization of atomic-scale processes with millisecond temporal resolutions in this decade. This, combined with progress in the transmission electron microscopy (TEM) specimen holder technology and nanofabrication techniques, allows comprehensive experiments on a wide range of materials in various phases via in situ methods. The development of ultrafast (sub-nanosecond) time-resolved TEM with ultrafast electron microscopy (UEM) has further pushed the envelope of in situ TEM to sub-nanosecond temporal resolution while maintaining sub-nanometer spatial resolution. A plethora of materials phenomena - including electron-phonon coupling, phonon transport, first-order phase transitions, bond rotation, plasmon dynamics, melting, and dopant atoms arrangement - are not yet clearly understood and could be benefitted with the current in situ TEM capabilities having atomic-level and ultrafast precision. Better understanding of these phenomena and intrinsic material dynamics (e.g. how phonons propagate in a material, what time-scales are involved in a first-order phase transition, how fast a material melts, where dopant atoms sit in a crystal) in new-generation and technologically important materials (e.g. two-dimensional layered materials, semiconductor and magnetic devices, rare-earth-element-free permanent magnets, unconventional superconductors) could bring a paradigm shift in their electronic, structural, magnetic, thermal and optical applications. Present research efforts, employing cutting-edge static and dynamic in situ electron microscopy resources at the University of Minnesota, are directed towards understanding the atomic-scale crystallographic structural transition and phonon transport in an iron-pnictide parent compound LaFeAsO, studying the mechanical stability of fast moving hard-drive heads in heat-assisted magnetic recording (HAMR) technology, exploring the possibility of ductile ceramics in magnesium oxide (MgO) nanomaterials, and revealing the atomic-structure of newly discovered rare-earth-element-free iron nitride (FeN) magnetic materials. Via atomic-resolution imaging and electron diffraction coupled with in situ TEM cooling on LaFeAsO, it was found that additional effects not related to the structural transition, namely dynamical scattering and electron channeling, can give signatures reminiscent of those typically associated with the symmetry change. UEM studies on LaFeAsO revealed direct, real-space imaging of the emergence and evolution of acoustic phonons and resolved dispersion behavior during propagation and scattering. Via UEM bright-field imaging, megahertz vibrational frequencies were observed upon laser-illumination in TEM specimens made out of HAMR devices which could be detrimental to their long-term thermal and structural reliability. Compression testing of 100-350 nm single-crystal MgO nanocubes shows size-dependent stresses and engineering strains of 4-13.8 GPa and 0.046-0.221 respectively at the first signs of yield accompanied by an absence of brittle fracture, which is a significant increase in plasticity of a brittle ceramic material. Atomic-scale characterization of FeN phases show that it is possible to detect interstitial locations of low atomic-number nitrogen atoms in iron crystal and hints at a development of novel routes (without involving rare-earth elements) for bulk permanent magnet synthesis.

Spectroscopic photon localization microscopy: breaking the resolution limit of single molecule localization microscopy (Conference Presentation)

NASA Astrophysics Data System (ADS)

Dong, Biqin; Almassalha, Luay Matthew; Urban, Ben E.; Nguyen, The-Quyen; Khuon, Satya; Chew, Teng-Leong; Backman, Vadim; Sun, Cheng; Zhang, Hao F.

2017-02-01

Distinguishing minute differences in spectroscopic signatures is crucial for revealing the fluorescence heterogeneity among fluorophores to achieve a high molecular specificity. Here we report spectroscopic photon localization microscopy (SPLM), a newly developed far-field spectroscopic imaging technique, to achieve nanoscopic resolution based on the principle of single-molecule localization microscopy while simultaneously uncovering the inherent molecular spectroscopic information associated with each stochastic event (Dong et al., Nature Communications 2016, in press). In SPLM, by using a slit-less monochromator, both the zero-order and the first-order diffractions from a grating were recorded simultaneously by an electron multiplying charge-coupled device to reveal the spatial distribution and the associated emission spectra of individual stochastic radiation events, respectively. As a result, the origins of photon emissions from different molecules can be identified according to their spectral differences with sub-nm spectral resolution, even when the molecules are within close proximity. With the newly developed algorithms including background subtraction and spectral overlap unmixing, we established and tested a method which can significantly extend the fundamental spatial resolution limit of single molecule localization microscopy by molecular discrimination through spectral regression. Taking advantage of this unique capability, we demonstrated improvement in spatial resolution of PALM/STORM up to ten fold with selected fluorophores. This technique can be readily adopted by other research groups to greatly enhance the optical resolution of single molecule localization microscopy without the need to modify their existing staining methods and protocols. This new resolving capability can potentially provide new insights into biological phenomena and enable significant research progress to be made in the life sciences.

Super-resolution imaging of subcortical white matter using stochastic optical reconstruction microscopy (STORM) and super-resolution optical fluctuation imaging (SOFI)

PubMed Central

Hainsworth, A. H.; Lee, S.; Patel, A.; Poon, W. W.; Knight, A. E.

2018-01-01

Aims The spatial resolution of light microscopy is limited by the wavelength of visible light (the ‘diffraction limit’, approximately 250 nm). Resolution of sub-cellular structures, smaller than this limit, is possible with super resolution methods such as stochastic optical reconstruction microscopy (STORM) and super-resolution optical fluctuation imaging (SOFI). We aimed to resolve subcellular structures (axons, myelin sheaths and astrocytic processes) within intact white matter, using STORM and SOFI. Methods Standard cryostat-cut sections of subcortical white matter from donated human brain tissue and from adult rat and mouse brain were labelled, using standard immunohistochemical markers (neurofilament-H, myelin-associated glycoprotein, glial fibrillary acidic protein, GFAP). Image sequences were processed for STORM (effective pixel size 8–32 nm) and for SOFI (effective pixel size 80 nm). Results In human, rat and mouse, subcortical white matter high-quality images for axonal neurofilaments, myelin sheaths and filamentous astrocytic processes were obtained. In quantitative measurements, STORM consistently underestimated width of axons and astrocyte processes (compared with electron microscopy measurements). SOFI provided more accurate width measurements, though with somewhat lower spatial resolution than STORM. Conclusions Super resolution imaging of intact cryo-cut human brain tissue is feasible. For quantitation, STORM can under-estimate diameters of thin fluorescent objects. SOFI is more robust. The greatest limitation for super-resolution imaging in brain sections is imposed by sample preparation. We anticipate that improved strategies to reduce autofluorescence and to enhance fluorophore performance will enable rapid expansion of this approach. PMID:28696566

Super-resolution imaging of subcortical white matter using stochastic optical reconstruction microscopy (STORM) and super-resolution optical fluctuation imaging (SOFI).

PubMed

Hainsworth, A H; Lee, S; Foot, P; Patel, A; Poon, W W; Knight, A E

2018-06-01

The spatial resolution of light microscopy is limited by the wavelength of visible light (the 'diffraction limit', approximately 250 nm). Resolution of sub-cellular structures, smaller than this limit, is possible with super resolution methods such as stochastic optical reconstruction microscopy (STORM) and super-resolution optical fluctuation imaging (SOFI). We aimed to resolve subcellular structures (axons, myelin sheaths and astrocytic processes) within intact white matter, using STORM and SOFI. Standard cryostat-cut sections of subcortical white matter from donated human brain tissue and from adult rat and mouse brain were labelled, using standard immunohistochemical markers (neurofilament-H, myelin-associated glycoprotein, glial fibrillary acidic protein, GFAP). Image sequences were processed for STORM (effective pixel size 8-32 nm) and for SOFI (effective pixel size 80 nm). In human, rat and mouse, subcortical white matter high-quality images for axonal neurofilaments, myelin sheaths and filamentous astrocytic processes were obtained. In quantitative measurements, STORM consistently underestimated width of axons and astrocyte processes (compared with electron microscopy measurements). SOFI provided more accurate width measurements, though with somewhat lower spatial resolution than STORM. Super resolution imaging of intact cryo-cut human brain tissue is feasible. For quantitation, STORM can under-estimate diameters of thin fluorescent objects. SOFI is more robust. The greatest limitation for super-resolution imaging in brain sections is imposed by sample preparation. We anticipate that improved strategies to reduce autofluorescence and to enhance fluorophore performance will enable rapid expansion of this approach. © 2017 British Neuropathological Society.

Nanodiamonds as multi-purpose labels for microscopy.

PubMed

Hemelaar, S R; de Boer, P; Chipaux, M; Zuidema, W; Hamoh, T; Martinez, F Perona; Nagl, A; Hoogenboom, J P; Giepmans, B N G; Schirhagl, R

2017-04-07

Nanodiamonds containing fluorescent nitrogen-vacancy centers are increasingly attracting interest for use as a probe in biological microscopy. This interest stems from (i) strong resistance to photobleaching allowing prolonged fluorescence observation times; (ii) the possibility to excite fluorescence using a focused electron beam (cathodoluminescence; CL) for high-resolution localization; and (iii) the potential use for nanoscale sensing. For all these schemes, the development of versatile molecular labeling using relatively small diamonds is essential. Here, we show the direct targeting of a biological molecule with nanodiamonds as small as 70 nm using a streptavidin conjugation and standard antibody labelling approach. We also show internalization of 40 nm sized nanodiamonds. The fluorescence from the nanodiamonds survives osmium-fixation and plastic embedding making them suited for correlative light and electron microscopy. We show that CL can be observed from epon-embedded nanodiamonds, while surface-exposed nanoparticles also stand out in secondary electron (SE) signal due to the exceptionally high diamond SE yield. Finally, we demonstrate the magnetic read-out using fluorescence from diamonds prior to embedding. Thus, our results firmly establish nanodiamonds containing nitrogen-vacancy centers as unique, versatile probes for combining and correlating different types of microscopy, from fluorescence imaging and magnetometry to ultrastructural investigation using electron microscopy.

Resolution Quality and Atom Positions in Sub-?ngstr?m Electron Microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

O'Keefe, Michael A.; Allard Jr, Lawrence Frederick; Blom, Douglas Allen

2005-01-01

John Cowley pioneered use of transmission electron microscopy (TEM) for high-resolution imaging and helped spur improvements in resolution that enabled researchers to pinpoint the positions of all but the lightest atoms within a crystal structure. Sub-{angstrom} capabilities allow imaging of even the lightest atoms. Initially achieved with software aberration correction (focal-series reconstruction of the specimen exit-surface wave), sub-{angstrom} imaging will become commonplace for next-generation electron microscopes with hardware-corrected lenses and monochromated electron beams. Currently, advanced HR-TEMs can image columns of light atoms (carbon, oxygen, nitrogen) in complex structures, including the lithium atoms present in battery materials. The ability to determinemore » whether an image peak represents one single atom (or atom column) instead of several depends on the resolution of the HR-(S)TEM. Rayleigh's resolution criterion, an accepted standard in optics, was derived as a means for judging when two image intensity peaks from two sources of light (stars) are distinguishable from a single source. Atom spacings closer than the Rayleigh limit have been resolved in HR-TEM, suggesting that it may be useful to consider other limits, such as the Sparrow resolution criterion. From the viewpoint of the materials scientist, it is important to be able to use the image to determine whether an image feature represents one or more atoms (resolution), and where the atoms (or atom columns) are positioned relative to one another (resolution quality). When atoms and the corresponding image peaks are separated by more than the Rayleigh limit of the HR-(S)TEM, it is possible to adjust imaging parameters so that relative peak positions in the image correspond to relative atom positions in the specimen. When atoms are closer than the Rayleigh limit, we must find the relationship of the peak position to the atom position by peak fitting or, if we have a suitable model, by image simulation.« less

Transmission electron microscopy: direct observation of crystal structure in refractory ceramics.

PubMed

Shaw, T M; Thomas, G

1978-11-10

Using high-resolution multibeam interference techniques in the transmission electron microscope, images have been obtained that make possible a real-space structure analysis of a beryllium-silicon-nitrogen compound. The results illustrate the usefulness of lattice imaging in the analysis of local crystal structure in these technologically promising ceramic materials.

Micro-CT scouting for transmission electron microscopy of human tissue specimens

DOE Office of Scientific and Technical Information (OSTI.GOV)

Morales, A. G.; Stempinski, E. S.; XIAO, X.

Transmission electron microscopy (TEM) provides sub-nanometre-scale details in volumetric samples. Samples such as pathology tissue specimens are often stained with a metal element to enhance contrast, which makes them opaque to optical microscopes. As a result, it can be a lengthy procedure to find the region of interest inside a sample through sectioning. Here, we describe micro-CT scouting for TEM that allows noninvasive identification of regions of interest within a block sample to guide the sectioning step. In a tissue pathology study, a bench-top micro-CT scanner with 10 m resolution was used to determine the location of patches of themore » mucous membrane in osmium-stained human nasal scraping samples. Furthermore, once the regions of interest were located, the sample block was sectioned to expose that location, followed by ultra-thin sectioning and TEM to inspect the internal structure of the cilia of the membrane epithelial cells with nanometre resolution. This method substantially reduced the time and labour of the search process from typically 20 sections for light microscopy to three sections with no added sample preparation. Lay description Electron microscopy provides very high levels of detail in a small area, and thus the question of where to look in an opaque sample, such as a stained tissue specimen, needs to be answered by sectioning the sample in small steps and examining the sections under a light microscope, until the region of interest is found. The search process can be lengthy and labor intensive, especially for a study involving a large number of samples. Small areas of interest can be missed in the process if not enough regions are examined. We also describe a method to directly locate the region of interest within a whole sample using micro-CT imaging, bypassing the need of blindly sectioning. Micro-CT enables locating the region within 3D space; this information provides a guide for sectioning the sample to expose that precise location for high resolution electron microscopy imaging. In a human tissue specimen study, this method considerably reduced the time and labor of the search process.« less

Micro-CT scouting for transmission electron microscopy of human tissue specimens

DOE PAGES

Morales, A. G.; Stempinski, E. S.; XIAO, X.; ...

2016-02-08

Transmission electron microscopy (TEM) provides sub-nanometre-scale details in volumetric samples. Samples such as pathology tissue specimens are often stained with a metal element to enhance contrast, which makes them opaque to optical microscopes. As a result, it can be a lengthy procedure to find the region of interest inside a sample through sectioning. Here, we describe micro-CT scouting for TEM that allows noninvasive identification of regions of interest within a block sample to guide the sectioning step. In a tissue pathology study, a bench-top micro-CT scanner with 10 m resolution was used to determine the location of patches of themore » mucous membrane in osmium-stained human nasal scraping samples. Furthermore, once the regions of interest were located, the sample block was sectioned to expose that location, followed by ultra-thin sectioning and TEM to inspect the internal structure of the cilia of the membrane epithelial cells with nanometre resolution. This method substantially reduced the time and labour of the search process from typically 20 sections for light microscopy to three sections with no added sample preparation. Lay description Electron microscopy provides very high levels of detail in a small area, and thus the question of where to look in an opaque sample, such as a stained tissue specimen, needs to be answered by sectioning the sample in small steps and examining the sections under a light microscope, until the region of interest is found. The search process can be lengthy and labor intensive, especially for a study involving a large number of samples. Small areas of interest can be missed in the process if not enough regions are examined. We also describe a method to directly locate the region of interest within a whole sample using micro-CT imaging, bypassing the need of blindly sectioning. Micro-CT enables locating the region within 3D space; this information provides a guide for sectioning the sample to expose that precise location for high resolution electron microscopy imaging. In a human tissue specimen study, this method considerably reduced the time and labor of the search process.« less

Low cost, high performance processing of single particle cryo-electron microscopy data in the cloud

PubMed Central

Cianfrocco, Michael A; Leschziner, Andres E

2015-01-01

The advent of a new generation of electron microscopes and direct electron detectors has realized the potential of single particle cryo-electron microscopy (cryo-EM) as a technique to generate high-resolution structures. Calculating these structures requires high performance computing clusters, a resource that may be limiting to many likely cryo-EM users. To address this limitation and facilitate the spread of cryo-EM, we developed a publicly available ‘off-the-shelf’ computing environment on Amazon's elastic cloud computing infrastructure. This environment provides users with single particle cryo-EM software packages and the ability to create computing clusters with 16–480+ CPUs. We tested our computing environment using a publicly available 80S yeast ribosome dataset and estimate that laboratories could determine high-resolution cryo-EM structures for $50 to $1500 per structure within a timeframe comparable to local clusters. Our analysis shows that Amazon's cloud computing environment may offer a viable computing environment for cryo-EM. DOI: http://dx.doi.org/10.7554/eLife.06664.001 PMID:25955969

Nanoscale imaging of whole cells using a liquid enclosure and a scanning transmission electron microscope.

PubMed

Peckys, Diana B; Veith, Gabriel M; Joy, David C; de Jonge, Niels

2009-12-14

Nanoscale imaging techniques are needed to investigate cellular function at the level of individual proteins and to study the interaction of nanomaterials with biological systems. We imaged whole fixed cells in liquid state with a scanning transmission electron microscope (STEM) using a micrometer-sized liquid enclosure with electron transparent windows providing a wet specimen environment. Wet-STEM images were obtained of fixed E. coli bacteria labeled with gold nanoparticles attached to surface membrane proteins. Mammalian cells (COS7) were incubated with gold-tagged epidermal growth factor and fixed. STEM imaging of these cells resulted in a resolution of 3 nm for the gold nanoparticles. The wet-STEM method has several advantages over conventional imaging techniques. Most important is the capability to image whole fixed cells in a wet environment with nanometer resolution, which can be used, e.g., to map individual protein distributions in/on whole cells. The sample preparation is compatible with that used for fluorescent microscopy on fixed cells for experiments involving nanoparticles. Thirdly, the system is rather simple and involves only minimal new equipment in an electron microscopy (EM) laboratory.

Structure and chemistry of epitaxial ceria thin films on yttria-stabilized zirconia substrates, studied by high resolution electron microscopy

DOE PAGES

Sinclair, Robert; Lee, Sang Chul; Shi, Yezhou; ...

2017-03-18

Here, we have applied aberration-corrected transmission electron microscopy (TEM) imaging and electron energy loss spectroscopy (EELS) to study the structure and chemistry of epitaxial ceria thin films, grown by pulsed laser deposition onto (001) yttria-stabilized zirconia (YSZ) substrates. There are few observable defects apart from the expected mismatch interfacial dislocations and so the films would be expected to have good potential for applications. Under high electron beam dose rate (above about 6000 e-/Å 2s) domains of an ordered structure appear and these are interpreted as being created by oxygen vacancy ordering. The ordered structure does not appear at lower losemore » rates (ca. 2600 e-/Å 2s) and can be removed by imaging under 1 mbar oxygen gas in an environmental TEM. EELS confirms that there is both oxygen deficiency and the associated increase in Ce 3+ versus Ce 4+ cations in the ordered domains. In situ high resolution TEM recordings show the formation of the ordered domains as well as atomic migration along the ceria thin film (001) surface.« less

Structure and chemistry of epitaxial ceria thin films on yttria-stabilized zirconia substrates, studied by high resolution electron microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sinclair, Robert; Lee, Sang Chul; Shi, Yezhou

Here, we have applied aberration-corrected transmission electron microscopy (TEM) imaging and electron energy loss spectroscopy (EELS) to study the structure and chemistry of epitaxial ceria thin films, grown by pulsed laser deposition onto (001) yttria-stabilized zirconia (YSZ) substrates. There are few observable defects apart from the expected mismatch interfacial dislocations and so the films would be expected to have good potential for applications. Under high electron beam dose rate (above about 6000 e-/Å 2s) domains of an ordered structure appear and these are interpreted as being created by oxygen vacancy ordering. The ordered structure does not appear at lower losemore » rates (ca. 2600 e-/Å 2s) and can be removed by imaging under 1 mbar oxygen gas in an environmental TEM. EELS confirms that there is both oxygen deficiency and the associated increase in Ce 3+ versus Ce 4+ cations in the ordered domains. In situ high resolution TEM recordings show the formation of the ordered domains as well as atomic migration along the ceria thin film (001) surface.« less

Using Synchrotron Radiation and Electron Microscopy to Map the Huge Structural Changes that Occur in Viruses During Their Life Cycle

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rossman, Michael

2011-09-07

The crystallographic techniques for structure determination of proteins and neucleic acids at near atomic resolution using synchrotron X-radiation has become almost automatic. However the limits of this procedure are determined by the availability of crystals. As the size and complexity of the molecular assemblies being studied increases, the likelihood of growing useful crystals diminishes. Cryo electron microscopy and tomography have extended the range of biological objects that can be determined at near atomic resolution. Furthermore it is now becoming apparent that the function of the molecular assemblies most often requires very large conformational changes that could never be contained withinmore » a crystal, Examples will be presented of the structural changes that occur in viruses as they assembly and prepare to infect new cells.« less

Preparation of superhydrophobic and transparent micro-nano hybrid coatings from polymethylhydroxysiloxane and silica ormosil aerogels

NASA Astrophysics Data System (ADS)

Nagappan, Saravanan; Park, Jin Joo; Park, Sung Soo; Ha, Chang-Sik

2014-12-01

Superhydrophobic and transparent polymethylhydroxysiloxane (PMHOS)/silica ormosil aerogel hybrids were prepared successfully by mixing of PMHOS with various weight percentages of silica ormosil aerogels (as synthesized from methyltriethoxysilane (MTES) and methyltrimethoxysilane (MTMS) precursors) in separate seal perfume glass vials. The hybrids were spin coated on glass substrate at 1000 rpm for 60 seconds and used for further analysis. The surface morphology and chemical compositions of the hybrids were analyzed by high resolution scanning electron microscopy, high resolution transmission electron microscopy, atomic force spectroscopy, adsorption and desorption isotherm, and X-ray photoelectron spectroscopy. The transparency, thermal decomposition and static contact angle (SCA) of each sample were measured by UV-Visible spectrophotometer, TGA and drop shape analysis system, respectively. The spin coated substrates showed good superhydrophobic properties, thermal stability as well as transparency on the glass substrates.

Simple synthesis of MoS{sub 2} inorganic fullerene-like nanomaterials from MoS{sub 2} amorphous nanoparticles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chang Lianxia; China Faw Group Corporation R and D Center, Changchun, 130011; Yang Haibin

The amorphous MoS{sub 2} nanoparticles have been synthesized by a simple oxidation-reduction reaction in an aqueous solution. A series of products with different morphologies, such as MoS{sub 2} nanospheres, inorganic fullerene-like nanospheres, nanorods and Mo bended rods, can be obtained by annealing the amorphous MoS{sub 2} nanoparticles under N{sub 2} atmosphere under 400-1200 deg. C. These products have been characterized by X-ray diffraction, field emission scanning electronic microscopy, transmission electron microscopy and high-resolution transmission electron microscopy in detail. The possible transformation mechanism for the structure has been discussed based on the experimental results. In addition, the optical properties of IF-MoS{submore » 2} have also been performed by UV-vis absorption spectroscopy.« less

Transfer doping of single isolated nanodiamonds, studied by scanning probe microscopy techniques.

PubMed

Bolker, Asaf; Saguy, Cecile; Kalish, Rafi

2014-09-26

The transfer doping of diamond surfaces has been applied in various novel two-dimensional electronic devices. Its extension to nanodiamonds (ND) is essential for ND-based applications in many fields. In particular, understanding the influence of the crystallite size on transfer doping is desirable. Here, we report the results of a detailed study of the electronic energetic band structure of single, isolated transfer-doped nanodiamonds with nanometric resolution using a combination of scanning tunneling spectroscopy and Kelvin force microscopy measurements. The results show how the band gap, the valence band maximum, the electron affinity and the work function all depend on the ND's size and nanoparticle surface properties. The present analysis, which combines information from both scanning tunneling spectroscopy and Kelvin force microscopy, should be applicable to any nanoparticle or surface that can be measured with scanning probe techniques.

Magnified pseudo-elemental map of atomic column obtained by Moiré method in scanning transmission electron microscopy.

PubMed

Kondo, Yukihito; Okunishi, Eiji

2014-10-01

Moiré method in scanning transmission electron microscopy allows observing a magnified two-dimensional atomic column elemental map of a higher pixel resolution with a lower electron dose unlike conventional atomic column mapping. The magnification of the map is determined by the ratio between the pixel size and the lattice spacing. With proper ratios for the x and y directions, we could observe magnified elemental maps, homothetic to the atomic arrangement in the sample of SrTiO3 [0 0 1]. The map showed peaks at all expected oxygen sites in SrTiO3 [0 0 1]. © The Author 2014. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Challenges in quantitative crystallographic characterization of 3D thin films by ACOM-TEM.

PubMed

Kobler, A; Kübel, C

2017-02-01

Automated crystal orientation mapping for transmission electron microscopy (ACOM-TEM) has become an easy to use method for the investigation of crystalline materials and complements other TEM methods by adding local crystallographic information over large areas. It fills the gap between high resolution electron microscopy and electron back scatter diffraction in terms of spatial resolution. Recent investigations showed that spot diffraction ACOM-TEM is a quantitative method with respect to sample parameters like grain size, twin density, orientation density and others. It can even be used in combination with in-situ tensile or thermal testing. However, there are limitations of the current method. In this paper we discuss some of the challenges and discuss solutions, e.g. we present an ambiguity filter that reduces the number of pixels with a '180° ambiguity problem'. For that an ACOM-TEM tilt series of nanocrystalline Pd thin films with overlapping crystallites was acquired and analyzed. Copyright © 2017. Published by Elsevier B.V.

Electronic and structural characteristics of zinc-blende wurtzite biphasic homostructure GaN nanowires

DOE PAGES

Jacobs, Benjamin W.; Ayres, Virginia M.; Petkov, Mihail P.; ...

2007-04-07

Here, we report a new biphasic crystalline wurtzite/zinc-blende homostructure in gallium nitride nanowires. Cathodoluminescence was used to quantitatively measure the wurtzite and zinc-blende band gaps. High-resolution transmission electron microscopy was used to identify distinct wurtzite and zinc-blende crystalline phases within single nanowires through the use of selected area electron diffraction, electron dispersive spectroscopy, electron energy loss spectroscopy, and fast Fourier transform techniques. A mechanism for growth is identified.

Electronic and structural characteristics of zinc-blende wurtzite biphasic homostructure GaN nanowires.

PubMed

Jacobs, Benjamin W; Ayres, Virginia M; Petkov, Mihail P; Halpern, Joshua B; He, Maoqi; Baczewski, Andrew D; McElroy, Kaylee; Crimp, Martin A; Zhang, Jiaming; Shaw, Harry C

2007-05-01

We report a new biphasic crystalline wurtzite/zinc-blende homostructure in gallium nitride nanowires. Cathodoluminescence was used to quantitatively measure the wurtzite and zinc-blende band gaps. High-resolution transmission electron microscopy was used to identify distinct wurtzite and zinc-blende crystalline phases within single nanowires through the use of selected area electron diffraction, electron dispersive spectroscopy, electron energy loss spectroscopy, and fast Fourier transform techniques. A mechanism for growth is identified.

Technical Review: Microscopy and Image Processing Tools to Analyze Plant Chromatin: Practical Considerations.

PubMed

Baroux, Célia; Schubert, Veit

2018-01-01

In situ nucleus and chromatin analyses rely on microscopy imaging that benefits from versatile, efficient fluorescent probes and proteins for static or live imaging. Yet the broad choice in imaging instruments offered to the user poses orientation problems. Which imaging instrument should be used for which purpose? What are the main caveats and what are the considerations to best exploit each instrument's ability to obtain informative and high-quality images? How to infer quantitative information on chromatin or nuclear organization from microscopy images? In this review, we present an overview of common, fluorescence-based microscopy systems and discuss recently developed super-resolution microscopy systems, which are able to bridge the resolution gap between common fluorescence microscopy and electron microscopy. We briefly present their basic principles and discuss their possible applications in the field, while providing experience-based recommendations to guide the user toward best-possible imaging. In addition to raw data acquisition methods, we discuss commercial and noncommercial processing tools required for optimal image presentation and signal evaluation in two and three dimensions.

Green synthesis of BiVO4 nanorods via aqueous extracts of Callistemon viminalis

NASA Astrophysics Data System (ADS)

Mohamed, H. E. A.; Sone, B. T.; Fuku, X. G.; Dhlamini, M. S.; Maaza, M.

2018-05-01

Nowadays, the development of efficient green chemistry methods for synthesis of metal oxides nanoparticles has become a major focus of researchers. These methods are being investigated in order to find an eco-friendly technique for production of well-characterized nanoparticles. In this contribution we report for the first time, the synthesis and structural characterization of n-type Bismuth vanadate (BiVO4) nanoparticles using aqueous extracts of Callistemon viminalis as a chelating agent. To ascertain the formation of BiVO4, X-Ray diffraction analysis (XRD), Scanning Electron Microscopy (SEM), High Resolution Transmission Electron Microscopy (TEM), Selected Area Electron Diffraction (SAED), Electron Dispersion X-ray Spectroscopy (EDS), Fourier Transform Infra-red Spectroscopy (FTIR), and Photoluminescence spectroscopy (PL) were carried out.

Nucleation of diamond by pure carbon ion bombardment—a transmission electron microscopy study

NASA Astrophysics Data System (ADS)

Yao, Y.; Liao, M. Y.; Wang, Z. G.; Lifshitz, Y.; Lee, S. T.

2005-08-01

A cross-sectional high-resolution transmission electron microscopy (HRTEM) study of a film deposited by a 1 keV mass-selected carbon ion beam onto silicon held at 800 °C is presented. Initially, a graphitic film with its basal planes perpendicular to the substrate is evolving. The precipitation of nanodiamond crystallites in upper layers is confirmed by HRTEM, selected area electron diffraction, and electron energy loss spectroscopy. The nucleation of diamond on graphitic edges as predicted by Lambrecht et al. [W. R. L. Lambrecht, C. H. Lee, B. Segall, J. C. Angus, Z. Li, and M. Sunkara, Nature, 364 607 (1993)] is experimentally confirmed. The results are discussed in terms of our recent subplantation-based diamond nucleation model.

Large-scale synthesis of monodisperse magnesium ferrite via an environmentally friendly molten salt route.

PubMed

Lou, Zhengsong; He, Minglong; Wang, Ruikun; Qin, Weiwei; Zhao, Dejian; Chen, Changle

2014-02-17

Sub-micrometer-sized magnesium ferrite spheres consisting of uniform small particles have been prepared using a facile, large-scale solid-state reaction employing a molten salt technique. Extensive structural characterization of the as-prepared samples has been performed using scanning electron microscope, transmission electron microscopy, high-resolution transmission electron microscopy, selected area electron diffraction, and X-ray diffraction. The yield of the magnesium ferrite sub-micrometer spheres is up to 90%, and these sub-micrometer spheres are made up of square and rectangular nanosheets. The magnetic properties of magnesium ferrite sub-micrometer spheres are investigated, and the magnetization saturation value is about 24.96 emu/g. Moreover, the possible growth mechanism is proposed based on the experimental results.

Conduction at domain walls in oxide multiferroics

NASA Astrophysics Data System (ADS)

Seidel, J.; Martin, L. W.; He, Q.; Zhan, Q.; Chu, Y.-H.; Rother, A.; Hawkridge, M. E.; Maksymovych, P.; Yu, P.; Gajek, M.; Balke, N.; Kalinin, S. V.; Gemming, S.; Wang, F.; Catalan, G.; Scott, J. F.; Spaldin, N. A.; Orenstein, J.; Ramesh, R.

2009-03-01

Domain walls may play an important role in future electronic devices, given their small size as well as the fact that their location can be controlled. Here, we report the observation of room-temperature electronic conductivity at ferroelectric domain walls in the insulating multiferroic BiFeO3. The origin and nature of the observed conductivity are probed using a combination of conductive atomic force microscopy, high-resolution transmission electron microscopy and first-principles density functional computations. Our analyses indicate that the conductivity correlates with structurally driven changes in both the electrostatic potential and the local electronic structure, which shows a decrease in the bandgap at the domain wall. Additionally, we demonstrate the potential for device applications of such conducting nanoscale features.

Conduction at domain walls in oxide multiferroics.

PubMed

Seidel, J; Martin, L W; He, Q; Zhan, Q; Chu, Y-H; Rother, A; Hawkridge, M E; Maksymovych, P; Yu, P; Gajek, M; Balke, N; Kalinin, S V; Gemming, S; Wang, F; Catalan, G; Scott, J F; Spaldin, N A; Orenstein, J; Ramesh, R

2009-03-01

Domain walls may play an important role in future electronic devices, given their small size as well as the fact that their location can be controlled. Here, we report the observation of room-temperature electronic conductivity at ferroelectric domain walls in the insulating multiferroic BiFeO(3). The origin and nature of the observed conductivity are probed using a combination of conductive atomic force microscopy, high-resolution transmission electron microscopy and first-principles density functional computations. Our analyses indicate that the conductivity correlates with structurally driven changes in both the electrostatic potential and the local electronic structure, which shows a decrease in the bandgap at the domain wall. Additionally, we demonstrate the potential for device applications of such conducting nanoscale features.

Experiments in electron microscopy: from metals to nerves

NASA Astrophysics Data System (ADS)

Unwin, Nigel

2015-04-01

Electron microscopy has advanced remarkably as a tool for biological structure research since the development of methods to examine radiation-sensitive unstained specimens and the introduction of cryo-techniques. Structures of biological molecules at near-atomic resolution can now be obtained from images of single particles as well as crystalline arrays. It has also become possible to analyze structures of molecules in their functional context, i.e. in their natural membrane or cellular setting, and in an ionic environment like that in living tissue. Electron microscopy is thus opening ways to answer definitively questions about physiological mechanisms. Here I recall a number of experiments contributing to, and benefiting from the technical advances that have taken place. I begin—in the spirit of this crystallography series—with some biographical background, and then sketch the path to an analysis by time-resolved microscopy of the opening mechanism of an ion channel (nicotinic acetylcholine receptor). This analysis illustrates how electron imaging can be combined with freeze-trapping to illuminate a transient biological event: in our case, chemical-to-electrical transduction at the nerve-muscle synapse.

Bandgap Inhomogeneity of a PbSe Quantum Dot Ensemble from Two-Dimensional Spectroscopy and Comparison to Size Inhomogeneity from Electron Microscopy

DOE PAGES

Park, Samuel D.; Baranov, Dmitry; Ryu, Jisu; ...

2017-01-03

Femtosecond two-dimensional Fourier transform spectroscopy is used to determine the static bandgap inhomogeneity of a colloidal quantum dot ensemble. The excited states of quantum dots absorb light, so their absorptive two-dimensional (2D) spectra will typically have positive and negative peaks. We show that the absorption bandgap inhomogeneity is robustly determined by the slope of the nodal line separating positive and negative peaks in the 2D spectrum around the bandgap transition; this nodal line slope is independent of excited state parameters not known from the absorption and emission spectra. The absorption bandgap inhomogeneity is compared to a size and shape distributionmore » determined by electron microscopy. The electron microscopy images are analyzed using new 2D histograms that correlate major and minor image projections to reveal elongated nanocrystals, a conclusion supported by grazing incidence small-angle X-ray scattering and high-resolution transmission electron microscopy. Lastly, the absorption bandgap inhomogeneity quantitatively agrees with the bandgap variations calculated from the size and shape distribution, placing upper bounds on any surface contributions.« less

Electron tomography of HEK293T cells using scanning electron microscope-based scanning transmission electron microscopy.

PubMed

You, Yun-Wen; Chang, Hsun-Yun; Liao, Hua-Yang; Kao, Wei-Lun; Yen, Guo-Ji; Chang, Chi-Jen; Tsai, Meng-Hung; Shyue, Jing-Jong

2012-10-01

Based on a scanning electron microscope operated at 30 kV with a homemade specimen holder and a multiangle solid-state detector behind the sample, low-kV scanning transmission electron microscopy (STEM) is presented with subsequent electron tomography for three-dimensional (3D) volume structure. Because of the low acceleration voltage, the stronger electron-atom scattering leads to a stronger contrast in the resulting image than standard TEM, especially for light elements. Furthermore, the low-kV STEM yields less radiation damage to the specimen, hence the structure can be preserved. In this work, two-dimensional STEM images of a 1-μm-thick cell section with projection angles between ±50° were collected, and the 3D volume structure was reconstructed using the simultaneous iterative reconstructive technique algorithm with the TomoJ plugin for ImageJ, which are both public domain software. Furthermore, the cross-sectional structure was obtained with the Volume Viewer plugin in ImageJ. Although the tilting angle is constrained and limits the resulting structural resolution, slicing the reconstructed volume generated the depth profile of the thick specimen with sufficient resolution to examine cellular uptake of Au nanoparticles, and the final position of these nanoparticles inside the cell was imaged.

Electron microscopy study of gold nanoparticles deposited on transition metal oxides.

PubMed

Akita, Tomoki; Kohyama, Masanori; Haruta, Masatake

2013-08-20

Many researchers have investigated the catalytic performance of gold nanoparticles (GNPs) supported on metal oxides for various catalytic reactions of industrial importance. These studies have consistently shown that the catalytic activity and selectivity depend on the size of GNPs, the kind of metal oxide supports, and the gold/metal oxide interface structure. Although researchers have proposed several structural models for the catalytically active sites and have identified the specific electronic structures of GNPs induced by the quantum effect, recent experimental and theoretical studies indicate that the perimeter around GNPs in contact with the metal oxide supports acts as an active site in many reactions. Thus, it is of immense importance to investigate the detailed structures of the perimeters and the contact interfaces of gold/metal oxide systems by using electron microscopy at an atomic scale. This Account describes our investigation, at the atomic scale using electron microscopy, of GNPs deposited on metal oxides. In particular, high-resolution transmission electron microscopy (HRTEM) and high-angle annular dark-field scanning transmission electron microscopy (HAADF-STEM) are valuable tools to observe local atomic structures, as has been successfully demonstrated for various nanoparticles, surfaces, and material interfaces. TEM can be applied to real powder catalysts as received without making special specimens, in contrast to what is typically necessary to observe bulk materials. For precise structure analyses at an atomic scale, model catalysts prepared by using well-defined single-crystalline substrates are also adopted for TEM observations. Moreover, aberration-corrected TEM, which has high spatial resolution under 0.1 nm, is a promising tool to observe the interface structure between GNPs and metal oxide supports including oxygen atoms at the interfaces. The oxygen atoms in particular play an important role in the behavior of gold/metal oxide interfaces, because they may participate in catalytic reaction steps. Detailed information about the interfacial structures between GNPs and metal oxides provides valuable structure models for theoretical calculations which can elucidate the local electronic structure effective for activating a reactant molecule. Based on our observations with HRTEM and HAADF-STEM, we report the detailed structure of gold/metal oxide interfaces.

eV-TEM: Transmission electron microscopy in a low energy cathode lens instrument.

PubMed

Geelen, Daniël; Thete, Aniket; Schaff, Oliver; Kaiser, Alexander; van der Molen, Sense Jan; Tromp, Rudolf

2015-12-01

We are developing a transmission electron microscope that operates at extremely low electron energies, 0-40 eV. We call this technique eV-TEM. Its feasibility is based on the fact that at very low electron energies the number of energy loss pathways decreases. Hence, the electron inelastic mean free path increases dramatically. eV-TEM will enable us to study elastic and inelastic interactions of electrons with thin samples. With the recent development of aberration correction in cathode lens instruments, a spatial resolution of a few nm appears within range, even for these very low electron energies. Such resolution will be highly relevant to study biological samples such as proteins and cell membranes. The low electron energies minimize adverse effects due to radiation damage. Copyright © 2015. Published by Elsevier B.V.

Mega-electron-volt ultrafast electron diffraction at SLAC National Accelerator Laboratory

DOE Office of Scientific and Technical Information (OSTI.GOV)

Weathersby, S. P.; Brown, G.; Chase, T. F.

Ultrafast electron probes are powerful tools, complementary to x-ray free-electron lasers, used to study structural dynamics in material, chemical, and biological sciences. High brightness, relativistic electron beams with femtosecond pulse duration can resolve details of the dynamic processes on atomic time and length scales. SLAC National Accelerator Laboratory recently launched the Ultrafast Electron Diffraction (UED) and microscopy Initiative aiming at developing the next generation ultrafast electron scattering instruments. As the first stage of the Initiative, a mega-electron-volt (MeV) UED system has been constructed and commissioned to serve ultrafast science experiments and instrumentation development. The system operates at 120-Hz repetition ratemore » with outstanding performance. In this paper, we report on the SLAC MeV UED system and its performance, including the reciprocal space resolution, temporal resolution, and machine stability.« less

Mega-electron-volt ultrafast electron diffraction at SLAC National Accelerator Laboratory.

PubMed

Weathersby, S P; Brown, G; Centurion, M; Chase, T F; Coffee, R; Corbett, J; Eichner, J P; Frisch, J C; Fry, A R; Gühr, M; Hartmann, N; Hast, C; Hettel, R; Jobe, R K; Jongewaard, E N; Lewandowski, J R; Li, R K; Lindenberg, A M; Makasyuk, I; May, J E; McCormick, D; Nguyen, M N; Reid, A H; Shen, X; Sokolowski-Tinten, K; Vecchione, T; Vetter, S L; Wu, J; Yang, J; Dürr, H A; Wang, X J

2015-07-01

Ultrafast electron probes are powerful tools, complementary to x-ray free-electron lasers, used to study structural dynamics in material, chemical, and biological sciences. High brightness, relativistic electron beams with femtosecond pulse duration can resolve details of the dynamic processes on atomic time and length scales. SLAC National Accelerator Laboratory recently launched the Ultrafast Electron Diffraction (UED) and microscopy Initiative aiming at developing the next generation ultrafast electron scattering instruments. As the first stage of the Initiative, a mega-electron-volt (MeV) UED system has been constructed and commissioned to serve ultrafast science experiments and instrumentation development. The system operates at 120-Hz repetition rate with outstanding performance. In this paper, we report on the SLAC MeV UED system and its performance, including the reciprocal space resolution, temporal resolution, and machine stability.

Reprint of: Atmospheric scanning electron microscope observes cells and tissues in open medium through silicon nitride film.

PubMed

Nishiyama, Hidetoshi; Suga, Mitsuo; Ogura, Toshihiko; Maruyama, Yuusuke; Koizumi, Mitsuru; Mio, Kazuhiro; Kitamura, Shinichi; Sato, Chikara

2010-11-01

Direct observation of subcellular structures and their characterization is essential for understanding their physiological functions. To observe them in open environment, we have developed an inverted scanning electron microscope with a detachable, open-culture dish, capable of 8 nm resolution, and combined with a fluorescence microscope quasi-simultaneously observing the same area from the top. For scanning electron microscopy from the bottom, a silicon nitride film window in the base of the dish maintains a vacuum between electron gun and open sample dish while allowing electrons to pass through. Electrons are backscattered from the sample and captured by a detector under the dish. Cells cultured on the open dish can be externally manipulated under optical microscopy, fixed, and observed using scanning electron microscopy. Once fine structures have been revealed by scanning electron microscopy, their component proteins may be identified by comparison with separately prepared fluorescence-labeled optical microscopic images of the candidate proteins, with their heavy-metal-labeled or stained ASEM images. Furthermore, cell nuclei in a tissue block stained with platinum-blue were successfully observed without thin-sectioning, which suggests the applicability of this inverted scanning electron microscope to cancer diagnosis. This microscope visualizes mesoscopic-scale structures, and is also applicable to non-bioscience fields including polymer chemistry. Copyright © 2010 Elsevier Inc. All rights reserved.

Epidermal growth factor receptor subunit locations determined in hydrated cells with environmental scanning electron microscopy.

PubMed

Peckys, Diana B; Baudoin, Jean-Pierre; Eder, Magdalena; Werner, Ulf; de Jonge, Niels

2013-01-01

Imaging single epidermal growth factor receptors (EGFR) in intact cells is presently limited by the available microscopy methods. Environmental scanning electron microscopy (ESEM) of whole cells in hydrated state in combination with specific labeling with gold nanoparticles was used to localize activated EGFRs in the plasma membranes of COS7 and A549 cells. The use of a scanning transmission electron microscopy (STEM) detector yielded a spatial resolution of 3 nm, sufficient to identify the locations of individual EGFR dimer subunits. The sizes and distribution of dimers and higher order clusters of EGFRs were determined. The distance between labels bound to dimers amounted to 19 nm, consistent with a molecular model. A fraction of the EGFRs was found in higher order clusters with sizes ranging from 32-56 nm. ESEM can be used for quantitative whole cell screening studies of membrane receptors, and for the study of nanoparticle-cell interactions in general.

Epidermal growth factor receptor subunit locations determined in hydrated cells with environmental scanning electron microscopy

PubMed Central

Peckys, Diana B.; Baudoin, Jean-Pierre; Eder, Magdalena; Werner, Ulf; de Jonge, Niels

2013-01-01

Imaging single epidermal growth factor receptors (EGFR) in intact cells is presently limited by the available microscopy methods. Environmental scanning electron microscopy (ESEM) of whole cells in hydrated state in combination with specific labeling with gold nanoparticles was used to localize activated EGFRs in the plasma membranes of COS7 and A549 cells. The use of a scanning transmission electron microscopy (STEM) detector yielded a spatial resolution of 3 nm, sufficient to identify the locations of individual EGFR dimer subunits. The sizes and distribution of dimers and higher order clusters of EGFRs were determined. The distance between labels bound to dimers amounted to 19 nm, consistent with a molecular model. A fraction of the EGFRs was found in higher order clusters with sizes ranging from 32–56 nm. ESEM can be used for quantitative whole cell screening studies of membrane receptors, and for the study of nanoparticle-cell interactions in general. PMID:24022088

Direct observation of dopant distribution in GaAs compound semiconductors using phase-shifting electron holography and Lorentz microscopy.

PubMed

Sasaki, Hirokazu; Otomo, Shinya; Minato, Ryuichiro; Yamamoto, Kazuo; Hirayama, Tsukasa

2014-06-01

Phase-shifting electron holography and Lorentz microscopy were used to map dopant distributions in GaAs compound semiconductors with step-like dopant concentration. Transmission electron microscope specimens were prepared using a triple beam focused ion beam (FIB) system, which combines a Ga ion beam, a scanning electron microscope, and an Ar ion beam to remove the FIB damaged layers. The p-n junctions were clearly observed in both under-focused and over-focused Lorentz microscopy images. A phase image was obtained by using a phase-shifting reconstruction method to simultaneously achieve high sensitivity and high spatial resolution. Differences in dopant concentrations between 1 × 10(19) cm(-3) and 1 × 10(18) cm(-3) regions were clearly observed by using phase-shifting electron holography. We also interpreted phase profiles quantitatively by considering inactive layers induced by ion implantation during the FIB process. The thickness of an inactive layer at different dopant concentration area can be measured from the phase image. © The Author 2014. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Active pixel sensor array as a detector for electron microscopy.

PubMed

Milazzo, Anna-Clare; Leblanc, Philippe; Duttweiler, Fred; Jin, Liang; Bouwer, James C; Peltier, Steve; Ellisman, Mark; Bieser, Fred; Matis, Howard S; Wieman, Howard; Denes, Peter; Kleinfelder, Stuart; Xuong, Nguyen-Huu

2005-09-01

A new high-resolution recording device for transmission electron microscopy (TEM) is urgently needed. Neither film nor CCD cameras are systems that allow for efficient 3-D high-resolution particle reconstruction. We tested an active pixel sensor (APS) array as a replacement device at 200, 300, and 400 keV using a JEOL JEM-2000 FX II and a JEM-4000 EX electron microscope. For this experiment, we used an APS prototype with an area of 64 x 64 pixels of 20 microm x 20 microm pixel pitch. Single-electron events were measured by using very low beam intensity. The histogram of the incident electron energy deposited in the sensor shows a Landau distribution at low energies, as well as unexpected events at higher absorbed energies. After careful study, we concluded that backscattering in the silicon substrate and re-entering the sensitive epitaxial layer a second time with much lower speed caused the unexpected events. Exhaustive simulation experiments confirmed the existence of these back-scattered electrons. For the APS to be usable, the back-scattered electron events must be eliminated, perhaps by thinning the substrate to less than 30 microm. By using experimental data taken with an APS chip with a standard silicon substrate (300 microm) and adjusting the results to take into account the effect of a thinned silicon substrate (30 microm), we found an estimate of the signal-to-noise ratio for a back-thinned detector in the energy range of 200-400 keV was about 10:1 and an estimate for the spatial resolution was about 10 microm.

Super-resolution optical microscopy study of telomere structure.

PubMed

Phipps, Mary Lisa; Goodwin, Peter M; Martinez, Jennifer S; Goodwin, Edwin H

2016-09-01

Chromosome ends are shielded from exonucleolytic attack and inappropriate end-joining by terminal structures called telomeres; these structures are potential targets for anticancer drugs. Telomeres are composed of a simple DNA sequence (5?-TTAGGG-3? in humans) repeated more than a thousand times, a short 3? single-stranded overhang, and numerous proteins. Electron microscopy has shown that the 3? overhang pairs with the complementary strand at an internal site creating a small displacement loop and a large double-stranded “t-loop.” Our goal is to determine whether all telomeres adopt the t-loop configuration, or whether there are two or more distinct configurations. Progress in optimizing super-resolution (SR) microscopy for this ongoing investigation is reported here. Results suggest that under certain conditions sample preparation procedures may disrupt chromatin by causing loss of nucleosomes. This finding may limit the use of SR microscopy in telomere studies.

Super-resolution optical microscopy study of telomere structure

NASA Astrophysics Data System (ADS)

Phipps, Mary Lisa; Goodwin, Peter M.; Martinez, Jennifer S.; Goodwin, Edwin H.

2016-09-01

Chromosome ends are shielded from exonucleolytic attack and inappropriate end-joining by terminal structures called telomeres; these structures are potential targets for anticancer drugs. Telomeres are composed of a simple DNA sequence (5‧-TTAGGG-3‧ in humans) repeated more than a thousand times, a short 3‧ single-stranded overhang, and numerous proteins. Electron microscopy has shown that the 3‧ overhang pairs with the complementary strand at an internal site creating a small displacement loop and a large double-stranded "t-loop." Our goal is to determine whether all telomeres adopt the t-loop configuration, or whether there are two or more distinct configurations. Progress in optimizing super-resolution (SR) microscopy for this ongoing investigation is reported here. Results suggest that under certain conditions sample preparation procedures may disrupt chromatin by causing loss of nucleosomes. This finding may limit the use of SR microscopy in telomere studies.

Synchrotron X-ray imaging of nanomagnetism in meteoritic metal (Invited)

NASA Astrophysics Data System (ADS)

Bryson, J. F.; Herrero Albillos, J.; Kronast, F.; Tyliszczak, T.; Redfern, S. A.; van der Laan, G.; Harrison, R. J.

2013-12-01

It is becoming increasingly apparent that a wealth of paleomagnetic information is stored at the nanoscale within natural samples. To date, this nanopaleomagetism has been investigated using high resolution magnetic microscopies, such as electron holography. Although unparalleled in its spatial resolution, electron holography produces images that are indirectly related to the magnetisation state of the sample, introducing ambiguity when interpreting magnetisation information. Holography also requires extensive off-line processing, making it unsuitable for studying dynamic processes, and the sample preparation negates the study of natural remanences. Here we demonstrate the capabilities of a new generation of nanomagnetic imaging methods using synchrotron X-ray radiation. X-rays tuned to an elemental absorption edge can display differing excitation probabilities depending on the orientation of an electron's magnetic moment relative to that of the X-ray beam. This is achieved by introducing an angular momentum to the photon through circular polarisation, resulting in an absorption signal that is proportional to the projection of the magnetic moment on to the X-ray beam direction. We introduce and compare two experimental set-ups capable of spatially resolving these signals to form a high-resolution magnetisation map: photoemission electron microscopy and scanning transmission electron microscopy. Both techniques provide measurements of magnetisation with 30-50nm resolution and elemental specificity. Photoemission electron microscopy can be used also to create maps of all three of the spatial components of magnetisation and investigate dynamic magnetic switching processes. The full capabilities of X-ray imaging are demonstrated through the application of both of these techniques to meteoritic metal. We show that the 'cloudy zone' within iron meteorites contains nanoscale islands of tetrataenite (FeNi) that are populated equally by all three possible magnetic easy axes, suggesting that there were no stray fields (either magnetic or stress) effecting the magnetisation during cloudy zone formation. This observation allows for dynamo field information to be extracted from X-ray nanomagnetic images of the cloudy zone in metallic inclusions within certain chondritic meteorites, as it implies that any deviation from the randomly populated easy axis distribution can be assigned to an external dynamo field. As the cloudy zone forms over 10-100 Ma, this observation suggests that X-ray imaging of the nanopaleomagentism in these meteorites could provide an elegant and concise relative measure of asteroid dynamo field direction and strength over this entire time period, revolutionising our understanding of dynamo processes and planetary formation.

High resolution electron microscopy of a small crack at the superficial layer of enamel.

PubMed

Hayashi, Y

1994-12-01

A small enamel crack was investigated using a high resolution electron microscope. The inside of the crack was filled with aggregates of irregularly oriented plate-like crystals. Amorphous mineral deposits were observed among these aggregates at a low magnification. Selected area electron diffractions indicated that the plate-like crystals consisted of hydroxyapatite (OH-AP), and that the amorphous mineral deposits were a mixture of OH-AP and whitlockite. These findings indicate that this crack may have been formed by occlusal and/or masticatory stress, and that a natural occlusion might occur through mineral deposition at the small crack such as in this case.

He-Ion Microscopy as a High-Resolution Probe for Complex Quantum Heterostructures in Core-Shell Nanowires.

PubMed

Pöpsel, Christian; Becker, Jonathan; Jeon, Nari; Döblinger, Markus; Stettner, Thomas; Gottschalk, Yeanitza Trujillo; Loitsch, Bernhard; Matich, Sonja; Altzschner, Marcus; Holleitner, Alexander W; Finley, Jonathan J; Lauhon, Lincoln J; Koblmüller, Gregor

2018-06-13

Core-shell semiconductor nanowires (NW) with internal quantum heterostructures are amongst the most complex nanostructured materials to be explored for assessing the ultimate capabilities of diverse ultrahigh-resolution imaging techniques. To probe the structure and composition of these materials in their native environment with minimal damage and sample preparation calls for high-resolution electron or ion microscopy methods, which have not yet been tested on such classes of ultrasmall quantum nanostructures. Here, we demonstrate that scanning helium ion microscopy (SHeIM) provides a powerful and straightforward method to map quantum heterostructures embedded in complex III-V semiconductor NWs with unique material contrast at ∼1 nm resolution. By probing the cross sections of GaAs-Al(Ga)As core-shell NWs with coaxial GaAs quantum wells as well as short-period GaAs/AlAs superlattice (SL) structures in the shell, the Al-rich and Ga-rich layers are accurately discriminated by their image contrast in excellent agreement with correlated, yet destructive, scanning transmission electron microscopy and atom probe tomography analysis. Most interestingly, quantitative He-ion dose-dependent SHeIM analysis of the ternary AlGaAs shell layers and of compositionally nonuniform GaAs/AlAs SLs reveals distinct alloy composition fluctuations in the form of Al-rich clusters with size distributions between ∼1-10 nm. In the GaAs/AlAs SLs the alloy clustering vanishes with increasing SL-period (>5 nm-GaAs/4 nm-AlAs), providing insights into critical size dimensions for atomic intermixing effects in short-period SLs within a NW geometry. The straightforward SHeIM technique therefore provides unique benefits in imaging the tiniest nanoscale features in topography, structure and composition of a multitude of diverse complex semiconductor nanostructures.

Evaluation of Characterization Techniques for Carbon-Carbon Composites

DTIC Science & Technology

1992-05-01

Enhancement of Resin (50X) 51 28 Confocal Image of Reticulated , Vitreous Carbon Foam 53 29 Schemmtic Principle of Backscattered Electron Microscopy for...future. 7.2 Confocal Microscopy Both carbon - carbon composites and reticulated vitreous carbon foams were submitted to Sarastro, Inc. to evaluate...indicate 1-micron resolutions are possible; however, the depth penetration is limited even further at these parameters. Six reticulated vitreous carbon

Atomic Structure and Properties of Extended Defects in Silicon

DOE Office of Scientific and Technical Information (OSTI.GOV)

Buczko, R.; Chisholm, M.F.; Kaplan, T.

1998-10-15

The Z-contrast technique represents a new approach to high-resolution electron microscopy allowing for the first time incoherent imaging of materials on the atomic scale. The key advantages of the technique, an intrinsically higher resolution limit and directly interpretable, compositionally sensitive imaging, allow a new level of insight into the atomic configurations of extended defects in silicon. This experimental technique has been combined with theoretical calculations (a combination of first principles, tight binding, and classical methods) to extend this level of insight by obtaining the energetic and electronic structure of the defects.

Visualization of early influenza A virus trafficking in human dendritic cells using STED microscopy.

PubMed

Baharom, Faezzah; Thomas, Oliver S; Lepzien, Rico; Mellman, Ira; Chalouni, Cécile; Smed-Sörensen, Anna

2017-01-01

Influenza A viruses (IAV) primarily target respiratory epithelial cells, but can also replicate in immune cells, including human dendritic cells (DCs). Super-resolution microscopy provides a novel method of visualizing viral trafficking by overcoming the resolution limit imposed by conventional light microscopy, without the laborious sample preparation of electron microscopy. Using three-color Stimulated Emission Depletion (STED) microscopy, we visualized input IAV nucleoprotein (NP), early and late endosomal compartments (EEA1 and LAMP1 respectively), and HLA-DR (DC membrane/cytosol) by immunofluorescence in human DCs. Surface bound IAV were internalized within 5 min of infection. The association of virus particles with early endosomes peaked at 5 min when 50% of NP+ signals were also EEA1+. Peak association with late endosomes occurred at 15 min when 60% of NP+ signals were LAMP1+. At 30 min of infection, the majority of NP signals were in the nucleus. Our findings illustrate that early IAV trafficking in human DCs proceeds via the classical endocytic pathway.

Vertically grown nanowire crystals of dibenzotetrathienocoronene (DBTTC) on large-area graphene

DOE PAGES

Kim, B.; Chiu, C. -Y.; Kang, S. J.; ...

2016-06-01

Here we demonstrate controlled growth of vertical organic crystal nanowires on single layer graphene. Using Scanning Electron Microscopy (SEM), high-resolution transition electron microscopy (TEM), and Grazing Incidence X-ray Diffraction (GIXD), we probe the microstructure and morphology of dibenzotetrathienocoronene (DBTTC) nanowires epitaxially grown on graphene. The investigation is performed at both the ensemble and single nanowire level, and as function of growth parameters, providing insight of and control over the formation mechanism. Finally, the size, density and height of the nanowires can be tuned via growth conditions, opening new avenues for tailoring three-dimensional (3-D) nanostructured architectures for organic electronics with improvedmore » functional performance.« less

Controlled Synthesis of Atomically Layered Hexagonal Boron Nitride via Chemical Vapor Deposition.

PubMed

Liu, Juanjuan; Kutty, R Govindan; Liu, Zheng

2016-11-29

Hexagonal boron nitrite (h-BN) is an attractive material for many applications including electronics as a complement to graphene, anti-oxidation coatings, light emitters, etc. However, the synthesis of high-quality h-BN is still a great challenge. In this work, via controlled chemical vapor deposition, we demonstrate the synthesis of h-BN films with a controlled thickness down to atomic layers. The quality of as-grown h-BN is confirmed by complementary characterizations including high-resolution transition electron microscopy, atomic force microscopy, Raman spectroscopy and X-ray photo-electron spectroscopy. This work will pave the way for production of large-scale and high-quality h-BN and its applications as well.

Electrical conduction at domain walls in multiferroic BiFeO3

NASA Astrophysics Data System (ADS)

Seidel, Jan; Martin, Lane; He, Qing; Zhan, Qian; Chu, Ying-Hao; Rother, Axel; Hawkridge, Michael; Maksymovych, Peter; Yu, Pu; Gajek, Martin; Balke, Nina; Kalinin, Sergei; Gemming, Sybille; Wang, Feng; Catalán, Gustau; Scott, James; Spaldin, Nicola; Orenstein, Joseph; Ramesh, Ramamoorthy

2009-03-01

We report the observation of room temperature electronic conductivity at ferroelectric domain walls in BiFeO3. The origin and nature of the observed conductivity is probed using a combination of conductive atomic force microscopy, high resolution transmission electron microscopy and first-principles density functional computations. We show that a structurally driven change in both the electrostatic potential and local electronic structure (i.e., a decrease in band gap) at the domain wall leads to the observed electrical conductivity. We estimate the conductivity in the wall to be several orders of magnitude higher than for the bulk material. Additionally we demonstrate the potential for device applications of such conducting nanoscale features.

Creating a single twin boundary between two CdTe (111) wafers with controlled rotation angle by wafer bonding

NASA Astrophysics Data System (ADS)

Sun, Ce; Lu, Ning; Wang, Jinguo; Lee, Jihyung; Peng, Xin; Klie, Robert F.; Kim, Moon J.

2013-12-01

The single twin boundary with crystallographic orientation relationship (1¯1¯1¯)//(111) [01¯1]//[011¯] was created by wafer bonding. Electron diffraction patterns and high-resolution transmission electron microscopy images demonstrated the well control of the rotation angle between the bonded pair. At the twin boundary, one unit of wurtzite structure was found between two zinc-blende matrices. High-angle annular dark-field scanning transmission electron microscopy images showed Cd- and Te-terminated for the two bonded portions, respectively. The I-V curve across the twin boundary showed increasingly nonlinear behavior, indicating a potential barrier at the bonded twin boundary.

Conventions and workflows for using Situs

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wriggers, Willy, E-mail: wriggers@biomachina.org

2012-04-01

Recent developments of the Situs software suite for multi-scale modeling are reviewed. Typical workflows and conventions encountered during processing of biophysical data from electron microscopy, tomography or small-angle X-ray scattering are described. Situs is a modular program package for the multi-scale modeling of atomic resolution structures and low-resolution biophysical data from electron microscopy, tomography or small-angle X-ray scattering. This article provides an overview of recent developments in the Situs package, with an emphasis on workflows and conventions that are important for practical applications. The modular design of the programs facilitates scripting in the bash shell that allows specific programs tomore » be combined in creative ways that go beyond the original intent of the developers. Several scripting-enabled functionalities, such as flexible transformations of data type, the use of symmetry constraints or the creation of two-dimensional projection images, are described. The processing of low-resolution biophysical maps in such workflows follows not only first principles but often relies on implicit conventions. Situs conventions related to map formats, resolution, correlation functions and feature detection are reviewed and summarized. The compatibility of the Situs workflow with CCP4 conventions and programs is discussed.« less

A Chemical and Structural Study of the A1N-Si Interface

NASA Technical Reports Server (NTRS)

George, T.; Beye, R.

1997-01-01

Samples of A1N grown on silicon [111] subtrates were examined using electron enery loss spectroscopy (EELS) and selected area diffraction (SAD) with high-resolution transmission electron microscopy (TEM) to determine the source of out-of-place tilts and in-plane rotations of the A1N crystallites at the Si interface.

An inexpensive approach for bright-field and dark-field imaging by scanning transmission electron microscopy in scanning electron microscopy.

PubMed

Patel, Binay; Watanabe, Masashi

2014-02-01

Scanning transmission electron microscopy in scanning electron microscopy (STEM-in-SEM) is a convenient technique for soft materials characterization. Various specimen-holder geometries and detector arrangements have been used for bright-field (BF) STEM-in-SEM imaging. In this study, to further the characterization potential of STEM-IN-SEM, a new specimen holder has been developed to facilitate direct detection of BF signals and indirect detection of dark-field (DF) signals without the need for substantial instrument modification. DF imaging is conducted with the use of a gold (Au)-coated copper (Cu) plate attached to the specimen holder which directs highly scattered transmitted electrons to an off-axis yttrium-aluminum-garnet (YAG) detector. A hole in the copper plate allows for BF imaging with a transmission electron (TE) detector. The inclusion of an Au-coated Cu plate enhanced DF signal intensity. Experiments validating the acquisition of true DF signals revealed that atomic number (Z) contrast may be achieved for materials with large lattice spacing. However, materials with small lattice spacing still exhibit diffraction contrast effects in this approach. The calculated theoretical fine probe size is 1.8 nm. At 30 kV, in this indirect approach, DF spatial resolution is limited to 3.2 nm as confirmed experimentally.

Mechanically adjustable single-molecule transistors and stencil mask nanofabrication of high-resolution scanning probes

NASA Astrophysics Data System (ADS)

Champagne, Alexandre

This dissertation presents the development of two original experimental techniques to probe nanoscale objects. The first one studies electronic transport in single organic molecule transistors in which the source-drain electrode spacing is mechanically adjustable. The second involves the fabrication of high-resolution scanning probe microscopy sensors using a stencil mask lithography technique. We describe the fabrication of transistors in which a single organic molecule can be incorporated. The source and drain leads of these transistors are freely suspended above a flexible substrate, and their spacing can be adjusted by bending the substrate. We detail the technology developed to carry out measurements on these samples. We study electronic transport in single C60 molecules at low temperature. We observe Coulomb blockaded transport and can resolve the discrete energy spectrum of the molecule. We are able to mechanically tune the spacing between the electrodes (over a range of 5 A) to modulate the lead-molecule coupling, and can electrostatically tune the energy levels on the molecule by up to 160 meV using a gate electrode. Initial progress in studying different transport regimes in other molecules is also discussed. We present a lithographic process that allows the deposition of metal nanostructures with a resolution down to 10 nm directly onto atomic force microscope (AFM) tips. We show that multiple layers of lithography can be deposited and aligned. We fabricate high-resolution magnetic force microscopy (MFM) probes using this method and discuss progress to fabricate other scanning probe microscopy (SPM) sensors.

High-resolution imaging of silicene on an Ag(111) surface by atomic force microscopy

NASA Astrophysics Data System (ADS)

Onoda, Jo; Yabuoshi, Keisuke; Miyazaki, Hiroki; Sugimoto, Yoshiaki

2017-12-01

Silicene, a two-dimensional (2D) honeycomb arrangement of Si atoms, is expected to have better electronic properties than graphene and has been mostly synthesized on Ag surfaces. Although scanning tunneling microscopy (STM) has been used for visualizing its atomic structure in real space, the interpretation of STM contrast is not straightforward and only the topmost Si atoms were observed on the (4 ×4 ) silicene/Ag(111) surface. Here, we demonstrate that high-resolution atomic force microscopy (AFM) can resolve all constituent Si atoms in the buckled honeycomb arrangement of the (4 ×4 ) silicene. Site-specific force spectroscopy attributes the origin of the high-resolution AFM images to chemical bonds between the AFM probe apex and the individual Si atoms on the (4 ×4 ) silicene. A detailed analysis of the geometric parameters suggests that the pulling up of lower-buckled Si atoms by the AFM tip could be a key for high-resolution AFM, implying a weakening of the Si-Ag interactions at the interface. We expect that high-resolution AFM will also unveil atomic structures of edges and defects of silicene, or other emerging 2D materials.

Electric field imaging of single atoms

PubMed Central

Shibata, Naoya; Seki, Takehito; Sánchez-Santolino, Gabriel; Findlay, Scott D.; Kohno, Yuji; Matsumoto, Takao; Ishikawa, Ryo; Ikuhara, Yuichi

2017-01-01

In scanning transmission electron microscopy (STEM), single atoms can be imaged by detecting electrons scattered through high angles using post-specimen, annular-type detectors. Recently, it has been shown that the atomic-scale electric field of both the positive atomic nuclei and the surrounding negative electrons within crystalline materials can be probed by atomic-resolution differential phase contrast STEM. Here we demonstrate the real-space imaging of the (projected) atomic electric field distribution inside single Au atoms, using sub-Å spatial resolution STEM combined with a high-speed segmented detector. We directly visualize that the electric field distribution (blurred by the sub-Å size electron probe) drastically changes within the single Au atom in a shape that relates to the spatial variation of total charge density within the atom. Atomic-resolution electric field mapping with single-atom sensitivity enables us to examine their detailed internal and boundary structures. PMID:28555629

Synthesis and Characteristics of ZnS Nanospheres for Heterojunction Photovoltaic Device

NASA Astrophysics Data System (ADS)

Chou, Sheng-Hung; Hsiao, Yu-Jen; Fang, Te-Hua; Chou, Po-Hsun

2015-06-01

The synthesis of ZnS nanospheres produced using the microwave hydrothermal method was studied. The microstructure and surface and optical properties of ZnS nanospheres on glass were characterized using scanning electron microscopy, high-resolution transmission electron microscopy, x-ray diffraction, and ultraviolet-visible spectroscopy. The influence of deposition time on the transmission and photovoltaic performance was determined. The power conversion efficiency of an Al-doped ZnO/ZnS nanosphere/textured p-Si device improved from 0.93 to 1.77% when the thickness of the ZnS nanostructured film was changed from 75 to 150 nm.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kendall, Amy; McDonald, Michele; Bian, Wen

Flexible filamentous viruses make up a large fraction of the known plant viruses, but in comparison with those of other viruses, very little is known about their structures. We have used fiber diffraction, cryo-electron microscopy, and scanning transmission electron microscopy to determine the symmetry of a potyvirus, soybean mosaic virus; to confirm the symmetry of a potexvirus, potato virus X; and to determine the low-resolution structures of both viruses. We conclude that these viruses and, by implication, most or all flexible filamentous plant viruses share a common coat protein fold and helical symmetry, with slightly less than 9 subunits permore » helical turn.« less

The Antibacterial Polyamide 6-ZnO Hierarchical Nanofibers Fabricated by Atomic Layer Deposition and Hydrothermal Growth

NASA Astrophysics Data System (ADS)

Wang, Zhengduo; Zhang, Li; Liu, Zhongwei; Sang, Lijun; Yang, Lizhen; Chen, Qiang

2017-06-01

In this paper, we report the combination of atomic layer deposition (ALD) with hydrothermal techniques to deposit ZnO on electrospun polyamide 6 (PA 6) nanofiber (NF) surface in the purpose of antibacterial application. The micro- and nanostructures of the hierarchical fibers are characterized by field emission scanning electron microscopy (FE-SEM), high-resolution transmission electron microscopy (HRTEM), and scanning transmission electron microscopy (STEM). We find that NFs can grow into "water lily"- and "caterpillar"-like shapes, which depend on the number of ALD cycles and the hydrothermal reaction period. It is believed that the thickness of ZnO seed layer by ALD process and the period in hydrothermal reaction have the same importance in crystalline growth and hierarchical fiber formation. The tests of antibacterial activity demonstrate that the ZnO/PA 6 core-shell composite fabricated by the combination of ALD with hydrothermal are markedly efficient in suppressing bacteria survivorship.

Synthesis, structural and optical properties of nanocrystalline vanadium doped zinc oxide aerogel

NASA Astrophysics Data System (ADS)

El Ghoul, J.; Barthou, C.; El Mir, L.

2012-06-01

We report the synthesis of vanadium-doped ZnO nanoparticles prepared by a sol-gel processing technique. In our approach, the water for hydrolysis was slowly released by esterification reaction followed by a supercritical drying in ethyl alcohol. Vanadium doping concentration of 10 at% has been investigated. After treatment in air at different temperatures, the obtained nanopowder was characterized by various techniques such as scanning electron microscopy (SEM), transmission electron microscopy (TEM), X-ray diffraction (XRD) and photoluminescence (PL). Analysis by scanning electron microscopy at high resolution shows that the grain size increases with increasing temperature. Thus, in the case of thermal treatment at 500 °C in air, the powder with an average particle size of 25 nm shows a strong luminescence band in the visible range. The intensity and energy position of the obtained PL band depends on the temperature measurement increase. The mechanism of this emission band is discussed.

A simple procedure to analyze positions of interest in infectious cell cultures by correlative light and electron microscopy.

PubMed

Madela, Kazimierz; Banhart, Sebastian; Zimmermann, Anja; Piesker, Janett; Bannert, Norbert; Laue, Michael

2014-01-01

Plastic cell culture dishes that contain a thin bottom of highest optical quality including an imprinted finder grid (μ-Dish Grid-500) are optimally suited for routine correlative light and electron microscopy using chemical fixation. Such dishes allow high-resolution fluorescence and bright-field imaging using fixed and living cells and are compatible with standard protocols for scanning and transmission electron microscopy. Ease of use during cell culture and imaging, as well as a tight cover render the dishes particularly suitable for working with infectious organisms up to the highest biosafety level. Detailed protocols are provided and demonstrated by showing two examples: monitoring the production of virus-like particles of the Human Endogenous Retrovirus HERV-K(HML-2) by HeLa cells and investigation of Rab11-positive membrane-compartments of HeLa cells after infection with Chlamydia trachomatis. © 2014 Elsevier Inc. All rights reserved.

One-pot template-free synthesis of uniform-sized fullerene-like magnetite hollow spheres

NASA Astrophysics Data System (ADS)

Zhu, Qing; Zhang, Yue; Liu, Zheng; Zhou, Xinrui; Zhang, Xinmei; Zeng, Lintao

2015-11-01

Uniform-sized Fe3O4 hollow spheres with average diameter of 250 nm and shell thickness of ∼50 nm have been successfully synthesized through a simple hydrothermal route with the presence of di-n-propylamine (DPA) as a weak-base. The reaction time and DPA amount play important roles in the formation of the magnetite hollow spheres. The structures of the products were characterized by X-ray diffraction, X-ray photoelectron spectroscopy, Fourier transform infrared spectra, scanning electron microscopy, transmission electron microscopy, and high-resolution transmission electron microscopy. The results show that the single-crystalline Fe3O4 hollow spheres are composed of well-aligned magnetite nanoparticles (NPs). The magnetic property investigation shows that these hollow spheres have a higher saturation magnetization (Ms) than the solid spheres. Furthermore, a possible mechanism for the formation of magnetite hollow spheres is proposed based on the experimental observations.

Transfer doping of single isolated nanodiamonds, studied by scanning probe microscopy techniques

NASA Astrophysics Data System (ADS)

Bolker, Asaf; Saguy, Cecile; Kalish, Rafi

2014-09-01

The transfer doping of diamond surfaces has been applied in various novel two-dimensional electronic devices. Its extension to nanodiamonds (ND) is essential for ND-based applications in many fields. In particular, understanding the influence of the crystallite size on transfer doping is desirable. Here, we report the results of a detailed study of the electronic energetic band structure of single, isolated transfer-doped nanodiamonds with nanometric resolution using a combination of scanning tunneling spectroscopy and Kelvin force microscopy measurements. The results show how the band gap, the valence band maximum, the electron affinity and the work function all depend on the ND’s size and nanoparticle surface properties. The present analysis, which combines information from both scanning tunneling spectroscopy and Kelvin force microscopy, should be applicable to any nanoparticle or surface that can be measured with scanning probe techniques.

Novel Solid Electrolytes for Li-Ion Batteries: A Perspective from Electron Microscopy Studies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ma, Cheng; Chi, Miaofang

2016-06-08

Solid electrolytes can simultaneously overcome two of the most formidable challenges of Li-ion batteries: the severe safety issues and insufficient energy densities. However, before they can be implemented in actual batteries, the ionic conductivity needs to be improved and the interface with electrodes must be optimized. The prerequisite for addressing these issues is a thorough understanding of the material’s behavior at the microscopic and/or the atomic level. (Scanning) transmission electron microscopy is a powerful tool for this purpose, as it can reach an ultrahigh spatial resolution. Here, we review recent electron microscopy investigations on the ion transport behavior in solidmore » electrolytes and their interfaces. Specifically, three aspects will be highlighted: the influence of grain interior atomic configuration on ionic conductivity, the contribution of grain boundaries, and the behavior of solid electrolyte/electrode interfaces. In conclusion, based on this, the perspectives for future research will be discussed.« less

Differentiation and Characterization of Excitatory and Inhibitory Synapses by Cryo-electron Tomography and Correlative Microscopy

PubMed Central

Sun, Rong; Zhang, Bin; Qi, Lei; Shivakoti, Sakar; Tian, Chong-Li; Lau, Pak-Ming

2018-01-01

As key functional units in neural circuits, different types of neuronal synapses play distinct roles in brain information processing, learning, and memory. Synaptic abnormalities are believed to underlie various neurological and psychiatric disorders. Here, by combining cryo-electron tomography and cryo-correlative light and electron microscopy, we distinguished intact excitatory and inhibitory synapses of cultured hippocampal neurons, and visualized the in situ 3D organization of synaptic organelles and macromolecules in their native state. Quantitative analyses of >100 synaptic tomograms reveal that excitatory synapses contain a mesh-like postsynaptic density (PSD) with thickness ranging from 20 to 50 nm. In contrast, the PSD in inhibitory synapses assumes a thin sheet-like structure ∼12 nm from the postsynaptic membrane. On the presynaptic side, spherical synaptic vesicles (SVs) of 25–60 nm diameter and discus-shaped ellipsoidal SVs of various sizes coexist in both synaptic types, with more ellipsoidal ones in inhibitory synapses. High-resolution tomograms obtained using a Volta phase plate and electron filtering and counting reveal glutamate receptor-like and GABAA receptor-like structures that interact with putative scaffolding and adhesion molecules, reflecting details of receptor anchoring and PSD organization. These results provide an updated view of the ultrastructure of excitatory and inhibitory synapses, and demonstrate the potential of our approach to gain insight into the organizational principles of cellular architecture underlying distinct synaptic functions. SIGNIFICANCE STATEMENT To understand functional properties of neuronal synapses, it is desirable to analyze their structure at molecular resolution. We have developed an integrative approach combining cryo-electron tomography and correlative fluorescence microscopy to visualize 3D ultrastructural features of intact excitatory and inhibitory synapses in their native state. Our approach shows that inhibitory synapses contain uniform thin sheet-like postsynaptic densities (PSDs), while excitatory synapses contain previously known mesh-like PSDs. We discovered “discus-shaped” ellipsoidal synaptic vesicles, and their distributions along with regular spherical vesicles in synaptic types are characterized. High-resolution tomograms further allowed identification of putative neurotransmitter receptors and their heterogeneous interaction with synaptic scaffolding proteins. The specificity and resolution of our approach enables precise in situ analysis of ultrastructural organization underlying distinct synaptic functions. PMID:29311144

Correlating Intravital Multi-Photon Microscopy to 3D Electron Microscopy of Invading Tumor Cells Using Anatomical Reference Points

PubMed Central

Karreman, Matthia A.; Mercier, Luc; Schieber, Nicole L.; Shibue, Tsukasa; Schwab, Yannick; Goetz, Jacky G.

2014-01-01

Correlative microscopy combines the advantages of both light and electron microscopy to enable imaging of rare and transient events at high resolution. Performing correlative microscopy in complex and bulky samples such as an entire living organism is a time-consuming and error-prone task. Here, we investigate correlative methods that rely on the use of artificial and endogenous structural features of the sample as reference points for correlating intravital fluorescence microscopy and electron microscopy. To investigate tumor cell behavior in vivo with ultrastructural accuracy, a reliable approach is needed to retrieve single tumor cells imaged deep within the tissue. For this purpose, fluorescently labeled tumor cells were subcutaneously injected into a mouse ear and imaged using two-photon-excitation microscopy. Using near-infrared branding, the position of the imaged area within the sample was labeled at the skin level, allowing for its precise recollection. Following sample preparation for electron microscopy, concerted usage of the artificial branding and anatomical landmarks enables targeting and approaching the cells of interest while serial sectioning through the specimen. We describe here three procedures showing how three-dimensional (3D) mapping of structural features in the tissue can be exploited to accurately correlate between the two imaging modalities, without having to rely on the use of artificially introduced markers of the region of interest. The methods employed here facilitate the link between intravital and nanoscale imaging of invasive tumor cells, enabling correlating function to structure in the study of tumor invasion and metastasis. PMID:25479106

Improving the oxidation resistance and stability of Ag nanoparticles by coating with multilayered reduced graphene oxide

NASA Astrophysics Data System (ADS)

Li, Yahui; Zhang, Huayu; Wu, Bowen; Guo, Zhuo

2017-12-01

A kind of coating nanostructure, Ag nanoparticles coated with multilayered reduced graphene oxide (RGO), is fabricated by employing a three-step reduction method in an orderly manner, which is significantly different from the conventional structures that are simply depositing or doping with Ag nanoparticles on RGO via chemical reduction. The as-prepared nanostructure is investigated by X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), transmission electron microscopy (TEM), high-resolution transmission electron microscopy (HRTEM), selected-area electronic diffraction (SEAD), scanning electron microscopy (SEM), and Fourier transform infrared spectroscopy (FTIR). The results show that the obtained Ag/RGO nanostructure is observed to be a perfect coating structure with well dispersed Ag particles, which is responsible for the remarkable oxidation resistance. The results of XPS spectra indicate the content of metallic Ag is far greater than that of Ag oxides despite of prolonged exposure to the air, which fully demonstrate the excellent stability of thus coating nanostructure.

A streaming multi-GPU implementation of image simulation algorithms for scanning transmission electron microscopy

DOE PAGES

Pryor, Alan; Ophus, Colin; Miao, Jianwei

2017-10-25

Simulation of atomic-resolution image formation in scanning transmission electron microscopy can require significant computation times using traditional methods. A recently developed method, termed plane-wave reciprocal-space interpolated scattering matrix (PRISM), demonstrates potential for significant acceleration of such simulations with negligible loss of accuracy. In this paper, we present a software package called Prismatic for parallelized simulation of image formation in scanning transmission electron microscopy (STEM) using both the PRISM and multislice methods. By distributing the workload between multiple CUDA-enabled GPUs and multicore processors, accelerations as high as 1000 × for PRISM and 15 × for multislice are achieved relative to traditionalmore » multislice implementations using a single 4-GPU machine. We demonstrate a potentially important application of Prismatic, using it to compute images for atomic electron tomography at sufficient speeds to include in the reconstruction pipeline. Prismatic is freely available both as an open-source CUDA/C++ package with a graphical user interface and as a Python package, PyPrismatic.« less

A streaming multi-GPU implementation of image simulation algorithms for scanning transmission electron microscopy.

PubMed

Pryor, Alan; Ophus, Colin; Miao, Jianwei

2017-01-01

Simulation of atomic-resolution image formation in scanning transmission electron microscopy can require significant computation times using traditional methods. A recently developed method, termed plane-wave reciprocal-space interpolated scattering matrix (PRISM), demonstrates potential for significant acceleration of such simulations with negligible loss of accuracy. Here, we present a software package called Prismatic for parallelized simulation of image formation in scanning transmission electron microscopy (STEM) using both the PRISM and multislice methods. By distributing the workload between multiple CUDA-enabled GPUs and multicore processors, accelerations as high as 1000 × for PRISM and 15 × for multislice are achieved relative to traditional multislice implementations using a single 4-GPU machine. We demonstrate a potentially important application of Prismatic , using it to compute images for atomic electron tomography at sufficient speeds to include in the reconstruction pipeline. Prismatic is freely available both as an open-source CUDA/C++ package with a graphical user interface and as a Python package, PyPrismatic .

Operando x-ray photoelectron emission microscopy for studying forward and reverse biased silicon p-n junctions.

PubMed

Barrett, N; Gottlob, D M; Mathieu, C; Lubin, C; Passicousset, J; Renault, O; Martinez, E

2016-05-01

Significant progress in the understanding of surfaces and interfaces of materials for new technologies requires operando studies, i.e., measurement of chemical, electronic, and magnetic properties under external stimulus (such as mechanical strain, optical illumination, or electric fields) applied in situ in order to approach real operating conditions. Electron microscopy attracts much interest, thanks to its ability to determine semiconductor doping at various scales in devices. Spectroscopic photoelectron emission microscopy (PEEM) is particularly powerful since it combines high spatial and energy resolution, allowing a comprehensive analysis of local work function, chemistry, and electronic structure using secondary, core level, and valence band electrons, respectively. Here we present the first operando spectroscopic PEEM study of a planar Si p-n junction under forward and reverse bias. The method can be used to characterize a vast range of materials at near device scales such as resistive oxides, conducting bridge memories and domain wall arrays in ferroelectrics photovoltaic devices.

A streaming multi-GPU implementation of image simulation algorithms for scanning transmission electron microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pryor, Alan; Ophus, Colin; Miao, Jianwei

Simulation of atomic-resolution image formation in scanning transmission electron microscopy can require significant computation times using traditional methods. A recently developed method, termed plane-wave reciprocal-space interpolated scattering matrix (PRISM), demonstrates potential for significant acceleration of such simulations with negligible loss of accuracy. In this paper, we present a software package called Prismatic for parallelized simulation of image formation in scanning transmission electron microscopy (STEM) using both the PRISM and multislice methods. By distributing the workload between multiple CUDA-enabled GPUs and multicore processors, accelerations as high as 1000 × for PRISM and 15 × for multislice are achieved relative to traditionalmore » multislice implementations using a single 4-GPU machine. We demonstrate a potentially important application of Prismatic, using it to compute images for atomic electron tomography at sufficient speeds to include in the reconstruction pipeline. Prismatic is freely available both as an open-source CUDA/C++ package with a graphical user interface and as a Python package, PyPrismatic.« less

CMC-coated Fe3O4 nanoparticles as new MRI probes for hepatocellular carcinoma

NASA Astrophysics Data System (ADS)

Sitthichai, Sudarat; Pilapong, Chalermchai; Thongtem, Titipun; Thongtem, Somchai

2015-11-01

Pure Fe3O4 nanoparticles and Fe3O4 magnetic nanoparticles (MNPs) coated with carboxymethyl cellulose (CMC) were successfully prepared by co-precipitating of FeCl2·4H2O and FeCl3·6H2O in the solutions containing ammonia at 80 °C for 3 h. Phase, morphology, particle-sized distribution, surface chemistry, and weight loss were characterized by X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), transmission electron microscopy (TEM) including high-resolution transmission electron microscopy (HRTEM) and selected area electron diffraction (SAED), thermogravimetric analysis (TGA), and Fourier transform infrared (FTIR) spectroscopy. In this research, CMC-coated Fe3O4 MNPs consisting of Fe2+ and Fe3+ ions with 543.3-mM-1 s-1 high relaxivity were detected and were able to be used for magnetic resonance imaging (MRI) application with very good contrast for targeting hepatocellular carcinoma (HCC) without any further vectorization.

Component analyses of urinary nanocrystallites of uric acid stone formers by combination of high-resolution transmission electron microscopy, fast Fourier transformation, energy dispersive X-ray spectroscopy, X-ray diffraction and Fourier transform infrared spectroscopy.

PubMed

Sun, Xin-Yuan; Xue, Jun-Fa; Xia, Zhi-Yue; Ouyang, Jian-Ming

2015-06-01

This study aimed to analyse the components of nanocrystallites in urines of patients with uric acid (UA) stones. X-ray diffraction (XRD), Fourier transform infrared spectroscopy, high-resolution transmission electron microscopy (HRTEM), fast Fourier transformation (FFT) of HRTEM, and energy dispersive X-ray spectroscopy (EDS) were performed to analyse the components of these nanocrystallites. XRD and FFT showed that the main component of urinary nanocrystallites was UA, which contains a small amount of calcium oxalate monohydrate and phosphates. EDS showed the characteristic absorption peaks of C, O, Ca and P. The formation of UA stones was closely related to a large number of UA nanocrystallites in urine. A combination of HRTEM, FFT, EDS and XRD analyses could be performed accurately to analyse the components of urinary nanocrystallites.

Variability of Protein Structure Models from Electron Microscopy.

PubMed

Monroe, Lyman; Terashi, Genki; Kihara, Daisuke

2017-04-04

An increasing number of biomolecular structures are solved by electron microscopy (EM). However, the quality of structure models determined from EM maps vary substantially. To understand to what extent structure models are supported by information embedded in EM maps, we used two computational structure refinement methods to examine how much structures can be refined using a dataset of 49 maps with accompanying structure models. The extent of structure modification as well as the disagreement between refinement models produced by the two computational methods scaled inversely with the global and the local map resolutions. A general quantitative estimation of deviations of structures for particular map resolutions are provided. Our results indicate that the observed discrepancy between the deposited map and the refined models is due to the lack of structural information present in EM maps and thus these annotations must be used with caution for further applications. Copyright © 2017 Elsevier Ltd. All rights reserved.

Effect of Ce doping on structural, optical and photocatalytic properties of ZnO nano-structures.

PubMed

Selvam, N Clament Sagaya; Vijaya, J Judith; Kennedy, L John

2014-03-01

A novel self-assembled pure and Ce doped ZnO nano-particles (NPs) were successfully synthesized by a simple low temperature co-precipitation method. The prepared photocatalysts were characterized by X-ray diffraction (XRD), High resolution scanning electron microscopy (HR-SEM), High resolution transmission electron microscopy (HR-TEM), diffuse reflectance spectroscopy (DRS) and Photoluminescence (PL) spectroscopy. The results indicated that the prepared photocatalysts shows a novel morphology, high crystallinity, uniform size distribution, and more defects. Photocatalytic degradation (PCD) of nonylphenol, a potent endocrine disrupting chemical in aqueous medium was investigated. Higher amount of oxygen defects exhibits enhanced PCD of nonylphenol. In addition, the influence of the Ce contents on the structure, morphology, absorption, emission and photocatalytic activity of ZnO nanoparticles (NPs) were investigated systematically. The relative PCD efficiency of pure ZnO, Ce-doped ZnO NPs and commercial TiO2 (Degussa P-25) have also been discussed.

Complementary Imaging of Silver Nanoparticle Interactions with Green Algae: Dark-Field Microscopy, Electron Microscopy, and Nanoscale Secondary Ion Mass Spectrometry.

PubMed

Sekine, Ryo; Moore, Katie L; Matzke, Marianne; Vallotton, Pascal; Jiang, Haibo; Hughes, Gareth M; Kirby, Jason K; Donner, Erica; Grovenor, Chris R M; Svendsen, Claus; Lombi, Enzo

2017-11-28

Increasing consumer use of engineered nanomaterials has led to significantly increased efforts to understand their potential impact on the environment and living organisms. Currently, no individual technique can provide all the necessary information such as their size, distribution, and chemistry in complex biological systems. Consequently, there is a need to develop complementary instrumental imaging approaches that provide enhanced understanding of these "bio-nano" interactions to overcome the limitations of individual techniques. Here we used a multimodal imaging approach incorporating dark-field light microscopy, high-resolution electron microscopy, and nanoscale secondary ion mass spectrometry (NanoSIMS). The aim was to gain insight into the bio-nano interactions of surface-functionalized silver nanoparticles (Ag-NPs) with the green algae Raphidocelis subcapitata, by combining the fidelity, spatial resolution, and elemental identification offered by the three techniques, respectively. Each technique revealed that Ag-NPs interact with the green algae with a dependence on the size (10 nm vs 60 nm) and surface functionality (tannic acid vs branched polyethylenimine, bPEI) of the NPs. Dark-field light microscopy revealed the presence of strong light scatterers on the algal cell surface, and SEM imaging confirmed their nanoparticulate nature and localization at nanoscale resolution. NanoSIMS imaging confirmed their chemical identity as Ag, with the majority of signal concentrated at the cell surface. Furthermore, SEM and NanoSIMS provided evidence of 10 nm bPEI Ag-NP internalization at higher concentrations (40 μg/L), correlating with the highest toxicity observed from these NPs. This multimodal approach thus demonstrated an effective approach to complement dose-response studies in nano-(eco)-toxicological investigations.

Crystallography and Morphology of Niobium Carbide in As-Cast HP-Niobium Reformer Tubes

NASA Astrophysics Data System (ADS)

Buchanan, Karl G.; Kral, Milo V.

2012-06-01

The microstructures of two as-cast heats of niobium-modified HP stainless steels were characterized. Particular attention was paid to the interdendritic niobium-rich carbides formed during solidification of these alloys. At low magnifications, these precipitates are grouped in colonies of similar lamellae. Higher magnifications revealed that the lamellae actually obtain two distinct morphologies. The type I morphology exhibits broad planar interfaces with a smooth platelike shape. Type II lamellae have undulating interfaces and an overall reticulated shape. To provide further insight into the origin of these two different morphologies, the microstructure and crystallography of each have been studied in detail using high resolution scanning electron microscopy, transmission electron microscopy, various electron diffraction methods (electron backscatter diffraction (EBSD), selected area diffraction (SAD), and convergent beam electron diffraction (CBED)), and energy dispersive X-ray spectroscopy.

Fundamental edge broadening effects during focused electron beam induced nanosynthesis

DOE PAGES

Schmied, Roland; Fowlkes, Jason Davidson; Winkler, Robert; ...

2015-02-16

In this study, we explore lateral broadening effects of 3D structures fabricated through focused electron beam induced deposition using MeCpPt(IV)Me 3 precursor. In particular, the scaling behavior of proximity effects as a function of the primary electron energy and the deposit height is investigated through experiments and validated through simulations. Correlated Kelvin force microscopy and conductive atomic force microscopy measurements identified conductive and non-conductive proximity regions. It was determined that the highest primary electron energies enable the highest edge sharpness while lower energies contain a complex convolution of broadening effects. In addition, it is demonstrated that intermediate energies lead tomore » even more complex proximity effects that significantly reduce lateral edge sharpness and thus should be avoided if desiring high lateral resolution.« less

Ultrafast transmission electron microscopy using a laser-driven field emitter: Femtosecond resolution with a high coherence electron beam.

PubMed

Feist, Armin; Bach, Nora; Rubiano da Silva, Nara; Danz, Thomas; Möller, Marcel; Priebe, Katharina E; Domröse, Till; Gatzmann, J Gregor; Rost, Stefan; Schauss, Jakob; Strauch, Stefanie; Bormann, Reiner; Sivis, Murat; Schäfer, Sascha; Ropers, Claus

2017-05-01

We present the development of the first ultrafast transmission electron microscope (UTEM) driven by localized photoemission from a field emitter cathode. We describe the implementation of the instrument, the photoemitter concept and the quantitative electron beam parameters achieved. Establishing a new source for ultrafast TEM, the Göttingen UTEM employs nano-localized linear photoemission from a Schottky emitter, which enables operation with freely tunable temporal structure, from continuous wave to femtosecond pulsed mode. Using this emission mechanism, we achieve record pulse properties in ultrafast electron microscopy of 9Å focused beam diameter, 200fs pulse duration and 0.6eV energy width. We illustrate the possibility to conduct ultrafast imaging, diffraction, holography and spectroscopy with this instrument and also discuss opportunities to harness quantum coherent interactions between intense laser fields and free-electron beams. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Development of a Fiber Laser with Independently Adjustable Properties for Optical Resolution Photoacoustic Microscopy.

PubMed

Aytac-Kipergil, Esra; Demirkiran, Aytac; Uluc, Nasire; Yavas, Seydi; Kayikcioglu, Tunc; Salman, Sarper; Karamuk, Sohret Gorkem; Ilday, Fatih Omer; Unlu, Mehmet Burcin

2016-12-08

Photoacoustic imaging is based on the detection of generated acoustic waves through thermal expansion of tissue illuminated by short laser pulses. Fiber lasers as an excitation source for photoacoustic imaging have recently been preferred for their high repetition frequencies. Here, we report a unique fiber laser developed specifically for multiwavelength photoacoustic microscopy system. The laser is custom-made for maximum flexibility in adjustment of its parameters; pulse duration (5-10 ns), pulse energy (up to 10 μJ) and repetition frequency (up to 1 MHz) independently from each other and covers a broad spectral region from 450 to 1100 nm and also can emit wavelengths of 532, 355, and 266 nm. The laser system consists of a master oscillator power amplifier, seeding two stages; supercontinuum and harmonic generation units. The laser is outstanding since the oscillator, amplifier and supercontinuum generation parts are all-fiber integrated with custom-developed electronics and software. To demonstrate the feasibility of the system, the images of several elements of standardized resolution test chart are acquired at multiple wavelengths. The lateral resolution of optical resolution photoacoustic microscopy system is determined as 2.68 μm. The developed system may pave the way for spectroscopic photoacoustic microscopy applications via widely tunable fiber laser technologies.

Development of a Fiber Laser with Independently Adjustable Properties for Optical Resolution Photoacoustic Microscopy

PubMed Central

Aytac-Kipergil, Esra; Demirkiran, Aytac; Uluc, Nasire; Yavas, Seydi; Kayikcioglu, Tunc; Salman, Sarper; Karamuk, Sohret Gorkem; Ilday, Fatih Omer; Unlu, Mehmet Burcin

2016-01-01

Photoacoustic imaging is based on the detection of generated acoustic waves through thermal expansion of tissue illuminated by short laser pulses. Fiber lasers as an excitation source for photoacoustic imaging have recently been preferred for their high repetition frequencies. Here, we report a unique fiber laser developed specifically for multiwavelength photoacoustic microscopy system. The laser is custom-made for maximum flexibility in adjustment of its parameters; pulse duration (5–10 ns), pulse energy (up to 10 μJ) and repetition frequency (up to 1 MHz) independently from each other and covers a broad spectral region from 450 to 1100 nm and also can emit wavelengths of 532, 355, and 266 nm. The laser system consists of a master oscillator power amplifier, seeding two stages; supercontinuum and harmonic generation units. The laser is outstanding since the oscillator, amplifier and supercontinuum generation parts are all-fiber integrated with custom-developed electronics and software. To demonstrate the feasibility of the system, the images of several elements of standardized resolution test chart are acquired at multiple wavelengths. The lateral resolution of optical resolution photoacoustic microscopy system is determined as 2.68 μm. The developed system may pave the way for spectroscopic photoacoustic microscopy applications via widely tunable fiber laser technologies. PMID:27929049

High throughput, detailed, cell-specific neuroanatomy of dendritic spines using microinjection and confocal microscopy

PubMed Central

Dumitriu, Dani; Rodriguez, Alfredo; Morrison, John H.

2012-01-01

Morphological features such as size, shape and density of dendritic spines have been shown to reflect important synaptic functional attributes and potential for plasticity. Here we describe in detail a protocol for obtaining detailed morphometric analysis of spines using microinjection of fluorescent dyes, high resolution confocal microscopy, deconvolution and image analysis using NeuronStudio. Recent technical advancements include better preservation of tissue resulting in prolonged ability to microinject, and algorithmic improvements that compensate for the residual Z-smear inherent in all optical imaging. Confocal imaging parameters were probed systematically for the identification of both optimal resolution as well as highest efficiency. When combined, our methods yield size and density measurements comparable to serial section transmission electron microscopy in a fraction of the time. An experiment containing 3 experimental groups with 8 subjects in each can take as little as one month if optimized for speed, or approximately 4 to 5 months if the highest resolution and morphometric detail is sought. PMID:21886104

The sea urchin egg jelly coat is a three-dimensional fibrous network as seen by intermediate voltage electron microscopy and deep etching analysis.

PubMed

Bonnell, B S; Larabell, C; Chandler, D E

1993-06-01

The egg jelly (EJ) coat which surrounds the unfertilized sea urchin egg undergoes extensive swelling upon contact with sea water, forming a three-dimensional network of interconnected fibers extending nearly 50 microns from the egg surface. Owing to its solubility, this coat has been difficult to visualize by light and electron microscopy. However, Lytechinus pictus EJ coats remain intact, if the fixation medium is maintained at pH 9. The addition of alcian blue during the final dehydration step of sample preparation stains the EJ for visualization of resin embedded eggs by both light and electron microscopy. Stereo pairs taken of thick sections prepared for intermediate voltage electron microscopy (IVEM) produce a three-dimensional image of the EJ network, consisting of interconnected fibers decorated along their length by globular jelly components. Using scanning electron microscopy (SEM), we have shown that before swelling, EJ exists in a tightly bound network of jelly fibers, 50-60 nm in diameter. In contrast, swollen EJ consists of a greatly extended network whose fibrous components measure 10 to 30 nm in diameter. High resolution stereo images of hydrated jelly produced by the quick-freeze/deep-etch/rotary-shadowing technique (QF/DE/RS) show nearly identical EJ networks, suggesting that dehydration does not markedly alter the structure of this extracellular matrix.

Modular Detection of GFP-Labeled Proteins for Rapid Screening by Electron Microscopy in Cells and Organisms.

PubMed

Ariotti, Nicholas; Hall, Thomas E; Rae, James; Ferguson, Charles; McMahon, Kerrie-Ann; Martel, Nick; Webb, Robyn E; Webb, Richard I; Teasdale, Rohan D; Parton, Robert G

2015-11-23

Reliable and quantifiable high-resolution protein localization is critical for understanding protein function. However, the time required to clone and characterize any protein of interest is a significant bottleneck, especially for electron microscopy (EM). We present a modular system for enzyme-based protein tagging that allows for improved speed and sampling for analysis of subcellular protein distributions using existing clone libraries to EM-resolution. We demonstrate that we can target a modified soybean ascorbate peroxidase (APEX) to any GFP-tagged protein of interest by engineering a GFP-binding peptide (GBP) directly to the APEX-tag. We demonstrate that APEX-GBP (1) significantly reduces the time required to characterize subcellular protein distributions of whole libraries to less than 3 days, (2) provides remarkable high-resolution localization of proteins to organelle subdomains, and (3) allows EM localization of GFP-tagged proteins, including proteins expressed at endogenous levels, in vivo by crossing existing GFP-tagged transgenic zebrafish lines with APEX-GBP transgenic lines. Copyright © 2015 Elsevier Inc. All rights reserved.

Wavefront propagation simulations for a UV/soft x-ray beamline: Electron Spectro-Microscopy beamline at NSLS-II

NASA Astrophysics Data System (ADS)

Canestrari, N.; Bisogni, V.; Walter, A.; Zhu, Y.; Dvorak, J.; Vescovo, E.; Chubar, O.

2014-09-01

A "source-to-sample" wavefront propagation analysis of the Electron Spectro-Microscopy (ESM) UV / soft X-ray beamline, which is under construction at the National Synchrotron Light Source II (NSLS-II) in the Brookhaven National Laboratory, has been conducted. All elements of the beamline - insertion device, mirrors, variable-line-spacing gratings and slits - are included in the simulations. Radiation intensity distributions at the sample position are displayed for representative photon energies in the UV range (20 - 100 eV) where diffraction effects are strong. The finite acceptance of the refocusing mirrors is the dominating factor limiting the spatial resolution at the sample (by ~3 μm at 20 eV). Absolute estimates of the radiation flux and energy resolution at the sample are also obtained from the electromagnetic calculations. The analysis of the propagated UV range undulator radiation at different deflection parameter values demonstrates that within the beamline angular acceptance a slightly "red-shifted" radiation provides higher flux at the sample and better energy resolution compared to the on-axis resonant radiation of the fundamental harmonic.

Defects in oxide surfaces studied by atomic force and scanning tunneling microscopy

PubMed Central

König, Thomas; Simon, Georg H; Heinke, Lars; Lichtenstein, Leonid

2011-01-01

Summary Surfaces of thin oxide films were investigated by means of a dual mode NC-AFM/STM. Apart from imaging the surface termination by NC-AFM with atomic resolution, point defects in magnesium oxide on Ag(001) and line defects in aluminum oxide on NiAl(110), respectively, were thoroughly studied. The contact potential was determined by Kelvin probe force microscopy (KPFM) and the electronic structure by scanning tunneling spectroscopy (STS). On magnesium oxide, different color centers, i.e., F0, F+, F2+ and divacancies, have different effects on the contact potential. These differences enabled classification and unambiguous differentiation by KPFM. True atomic resolution shows the topography at line defects in aluminum oxide. At these domain boundaries, STS and KPFM verify F2+-like centers, which have been predicted by density functional theory calculations. Thus, by determining the contact potential and the electronic structure with a spatial resolution in the nanometer range, NC-AFM and STM can be successfully applied on thin oxide films beyond imaging the topography of the surface atoms. PMID:21977410

High-resolution x-ray diffraction and transmission electron microscopy of multiferroic BiFeO3 films

NASA Astrophysics Data System (ADS)

Qi, Xiaoding; Wei, Ming; Lin, Yuan; Jia, Quanxi; Zhi, Dan; Dho, Joonghoe; Blamire, Mark G.; MacManus-Driscoll, Judith L.

2005-02-01

High-resolution x-ray diffraction and transmission electron microscopy (TEM) have been used to study BiFeO3 thin films grown on the bare and SrRuO3 buffered (001) SrTiO3 substrates. Reciprocal space mapping (RSM) around (002) and (103) reflections revealed that BFO films with a thickness of about 200 nm were almost fully relaxed and had a rhombohedral structure. Cross-sectional, high-resolution TEM showed that the films started to relax at a very early stage of growth, which was consistent with the RSM results. A thin intermediate layer of about 2 nm was observed at the interface, which had a smaller lattice than the overgrown film. Twist distortions about the c axis to release the shear strain introduced by the growth of rhombic (001) BiFeO3 on cubic (001) SrTiO3 were also observed. The results indicate that a strained, coherent BiFeO3 film on (001) SrTiO3 is very difficult to maintain and (111) STO substrates are preferable.

Gold nanoparticle uptake in whole cells in liquid examined by environmental scanning electron microscopy.

PubMed

Peckys, Diana B; de Jonge, Niels

2014-02-01

The size of gold nanoparticles (AuNPs) can influence various aspects of their cellular uptake. Light microscopy is not capable of resolving most AuNPs, while electron microscopy (EM) is not practically capable of acquiring the necessary statistical data from many cells and the results may suffer from various artifacts. Here, we demonstrate the use of a fast EM method for obtaining high-resolution data from a much larger population of cells than is usually feasible with conventional EM. A549 (human lung carcinoma) cells were subjected to uptake protocols with 10, 15, or 30 nm diameter AuNPs with adsorbed serum proteins. After 20 min, 24 h, or 45 h, the cells were fixed and imaged in whole in a thin layer of liquid water with environmental scanning electron microscopy equipped with a scanning transmission electron microscopy detector. The fast preparation and imaging of 145 whole cells in liquid allowed collection of nanoscale data within an exceptionally small amount of time of ~80 h. Analysis of 1,041 AuNP-filled vesicles showed that the long-term AuNP storing lysosomes increased their average size by 80 nm when AuNPs with 30 nm diameter were uptaken, compared to lysosomes of cells incubated with AuNPs of 10 and 15 nm diameter.

Quantitative Imaging of Single Unstained Magnetotactic Bacteria by Coherent X-ray Diffraction Microscopy.

PubMed

Fan, Jiadong; Sun, Zhibin; Zhang, Jian; Huang, Qingjie; Yao, Shengkun; Zong, Yunbing; Kohmura, Yoshiki; Ishikawa, Tetsuya; Liu, Hong; Jiang, Huaidong

2015-06-16

Novel coherent diffraction microscopy provides a powerful lensless imaging method to obtain a better understanding of the microorganism at the nanoscale. Here we demonstrated quantitative imaging of intact unstained magnetotactic bacteria using coherent X-ray diffraction microscopy combined with an iterative phase retrieval algorithm. Although the signal-to-noise ratio of the X-ray diffraction pattern from single magnetotactic bacterium is weak due to low-scattering ability of biomaterials, an 18.6 nm half-period resolution of reconstructed image was achieved by using a hybrid input-output phase retrieval algorithm. On the basis of the quantitative reconstructed images, the morphology and some intracellular structures, such as nucleoid, polyβ-hydroxybutyrate granules, and magnetosomes, were identified, which were also confirmed by scanning electron microscopy and energy dispersive spectroscopy. With the benefit from the quantifiability of coherent diffraction imaging, for the first time to our knowledge, an average density of magnetotactic bacteria was calculated to be ∼1.19 g/cm(3). This technique has a wide range of applications, especially in quantitative imaging of low-scattering biomaterials and multicomponent materials at nanoscale resolution. Combined with the cryogenic technique or X-ray free electron lasers, the method could image cells in a hydrated condition, which helps to maintain their natural structure.

Transmission electron microscope studies of extraterrestrial materials

NASA Technical Reports Server (NTRS)

Keller, Lindsay P.

1995-01-01

Transmission Electron Microscopy, X-Ray spectrometry and electron-energy-loss spectroscopy are used to analyse carbon in interplanetary dust particles. Optical micrographs are shown depicting cross sections of the dust particles embedded in sulphur. Selected-area electron diffraction patterns are shown. Transmission Electron Microscope specimens of lunar soil were prepared using two methods: ion-milling and ultramicrotomy. A combination of high resolution TEM imaging and electron diffraction is used to characterize the opaque assemblages. The opaque assemblages analyzed in this study are dominated by ilmenite with lesser rutile and spinel exsolutions, and traces of Fe metal.

Atomic force microscopic imaging of Acanthamoeba castellanii and Balamuthia mandrillaris trophozoites and cysts.

PubMed

Aqeel, Yousuf; Siddiqui, Ruqaiyyah; Ateeq, Muhammad; Raza Shah, Muhammad; Kulsoom, Huma; Khan, Naveed Ahmed

2015-01-01

Light microscopy and electron microscopy have been successfully used in the study of microbes, as well as free-living protists. Unlike light microscopy, which enables us to observe living organisms or the electron microscope which provides a two-dimensional image, atomic force microscopy provides a three-dimensional surface profile. Here, we observed two free-living amoebae, Acanthamoeba castellanii and Balamuthia mandrillaris under the phase contrast inverted microscope, transmission electron microscope and atomic force microscope. Although light microscopy was of lower magnification, it revealed functional biology of live amoebae such as motility and osmoregulation using contractile vacuoles of the trophozoite stage, but it is of limited value in defining the cyst stage. In contrast, transmission electron microscopy showed significantly greater magnification and resolution to reveal the ultra-structural features of trophozoites and cysts including intracellular organelles and cyst wall characteristics but it only produced a snapshot in time of a dead amoeba cell. Atomic force microscopy produced three-dimensional images providing detailed topographic description of shape and surface, phase imaging measuring boundary stiffness, and amplitude measurements including width, height and length of A. castellanii and B. mandrillaris trophozoites and cysts. These results demonstrate the importance of the application of various microscopic methods in the biological and structural characterization of the whole cell, ultra-structural features, as well as surface components and cytoskeleton of protist pathogens. © 2014 The Author(s) Journal of Eukaryotic Microbiology © 2014 International Society of Protistologists.

Direct Detection Electron Energy-Loss Spectroscopy: A Method to Push the Limits of Resolution and Sensitivity.

PubMed

Hart, James L; Lang, Andrew C; Leff, Asher C; Longo, Paolo; Trevor, Colin; Twesten, Ray D; Taheri, Mitra L

2017-08-15

In many cases, electron counting with direct detection sensors offers improved resolution, lower noise, and higher pixel density compared to conventional, indirect detection sensors for electron microscopy applications. Direct detection technology has previously been utilized, with great success, for imaging and diffraction, but potential advantages for spectroscopy remain unexplored. Here we compare the performance of a direct detection sensor operated in counting mode and an indirect detection sensor (scintillator/fiber-optic/CCD) for electron energy-loss spectroscopy. Clear improvements in measured detective quantum efficiency and combined energy resolution/energy field-of-view are offered by counting mode direct detection, showing promise for efficient spectrum imaging, low-dose mapping of beam-sensitive specimens, trace element analysis, and time-resolved spectroscopy. Despite the limited counting rate imposed by the readout electronics, we show that both core-loss and low-loss spectral acquisition are practical. These developments will benefit biologists, chemists, physicists, and materials scientists alike.

Materials characterisation by angle-resolved scanning transmission electron microscopy.

PubMed

Müller-Caspary, Knut; Oppermann, Oliver; Grieb, Tim; Krause, Florian F; Rosenauer, Andreas; Schowalter, Marco; Mehrtens, Thorsten; Beyer, Andreas; Volz, Kerstin; Potapov, Pavel

2016-11-16

Solid-state properties such as strain or chemical composition often leave characteristic fingerprints in the angular dependence of electron scattering. Scanning transmission electron microscopy (STEM) is dedicated to probe scattered intensity with atomic resolution, but it drastically lacks angular resolution. Here we report both a setup to exploit the explicit angular dependence of scattered intensity and applications of angle-resolved STEM to semiconductor nanostructures. Our method is applied to measure nitrogen content and specimen thickness in a GaN x As 1-x layer independently at atomic resolution by evaluating two dedicated angular intervals. We demonstrate contrast formation due to strain and composition in a Si- based metal-oxide semiconductor field effect transistor (MOSFET) with Ge x Si 1-x stressors as a function of the angles used for imaging. To shed light on the validity of current theoretical approaches this data is compared with theory, namely the Rutherford approach and contemporary multislice simulations. Inconsistency is found for the Rutherford model in the whole angular range of 16-255 mrad. Contrary, the multislice simulations are applicable for angles larger than 35 mrad whereas a significant mismatch is observed at lower angles. This limitation of established simulations is discussed particularly on the basis of inelastic scattering.

Facing the phase problem in Coherent Diffractive Imaging via Memetic Algorithms.

PubMed

Colombo, Alessandro; Galli, Davide Emilio; De Caro, Liberato; Scattarella, Francesco; Carlino, Elvio

2017-02-09

Coherent Diffractive Imaging is a lensless technique that allows imaging of matter at a spatial resolution not limited by lens aberrations. This technique exploits the measured diffraction pattern of a coherent beam scattered by periodic and non-periodic objects to retrieve spatial information. The diffracted intensity, for weak-scattering objects, is proportional to the modulus of the Fourier Transform of the object scattering function. Any phase information, needed to retrieve its scattering function, has to be retrieved by means of suitable algorithms. Here we present a new approach, based on a memetic algorithm, i.e. a hybrid genetic algorithm, to face the phase problem, which exploits the synergy of deterministic and stochastic optimization methods. The new approach has been tested on simulated data and applied to the phasing of transmission electron microscopy coherent electron diffraction data of a SrTiO 3 sample. We have been able to quantitatively retrieve the projected atomic potential, and also image the oxygen columns, which are not directly visible in the relevant high-resolution transmission electron microscopy images. Our approach proves to be a new powerful tool for the study of matter at atomic resolution and opens new perspectives in those applications in which effective phase retrieval is necessary.

Microscopy based studies on the interaction of bio-based silver nanoparticles with Bombyx mori Nuclear Polyhedrosis virus.

PubMed

Tamilselvan, Selvaraj; Ashokkumar, Thirunavukkarasu; Govindaraju, Kasivelu

2017-04-01

In the present investigation, silver nanoparticles (AgNPs) interactions with Bombyx mori Nuclear Polyhedrosis virus (BmNPV) were characterized using High-Resolution Scanning Electron Microscopy (HR-SEM), Energy Dispersive X-ray Analysis (EDAX), Transmission Electron Microscopy (TEM), Atomic Force Microcopy (AFM) and Confocal Microscope (CM). HR-SEM study reveals that the biosynthesized AgNPs have interacted with BmNPV and were found on the surface. TEM micrographs of normal and viral polyhedra treated with AgNPs showed that the nanoparticles were accumulated in the membrane and it was noted that some of the AgNPs successfully penetrated the membrane by reaching the capsid of BmNPV. AFM and confocal microscopy studies reveal that the disruption in the shell membrane tends to lose its stability due to exposure of AgNPs to BmNPV. Copyright © 2017 Elsevier B.V. All rights reserved.

Study of structural colour of Hebomoia glaucippe butterfly wing scales

NASA Astrophysics Data System (ADS)

Shur, V. Ya; Kuznetsov, D. K.; Pryakhina, V. I.; Kosobokov, M. S.; Zubarev, I. V.; Boymuradova, S. K.; Volchetskaya, K. V.

2017-10-01

Structural colours of Hebomoia glaucippe butterfly wing scales have been studied experimentally using high resolution scanning electron microscopy. Visualization of scales structures and computer simulation allowed distinguishing correlation between nanostructures on the scales and their colour.

Carbon Nanotubes as an Ultrafast Emitter with a Narrow Energy Spread at Optical Frequency.

PubMed

Li, Chi; Zhou, Xu; Zhai, Feng; Li, Zhenjun; Yao, Fengrui; Qiao, Ruixi; Chen, Ke; Cole, Matthew Thomas; Yu, Dapeng; Sun, Zhipei; Liu, Kaihui; Dai, Qing

2017-08-01

Ultrafast electron pulses, combined with laser-pump and electron-probe technologies, allow ultrafast dynamics to be characterized in materials. However, the pursuit of simultaneous ultimate spatial and temporal resolution of microscopy and spectroscopy is largely subdued by the low monochromaticity of the electron pulses and their poor phase synchronization to the optical excitation pulses. Field-driven photoemission from metal tips provides high light-phase synchronization, but suffers large electron energy spreads (3-100 eV) as driven by a long wavelength laser (>800 nm). Here, ultrafast electron emission from carbon nanotubes (≈1 nm radius) excited by a 410 nm femtosecond laser is realized in the field-driven regime. In addition, the emitted electrons have great monochromaticity with energy spread as low as 0.25 eV. This great performance benefits from the extraordinarily high field enhancement and great stability of carbon nanotubes, superior to metal tips. The new nanotube-based ultrafast electron source opens exciting prospects for extending current characterization to sub-femtosecond temporal resolution as well as sub-nanometer spatial resolution. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

High Resolution Orientation Imaging Microscopy

DTIC Science & Technology

2012-05-02

Structure of In-Situ Deformations of Steel , TMS, San Diego, 2011 13. Jay Basinger, David Fullwood, Brent Adams, EBSD Detail Extraction for Greater Spatial...Its use has contributed to the development of new steels , aluminum alloys, high TC superconductors, electronic materials, lead-free solders, optical...Resolution The simulated pattern method has been used to recover lattice tetragonality in high-strength low- alloy steels . Since the level of

Design principles and applications of a cooled CCD camera for electron microscopy.

PubMed

Faruqi, A R

1998-01-01

Cooled CCD cameras offer a number of advantages in recording electron microscope images with CCDs rather than film which include: immediate availability of the image in a digital format suitable for further computer processing, high dynamic range, excellent linearity and a high detective quantum efficiency for recording electrons. In one important respect however, film has superior properties: the spatial resolution of CCD detectors tested so far (in terms of point spread function or modulation transfer function) are inferior to film and a great deal of our effort has been spent in designing detectors with improved spatial resolution. Various instrumental contributions to spatial resolution have been analysed and in this paper we discuss the contribution of the phosphor-fibre optics system in this measurement. We have evaluated the performance of a number of detector components and parameters, e.g. different phosphors (and a scintillator), optical coupling with lens or fibre optics with various demagnification factors, to improve the detector performance. The camera described in this paper, which is based on this analysis, uses a tapered fibre optics coupling between the phosphor and the CCD and is installed on a Philips CM12 electron microscope equipped to perform cryo-microscopy. The main use of the camera so far has been in recording electron diffraction patterns from two dimensional crystals of bacteriorhodopsin--from wild type and from different trapped states during the photocycle. As one example of the type of data obtained with the CCD camera a two dimensional Fourier projection map from the trapped O-state is also included. With faster computers, it will soon be possible to undertake this type of work on an on-line basis. Also, with improvements in detector size and resolution, CCD detectors, already ideal for diffraction, will be able to compete with film in the recording of high resolution images.

Selected-zone dark-field electron microscopy.

NASA Technical Reports Server (NTRS)

Heinemann, K.; Poppa, H.

1972-01-01

Description of a new method which makes it possible to reduce drastically the resolution-limiting influence of chromatic aberration, and thus to obtain high-quality images, by selecting the image-forming electrons that have passed through a small annular zone of an objective lens. In addition, the manufacture of special objective-lens aperture diaphragms that are needed for this method is also described.

Synthesis, Structure And Properties of Electrochemically Active Nanocomposites

DTIC Science & Technology

2003-05-01

milling. Detailed systematic impedance analysis , electronic conductivity measurement and high-resolution electron microscopy studies have shown that...carbon particles determined by TEM analysis . Results of the studies so far have shown that Sn and Si-based nanocomposites appear to be quite promising... Analysis of the As-milled Powders 117 2. Electrochemical Characteristics of Si/SiC Nanocomposites 120 3. Microstructural/Morphological Analysis of

Electron beam detection of a Nanotube Scanning Force Microscope.

PubMed

Siria, Alessandro; Niguès, Antoine

2017-09-14

Atomic Force Microscopy (AFM) allows to probe matter at atomic scale by measuring the perturbation of a nanomechanical oscillator induced by near-field interaction forces. The quest to improve sensitivity and resolution of AFM forced the introduction of a new class of resonators with dimensions at the nanometer scale. In this context, nanotubes are the ultimate mechanical oscillators because of their one dimensional nature, small mass and almost perfect crystallinity. Coupled to the possibility of functionalisation, these properties make them the perfect candidates as ultra sensitive, on-demand force sensors. However their dimensions make the measurement of the mechanical properties a challenging task in particular when working in cavity free geometry at ambient temperature. By using a focused electron beam, we show that the mechanical response of nanotubes can be quantitatively measured while approaching to a surface sample. By coupling electron beam detection of individual nanotubes with a custom AFM we image the surface topography of a sample by continuously measuring the mechanical properties of the nanoresonators. The combination of very small size and mass together with the high resolution of the electron beam detection method offers unprecedented opportunities for the development of a new class of nanotube-based scanning force microscopy.

High-resolution electron microscopy observation of a new crystalline approximant W' of Mg-Zn-Y icosahedral quasicrystal

PubMed

Luo; Hashimoto

2000-10-01

A new ordered structure W' with a lattice parameter (a = 2.05 nm) about three times as large as that of the fundamental face-centered cubic W phase (a = 0.6848 nm) has been found in the Mg-Zn-Y system by means of transmission electron microscopy. The W' and W phases have the cube-to-cube orientation relationship. Moreover, the strong electron diffraction spots of the W' phase showed pseudoicosahedral symmetry, implying that it is a crystalline approximant of the Mg-Zn-Y icosahedral quasicrystal. In the high-resolution electron microscopic images of the W' phase, Penrose tiles of pentagons and boats with an edge length of a(p) = 0.481 nm can be identified. A binary tile of crown subunit has also been deduced from such a tiling. Translation domains of the W' phase have also been observed and the translation vectors at the domain boundary are: a(p), tau x a(p) and (1 + tau) x a(p), respectively, where (1 + tau) x a(p) equals to the edge length a(r) of the big obtuse rhombus of the W' phase and tau = (1 + square root of 5)/2, is the golden ratio.

Nanoscale Imaging of Whole Cells Using a Liquid Enclosure and a Scanning Transmission Electron Microscope

PubMed Central

Peckys, Diana B.; Veith, Gabriel M.; Joy, David C.; de Jonge, Niels

2009-01-01

Nanoscale imaging techniques are needed to investigate cellular function at the level of individual proteins and to study the interaction of nanomaterials with biological systems. We imaged whole fixed cells in liquid state with a scanning transmission electron microscope (STEM) using a micrometer-sized liquid enclosure with electron transparent windows providing a wet specimen environment. Wet-STEM images were obtained of fixed E. coli bacteria labeled with gold nanoparticles attached to surface membrane proteins. Mammalian cells (COS7) were incubated with gold-tagged epidermal growth factor and fixed. STEM imaging of these cells resulted in a resolution of 3 nm for the gold nanoparticles. The wet-STEM method has several advantages over conventional imaging techniques. Most important is the capability to image whole fixed cells in a wet environment with nanometer resolution, which can be used, e.g., to map individual protein distributions in/on whole cells. The sample preparation is compatible with that used for fluorescent microscopy on fixed cells for experiments involving nanoparticles. Thirdly, the system is rather simple and involves only minimal new equipment in an electron microscopy (EM) laboratory. PMID:20020038

Nanouric acid or nanocalcium phosphate as central nidus to induce calcium oxalate stone formation: a high-resolution transmission electron microscopy study on urinary nanocrystallites

PubMed Central

Gao, Jie; Xue, Jun-Fa; Xu, Meng; Gui, Bao-Song; Wang, Feng-Xin; Ouyang, Jian-Ming

2014-01-01

Purpose This study aimed to accurately analyze the relationship between calcium oxalate (CaOx) stone formation and the components of urinary nanocrystallites. Method High-resolution transmission electron microscopy (HRTEM), selected area electron diffraction, fast Fourier transformation of HRTEM, and energy dispersive X-ray spectroscopy were performed to analyze the components of these nanocrystallites. Results The main components of CaOx stones are calcium oxalate monohydrate and a small amount of dehydrate, while those of urinary nanocrystallites are calcium oxalate monohydrate, uric acid, and calcium phosphate. The mechanism of formation of CaOx stones was discussed based on the components of urinary nanocrystallites. Conclusion The formation of CaOx stones is closely related both to the properties of urinary nanocrystallites and to the urinary components. The combination of HRTEM, fast Fourier transformation, selected area electron diffraction, and energy dispersive X-ray spectroscopy could be accurately performed to analyze the components of single urinary nanocrystallites. This result provides evidence for nanouric acid and/or nanocalcium phosphate crystallites as the central nidus to induce CaOx stone formation. PMID:25258530

Near-Atomic Resolution Structure of a Highly Neutralizing Fab Bound to Canine Parvovirus.

PubMed

Organtini, Lindsey J; Lee, Hyunwook; Iketani, Sho; Huang, Kai; Ashley, Robert E; Makhov, Alexander M; Conway, James F; Parrish, Colin R; Hafenstein, Susan

2016-11-01

Canine parvovirus (CPV) is a highly contagious pathogen that causes severe disease in dogs and wildlife. Previously, a panel of neutralizing monoclonal antibodies (MAb) raised against CPV was characterized. An antibody fragment (Fab) of MAb E was found to neutralize the virus at low molar ratios. Using recent advances in cryo-electron microscopy (cryo-EM), we determined the structure of CPV in complex with Fab E to 4.1 Å resolution, which allowed de novo building of the Fab structure. The footprint identified was significantly different from the footprint obtained previously from models fitted into lower-resolution maps. Using single-chain variable fragments, we tested antibody residues that control capsid binding. The near-atomic structure also revealed that Fab binding had caused capsid destabilization in regions containing key residues conferring receptor binding and tropism, which suggests a mechanism for efficient virus neutralization by antibody. Furthermore, a general technical approach to solving the structures of small molecules is demonstrated, as binding the Fab to the capsid allowed us to determine the 50-kDa Fab structure by cryo-EM. Using cryo-electron microscopy and new direct electron detector technology, we have solved the 4 Å resolution structure of a Fab molecule bound to a picornavirus capsid. The Fab induced conformational changes in regions of the virus capsid that control receptor binding. The antibody footprint is markedly different from the previous one identified by using a 12 Å structure. This work emphasizes the need for a high-resolution structure to guide mutational analysis and cautions against relying on older low-resolution structures even though they were interpreted with the best methodology available at the time. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Near-Atomic Resolution Structure of a Highly Neutralizing Fab Bound to Canine Parvovirus

PubMed Central

Organtini, Lindsey J.; Lee, Hyunwook; Iketani, Sho; Huang, Kai; Ashley, Robert E.; Makhov, Alexander M.; Conway, James F.

2016-01-01

ABSTRACT Canine parvovirus (CPV) is a highly contagious pathogen that causes severe disease in dogs and wildlife. Previously, a panel of neutralizing monoclonal antibodies (MAb) raised against CPV was characterized. An antibody fragment (Fab) of MAb E was found to neutralize the virus at low molar ratios. Using recent advances in cryo-electron microscopy (cryo-EM), we determined the structure of CPV in complex with Fab E to 4.1 Å resolution, which allowed de novo building of the Fab structure. The footprint identified was significantly different from the footprint obtained previously from models fitted into lower-resolution maps. Using single-chain variable fragments, we tested antibody residues that control capsid binding. The near-atomic structure also revealed that Fab binding had caused capsid destabilization in regions containing key residues conferring receptor binding and tropism, which suggests a mechanism for efficient virus neutralization by antibody. Furthermore, a general technical approach to solving the structures of small molecules is demonstrated, as binding the Fab to the capsid allowed us to determine the 50-kDa Fab structure by cryo-EM. IMPORTANCE Using cryo-electron microscopy and new direct electron detector technology, we have solved the 4 Å resolution structure of a Fab molecule bound to a picornavirus capsid. The Fab induced conformational changes in regions of the virus capsid that control receptor binding. The antibody footprint is markedly different from the previous one identified by using a 12 Å structure. This work emphasizes the need for a high-resolution structure to guide mutational analysis and cautions against relying on older low-resolution structures even though they were interpreted with the best methodology available at the time. PMID:27535057

Studying the Stoichiometry of Epidermal Growth Factor Receptor in Intact Cells using Correlative Microscopy.

PubMed

Peckys, Diana B; de Jonge, Niels

2015-09-11

This protocol describes the labeling of epidermal growth factor receptor (EGFR) on COS7 fibroblast cells, and subsequent correlative fluorescence microscopy and environmental scanning electron microscopy (ESEM) of whole cells in hydrated state. Fluorescent quantum dots (QDs) were coupled to EGFR via a two-step labeling protocol, providing an efficient and specific protein labeling, while avoiding label-induced clustering of the receptor. Fluorescence microscopy provided overview images of the cellular locations of the EGFR. The scanning transmission electron microscopy (STEM) detector was used to detect the QD labels with nanoscale resolution. The resulting correlative images provide data of the cellular EGFR distribution, and the stoichiometry at the single molecular level in the natural context of the hydrated intact cell. ESEM-STEM images revealed the receptor to be present as monomer, as homodimer, and in small clusters. Labeling with two different QDs, i.e., one emitting at 655 nm and at 800 revealed similar characteristic results.

High resolution IVEM tomography of biological specimens

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sedat, J.W.; Agard, D.A.

Electron tomography is a powerful tool for elucidating the three-dimensional architecture of large biological complexes and subcellular organelles. The introduction of intermediate voltage electron microscopes further extended the technique by providing the means to examine very large and non-symmetrical subcellular organelles, at resolutions beyond what would be possible using light microscopy. Recent studies using electron tomography on a variety of cellular organelles and assemblies such as centrosomes, kinetochores, and chromatin have clearly demonstrated the power of this technique for obtaining 3D structural information on non-symmetric cell components. When combined with biochemical and molecular observations, these 3D reconstructions have provided significantmore » new insights into biological function.« less

Optical magnetic imaging of living cells

PubMed Central

Le Sage, D.; Arai, K.; Glenn, D. R.; DeVience, S. J.; Pham, L. M.; Rahn-Lee, L.; Lukin, M. D.; Yacoby, A.; Komeili, A.; Walsworth, R. L.

2013-01-01

Magnetic imaging is a powerful tool for probing biological and physical systems. However, existing techniques either have poor spatial resolution compared to optical microscopy and are hence not generally applicable to imaging of sub-cellular structure (e.g., magnetic resonance imaging [MRI]1), or entail operating conditions that preclude application to living biological samples while providing sub-micron resolution (e.g., scanning superconducting quantum interference device [SQUID] microscopy2, electron holography3, and magnetic resonance force microscopy [MRFM]4). Here we demonstrate magnetic imaging of living cells (magnetotactic bacteria) under ambient laboratory conditions and with sub-cellular spatial resolution (400 nm), using an optically-detected magnetic field imaging array consisting of a nanoscale layer of nitrogen-vacancy (NV) colour centres implanted at the surface of a diamond chip. With the bacteria placed on the diamond surface, we optically probe the NV quantum spin states and rapidly reconstruct images of the vector components of the magnetic field created by chains of magnetic nanoparticles (magnetosomes) produced in the bacteria, and spatially correlate these magnetic field maps with optical images acquired in the same apparatus. Wide-field sCMOS acquisition allows parallel optical and magnetic imaging of multiple cells in a population with sub-micron resolution and >100 micron field-of-view. Scanning electron microscope (SEM) images of the bacteria confirm that the correlated optical and magnetic images can be used to locate and characterize the magnetosomes in each bacterium. The results provide a new capability for imaging bio-magnetic structures in living cells under ambient conditions with high spatial resolution, and will enable the mapping of a wide range of magnetic signals within cells and cellular networks5, 6. PMID:23619694

Correlation Functions Quantify Super-Resolution Images and Estimate Apparent Clustering Due to Over-Counting

PubMed Central

Veatch, Sarah L.; Machta, Benjamin B.; Shelby, Sarah A.; Chiang, Ethan N.; Holowka, David A.; Baird, Barbara A.

2012-01-01

We present an analytical method using correlation functions to quantify clustering in super-resolution fluorescence localization images and electron microscopy images of static surfaces in two dimensions. We use this method to quantify how over-counting of labeled molecules contributes to apparent self-clustering and to calculate the effective lateral resolution of an image. This treatment applies to distributions of proteins and lipids in cell membranes, where there is significant interest in using electron microscopy and super-resolution fluorescence localization techniques to probe membrane heterogeneity. When images are quantified using pair auto-correlation functions, the magnitude of apparent clustering arising from over-counting varies inversely with the surface density of labeled molecules and does not depend on the number of times an average molecule is counted. In contrast, we demonstrate that over-counting does not give rise to apparent co-clustering in double label experiments when pair cross-correlation functions are measured. We apply our analytical method to quantify the distribution of the IgE receptor (FcεRI) on the plasma membranes of chemically fixed RBL-2H3 mast cells from images acquired using stochastic optical reconstruction microscopy (STORM/dSTORM) and scanning electron microscopy (SEM). We find that apparent clustering of FcεRI-bound IgE is dominated by over-counting labels on individual complexes when IgE is directly conjugated to organic fluorophores. We verify this observation by measuring pair cross-correlation functions between two distinguishably labeled pools of IgE-FcεRI on the cell surface using both imaging methods. After correcting for over-counting, we observe weak but significant self-clustering of IgE-FcεRI in fluorescence localization measurements, and no residual self-clustering as detected with SEM. We also apply this method to quantify IgE-FcεRI redistribution after deliberate clustering by crosslinking with two distinct trivalent ligands of defined architectures, and we evaluate contributions from both over-counting of labels and redistribution of proteins. PMID:22384026

Interlaced zone plate optics for hard X-ray imaging in the 10 nm range

DOE PAGES

Mohacsi, Istvan; Vartiainen, Ismo; Rosner, Benedikt; ...

2017-03-08

Multi-keV X-ray microscopy has been particularly successful in bridging the resolution gap between optical and electron microscopy. However, resolutions below 20 nm are still considered challenging, as high throughput direct imaging methods are limited by the availability of suitable optical elements. In order to bridge this gap, we present a new type of Fresnel zone plate lenses aimed at the sub-20 and the sub-10 nm resolution range. By extending the concept of double-sided zone plate stacking, we demonstrate the doubling of the effective line density and thus the resolution and provide large aperture, single- chip optical devices with 15 andmore » 7 nm smallest zone widths. The detailed characterization of these lenses shows excellent optical properties with focal spots down to 7.8 nm. Furthermore, beyond wave front characterization, the zone plates also excel in typical imaging scenarios, verifying their resolution close to their diffraction limited optical performance.« less

Interlaced zone plate optics for hard X-ray imaging in the 10 nm range

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mohacsi, Istvan; Vartiainen, Ismo; Rosner, Benedikt

Multi-keV X-ray microscopy has been particularly successful in bridging the resolution gap between optical and electron microscopy. However, resolutions below 20 nm are still considered challenging, as high throughput direct imaging methods are limited by the availability of suitable optical elements. In order to bridge this gap, we present a new type of Fresnel zone plate lenses aimed at the sub-20 and the sub-10 nm resolution range. By extending the concept of double-sided zone plate stacking, we demonstrate the doubling of the effective line density and thus the resolution and provide large aperture, single- chip optical devices with 15 andmore » 7 nm smallest zone widths. The detailed characterization of these lenses shows excellent optical properties with focal spots down to 7.8 nm. Furthermore, beyond wave front characterization, the zone plates also excel in typical imaging scenarios, verifying their resolution close to their diffraction limited optical performance.« less

Interlaced zone plate optics for hard X-ray imaging in the 10 nm range

PubMed Central

Mohacsi, Istvan; Vartiainen, Ismo; Rösner, Benedikt; Guizar-Sicairos, Manuel; Guzenko, Vitaliy A.; McNulty, Ian; Winarski, Robert; Holt, Martin V.; David, Christian

2017-01-01

Multi-keV X-ray microscopy has been particularly successful in bridging the resolution gap between optical and electron microscopy. However, resolutions below 20 nm are still considered challenging, as high throughput direct imaging methods are limited by the availability of suitable optical elements. In order to bridge this gap, we present a new type of Fresnel zone plate lenses aimed at the sub-20 and the sub-10 nm resolution range. By extending the concept of double-sided zone plate stacking, we demonstrate the doubling of the effective line density and thus the resolution and provide large aperture, singlechip optical devices with 15 and 7 nm smallest zone widths. The detailed characterization of these lenses shows excellent optical properties with focal spots down to 7.8 nm. Beyond wave front characterization, the zone plates also excel in typical imaging scenarios, verifying their resolution close to their diffraction limited optical performance.

Interlaced zone plate optics for hard X-ray imaging in the 10 nm range

NASA Astrophysics Data System (ADS)

Mohacsi, Istvan; Vartiainen, Ismo; Rösner, Benedikt; Guizar-Sicairos, Manuel; Guzenko, Vitaliy A.; McNulty, Ian; Winarski, Robert; Holt, Martin V.; David, Christian

2017-03-01

Multi-keV X-ray microscopy has been particularly successful in bridging the resolution gap between optical and electron microscopy. However, resolutions below 20 nm are still considered challenging, as high throughput direct imaging methods are limited by the availability of suitable optical elements. In order to bridge this gap, we present a new type of Fresnel zone plate lenses aimed at the sub-20 and the sub-10 nm resolution range. By extending the concept of double-sided zone plate stacking, we demonstrate the doubling of the effective line density and thus the resolution and provide large aperture, singlechip optical devices with 15 and 7 nm smallest zone widths. The detailed characterization of these lenses shows excellent optical properties with focal spots down to 7.8 nm. Beyond wave front characterization, the zone plates also excel in typical imaging scenarios, verifying their resolution close to their diffraction limited optical performance.

Effective reduction of p-nitrophenol by silver nanoparticle loaded on magnetic Fe3O4/ATO nano-composite

NASA Astrophysics Data System (ADS)

Karki, Hem Prakash; Ojha, Devi Prashad; Joshi, Mahesh Kumar; Kim, Han Joo

2018-03-01

A silver loaded hematite (Fe3O4) and antimony doped tin oxide (ATO) magnetic nano-composite (Ag-Fe3O4/ATO) was successfully synthesized by in situ one pot green and facile hydrothermal process. The formation of nano-composite, its structure, morphology, and stability were characterized by field emission scanning electron microscopy (FE-SEM), high resolution transmission electron microscopy (HRTEM), electron diffraction spectroscopy (EDS), elemental mapping by high resolution scanning transmission electron microscopy (STEM), X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), Fourier transform infra-red spectroscopy (FTIR). UV-vis spectroscopy was used to monitor the catalytic reduction of p-nitrophenol (PNP) into p-aminophenol (PAP) in presence of Ag-Fe3O4/ATO nano-composite with excess of sodium borohydride (NaBH4). The pseudo-first order kinetic equation could describe the reduction of p-nitrophenol with excess of NaBH4. For the first time, ATO surface was used for hydrothermal growth of silver and iron oxide magnetic nanoparticles. The in situ growth of these nanoparticles provided an effective bonding of components of the nano-composite over the surface of ATO nanoparticles. This nano-composite exhibited easy synthesis, high stability, cost effective and rapid separation using external magnet. The excellent catalytic and anti-bacterial activity of as-synthesized silver nano-composite makes it potential nano-catalyst for waste water treatment as well as biomedical application.

Characterization of twin boundaries in an Fe–17.5Mn–0.56C twinning induced plasticity steel

DOE Office of Scientific and Technical Information (OSTI.GOV)

Patterson, Erin E., E-mail: erin.diedrich@yahoo.com; Field, David P., E-mail: dfield@wsu.edu; Zhang, Yudong, E-mail: yudong.zhang@univ-metz.fr

2013-11-15

A twinning-induced plasticity steel of composition Fe–17.5 wt.% Mn–0.56 wt.% C–1.39 wt.% Al–0.24 wt.% Si was analyzed for the purpose of characterizing the relationship between tensile strain and deformation twinning. Tensile samples achieved a maximum of 0.46 true strain at failure, and a maximum ultimate tensile strength of 1599 MPa. Electron backscatter diffraction (EBSD) analysis showed that the grain orientation rotated heavily to < 111 > parallel to the tensile axis above 0.3 true strain. Sigma 3 misorientations, as identified by EBSD orientation measurements, and using the image quality maps were used to quantify the number of twins present inmore » the scanned areas of the samples. The image quality method yielded a distinct positive correlation between the twin area density and deformation, but the orientation measurements were unreliable in quantifying twin density in these structures. Quantitative analysis of the twin fraction is limited from orientation information because of the poor spatial resolution of EBSD in relation to the twin thickness. The EBSD orientation maps created for a thin foil sample showed some improvement in the resolution of the twins, but not enough to be significant. Measurements of the twins in the transmission electron microscopy micrographs yielded an average thickness of 23 nm, which is near the resolution capabilities of EBSD on this material for the instrumentation used. Electron channeling contrast imaging performed on one bulk tensile specimen of 0.34 true strain, using a method of controlled diffraction, yielded several images of twinning, dislocation structures and strain fields. A twin thickness of 66 nm was measured by the same method used for the transmission electron microscopy measurement. It is apparent that the results obtain by electron channeling contrast imaging were better than those by EBSD but did not capture all information on the twin boundaries such as was observed by transmission electron microscopy. - Highlights: • Performed tensile tests to assess mechanical performance of TWIP alloy • Analyzed tensile specimens using EBSD, TEM, and ECCI • EBSD showed that most twinning occurred at or near the < 111 >//TA orientation. • EBSD, TEM and ECCI were used to measure average twin density. • Compared spatial resolution of EBSD, ECCI and TEM for the instrumentation used.« less

Detection of local chemical states of lithium and their spatial mapping by scanning transmission electron microscopy, electron energy-loss spectroscopy and hyperspectral image analysis.

PubMed

Muto, Shunsuke; Tatsumi, Kazuyoshi

2017-02-08

Advancements in the field of renewable energy resources have led to a growing demand for the analysis of light elements at the nanometer scale. Detection of lithium is one of the key issues to be resolved for providing guiding principles for the synthesis of cathode active materials, and degradation analysis after repeated use of those materials. We have reviewed the different techniques currently used for the characterization of light elements such as high-resolution transmission electron microscopy, scanning transmission electron microscopy (STEM) and electron energy-loss spectroscopy (EELS). In the present study, we have introduced a methodology to detect lithium in solid materials, particularly for cathode active materials used in lithium-ion battery. The chemical states of lithium were isolated and analyzed from the overlapping multiple spectral profiles, using a suite of STEM, EELS and hyperspectral image analysis. The method was successfully applied in the chemical state analyses of hetero-phases near the surface and grain boundary regions of the active material particles formed by chemical reactions between the electrolyte and the active materials. © The Author 2016. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

The quest for four-dimensional imaging in plant cell biology: it's just a matter of time

PubMed Central

Domozych, David S.

2012-01-01

Background Analysis of plant cell dynamics over time, or four-dimensional imaging (4-DI), represents a major goal of plant science. The ability to resolve structures in the third dimension within the cell or tissue during developmental events or in response to environmental or experimental stresses (i.e. 4-DI) is critical to our understanding of gene expression, post-expression modulations of macromolecules and sub-cellular system interactions. Scope Microscopy-based technologies have been profoundly integral to this type of investigation, and new and refined microscopy technologies now allow for the visualization of cell dynamics with unprecedented resolution, contrast and experimental versatility. However, certain realities of light and electron microscopy, choice of specimen and specimen preparation techniques limit the scope of readily attaining 4-DI. Today, the plant microscopist must use a combinatorial strategy whereby multiple microscopy-based investigations are used. Modern fluorescence, confocal laser scanning, transmission electron and scanning electron microscopy provide effective conduits for synthesizing data detailing live cell dynamics and highly resolved snapshots of specific cell structures that will ultimately lead to 4-DI. This review provides a synopsis of such technologies available. PMID:22628381

Whole-brain serial-section electron microscopy in larval zebrafish.

PubMed

Hildebrand, David Grant Colburn; Cicconet, Marcelo; Torres, Russel Miguel; Choi, Woohyuk; Quan, Tran Minh; Moon, Jungmin; Wetzel, Arthur Willis; Scott Champion, Andrew; Graham, Brett Jesse; Randlett, Owen; Plummer, George Scott; Portugues, Ruben; Bianco, Isaac Henry; Saalfeld, Stephan; Baden, Alexander David; Lillaney, Kunal; Burns, Randal; Vogelstein, Joshua Tzvi; Schier, Alexander Franz; Lee, Wei-Chung Allen; Jeong, Won-Ki; Lichtman, Jeff William; Engert, Florian

2017-05-18

High-resolution serial-section electron microscopy (ssEM) makes it possible to investigate the dense meshwork of axons, dendrites, and synapses that form neuronal circuits. However, the imaging scale required to comprehensively reconstruct these structures is more than ten orders of magnitude smaller than the spatial extents occupied by networks of interconnected neurons, some of which span nearly the entire brain. Difficulties in generating and handling data for large volumes at nanoscale resolution have thus restricted vertebrate studies to fragments of circuits. These efforts were recently transformed by advances in computing, sample handling, and imaging techniques, but high-resolution examination of entire brains remains a challenge. Here, we present ssEM data for the complete brain of a larval zebrafish (Danio rerio) at 5.5 days post-fertilization. Our approach utilizes multiple rounds of targeted imaging at different scales to reduce acquisition time and data management requirements. The resulting dataset can be analysed to reconstruct neuronal processes, permitting us to survey all myelinated axons (the projectome). These reconstructions enable precise investigations of neuronal morphology, which reveal remarkable bilateral symmetry in myelinated reticulospinal and lateral line afferent axons. We further set the stage for whole-brain structure-function comparisons by co-registering functional reference atlases and in vivo two-photon fluorescence microscopy data from the same specimen. All obtained images and reconstructions are provided as an open-access resource.

Whole-brain serial-section electron microscopy in larval zebrafish

NASA Astrophysics Data System (ADS)

Hildebrand, David Grant Colburn; Cicconet, Marcelo; Torres, Russel Miguel; Choi, Woohyuk; Quan, Tran Minh; Moon, Jungmin; Wetzel, Arthur Willis; Scott Champion, Andrew; Graham, Brett Jesse; Randlett, Owen; Plummer, George Scott; Portugues, Ruben; Bianco, Isaac Henry; Saalfeld, Stephan; Baden, Alexander David; Lillaney, Kunal; Burns, Randal; Vogelstein, Joshua Tzvi; Schier, Alexander Franz; Lee, Wei-Chung Allen; Jeong, Won-Ki; Lichtman, Jeff William; Engert, Florian

2017-05-01

High-resolution serial-section electron microscopy (ssEM) makes it possible to investigate the dense meshwork of axons, dendrites, and synapses that form neuronal circuits. However, the imaging scale required to comprehensively reconstruct these structures is more than ten orders of magnitude smaller than the spatial extents occupied by networks of interconnected neurons, some of which span nearly the entire brain. Difficulties in generating and handling data for large volumes at nanoscale resolution have thus restricted vertebrate studies to fragments of circuits. These efforts were recently transformed by advances in computing, sample handling, and imaging techniques, but high-resolution examination of entire brains remains a challenge. Here, we present ssEM data for the complete brain of a larval zebrafish (Danio rerio) at 5.5 days post-fertilization. Our approach utilizes multiple rounds of targeted imaging at different scales to reduce acquisition time and data management requirements. The resulting dataset can be analysed to reconstruct neuronal processes, permitting us to survey all myelinated axons (the projectome). These reconstructions enable precise investigations of neuronal morphology, which reveal remarkable bilateral symmetry in myelinated reticulospinal and lateral line afferent axons. We further set the stage for whole-brain structure-function comparisons by co-registering functional reference atlases and in vivo two-photon fluorescence microscopy data from the same specimen. All obtained images and reconstructions are provided as an open-access resource.

Rigorous quantitative elemental microanalysis by scanning electron microscopy/energy dispersive x-ray spectrometry (SEM/EDS) with spectrum processing by NIST DTSA-II

NASA Astrophysics Data System (ADS)

Newbury, Dale E.; Ritchie, Nicholas W. M.

2014-09-01

Quantitative electron-excited x-ray microanalysis by scanning electron microscopy/silicon drift detector energy dispersive x-ray spectrometry (SEM/SDD-EDS) is capable of achieving high accuracy and high precision equivalent to that of the high spectral resolution wavelength dispersive x-ray spectrometer even when severe peak interference occurs. The throughput of the SDD-EDS enables high count spectra to be measured that are stable in calibration and resolution (peak shape) across the full deadtime range. With this high spectral stability, multiple linear least squares peak fitting is successful for separating overlapping peaks and spectral background. Careful specimen preparation is necessary to remove topography on unknowns and standards. The standards-based matrix correction procedure embedded in the NIST DTSA-II software engine returns quantitative results supported by a complete error budget, including estimates of the uncertainties from measurement statistics and from the physical basis of the matrix corrections. NIST DTSA-II is available free for Java-platforms at: http://www.cstl.nist.gov/div837/837.02/epq/dtsa2/index.html).

High-resolution imaging of living mammalian cells bound by nanobeads-connected antibodies in a medium using scanning electron-assisted dielectric microscopy

NASA Astrophysics Data System (ADS)

Okada, Tomoko; Ogura, Toshihiko

2017-02-01

Nanometre-scale-resolution imaging technologies for liquid-phase specimens are indispensable tools in various scientific fields. In biology, observing untreated living cells in a medium is essential for analysing cellular functions. However, nanoparticles that bind living cells in a medium are hard to detect directly using traditional optical or electron microscopy. Therefore, we previously developed a novel scanning electron-assisted dielectric microscope (SE-ADM) capable of nanoscale observations. This method enables observation of intact cells in aqueous conditions. Here, we use this SE-ADM system to clearly observe antibody-binding nanobeads in liquid-phase. We also report the successful direct detection of streptavidin-conjugated nanobeads binding to untreated cells in a medium via a biotin-conjugated anti-CD44 antibody. Our system is capable of obtaining clear images of cellular organelles and beads on the cells at the same time. The direct observation of living cells with nanoparticles in a medium allowed by our system may contribute the development of carriers for drug delivery systems (DDS).

High spatial resolution PEELS characterization of FeAl nanograins prepared by mechanical alloying

DOE Office of Scientific and Technical Information (OSTI.GOV)

Valdre, G.; Botton, G.A.; Brown, L.M.

The authors investigate the nanograin ``chemical`` structure in a nanostructured material of possible industrial application (Fe-Al system) prepared by conventional mechanical alloying via ball milling in argon atmosphere. They restrict themselves to the structural and nanochemical behavior of ball-milled nanocrystalline Fe-Al powders with atomic composition Fe{sub 3}Al, corresponding to a well-known intermetallic compound of the Fe-Al system. Scanning transmission electron microscopy (STEM) equipped with a parallel detection electron energy loss spectrometer (PEELS) has provided an insight on the ``chemical`` structure of both nanograins and their surface at a spatial resolution of better than 1 nm. The energy loss near edgemore » structure of the Al L loss reveals that the Al coordination is similar to a B2 compound and the oxidation of the powder during processing may play a significant role in the stabilization of the intermetallic phases. Conventional transmission electron microscopy (TEM) was used for the structural characterization of the material after the ball milling; powder X-ray diffraction (XRD) aided the investigation.« less

High Resolution Transmission Electron Microscopy (HRTEM) of nanophase ferric oxides

NASA Technical Reports Server (NTRS)

Golden, D. C.; Morris, R. V.; Ming, D. W.; Lauer, H. V., Jr.

1994-01-01

Iron oxide minerals are the prime candidates for Fe(III) signatures in remotely sensed Martian surface spectra. Magnetic, Mossbauer, and reflectance spectroscopy have been carried out in the laboratory in order to understand the mineralogical nature of Martian analog ferric oxide minerals of submicron or nanometer size range. Out of the iron oxide minerals studied, nanometer sized ferric oxides are promising candidates for possible Martian spectral analogs. 'Nanophase ferric oxide (np-Ox)' is a generic term for ferric oxide/oxihydroxide particles having nanoscale (less than 10 nm) particle dimensions. Ferrihydrite, superparamagnetic particles of hematite, maghemite and goethite, and nanometer sized particles of inherently paramagnetic lepidocrocite are all examples of nanophase ferric oxides. np-Ox particles in general do not give X-ray diffraction (XRD) patterns with well defined peaks and would often be classified as X-ray amorphous. Therefore, different np-Oxs preparations should be characterized using a more sensitive technique e.g., high resolution transmission electron microscopy (HRTEM). The purpose of this study is to report the particle size, morphology and crystalline order, of five np-Ox samples by HRTEM imaging and electron diffraction (ED).

High-resolution electron microscopic evidence for the filamentous structure of the cyst wall in Giardia muris and Giardia duodenalis.

PubMed

Erlandsen, S L; Bemrick, W J; Pawley, J

1989-10-01

High-resolution morphological studies of the cyst wall of Giardia spp. were performed using low-voltage scanning electron microscopy (LVSEM) and transmission electron microscopy (TEM). The cyst wall was composed of membranous and filamentous layers. The membranous layer consisted of an inner and an outer cyst membrane separated by a thin layer of cytoplasm. The filamentous layer contained individual filaments that ranged from 7 to 20 nm in diameter when measured by LVSEM, formed a dense meshwork with branches or interconnections, and were occasionally arranged on the surface in whorled patterns. Cysts of Giardia muris from mice, Giardia duodenalis from dogs, pigs, voles, beavers, muskrats, and humans, and Giardia psittaci from a bird (parakeet), possessed an essentially identical wall composed of filaments. Inducement of excystation in viable Giardia cysts produced a dramatic increase in the interfilament spacing over an entire cyst, but none was observed in heat-killed or chemically fixed control cysts. These results demonstrated that the cyst wall of Giardia spp. was composed of a complex arrangement of filaments, presumably formed during the process of encystment.

3-D Ultrastructure of O. tauri: Electron Cryotomography of an Entire Eukaryotic Cell

PubMed Central

Henderson, Gregory P.; Gan, Lu; Jensen, Grant J.

2007-01-01

The hallmark of eukaryotic cells is their segregation of key biological functions into discrete, membrane-bound organelles. Creating accurate models of their ultrastructural complexity has been difficult in part because of the limited resolution of light microscopy and the artifact-prone nature of conventional electron microscopy. Here we explored the potential of the emerging technology electron cryotomography to produce three-dimensional images of an entire eukaryotic cell in a near-native state. Ostreococcus tauri was chosen as the specimen because as a unicellular picoplankton with just one copy of each organelle, it is the smallest known eukaryote and was therefore likely to yield the highest resolution images. Whole cells were imaged at various stages of the cell cycle, yielding 3-D reconstructions of complete chloroplasts, mitochondria, endoplasmic reticula, Golgi bodies, peroxisomes, microtubules, and putative ribosome distributions in-situ. Surprisingly, the nucleus was seen to open long before mitosis, and while one microtubule (or two in some predivisional cells) was consistently present, no mitotic spindle was ever observed, prompting speculation that a single microtubule might be sufficient to segregate multiple chromosomes. PMID:17710148

Long helical filaments are not seen encircling cells in electron cryotomograms of rod-shaped bacteria

DOE Office of Scientific and Technical Information (OSTI.GOV)

Swulius, Matthew T.; Chen, Songye; Jane Ding, H.

2011-04-22

Highlights: {yields} No long helical filaments are seen near or along rod-shaped bacterial inner membranes by electron cryo-tomography. {yields} Electron cryo-tomography has the resolution to detect single filaments in vivo. -- Abstract: How rod-shaped bacteria form and maintain their shape is an important question in bacterial cell biology. Results from fluorescent light microscopy have led many to believe that the actin homolog MreB and a number of other proteins form long helical filaments along the inner membrane of the cell. Here we show using electron cryotomography of six different rod-shaped bacterial species, at macromolecular resolution, that no long (>80 nm)more » helical filaments exist near or along either surface of the inner membrane. We also use correlated cryo-fluorescent light microscopy (cryo-fLM) and electron cryo-tomography (ECT) to identify cytoplasmic bundles of MreB, showing that MreB filaments are detectable by ECT. In light of these results, the structure and function of MreB must be reconsidered: instead of acting as a large, rigid scaffold that localizes cell-wall synthetic machinery, moving MreB complexes may apply tension to growing peptidoglycan strands to ensure their orderly, linear insertion.« less

Fine structural features of nanoscale zero-valent iron characterized by spherical aberration corrected scanning transmission electron microscopy (Cs-STEM).

PubMed

Liu, Airong; Zhang, Wei-xian

2014-09-21

An angstrom-resolution physical model of nanoscale zero-valent iron (nZVI) is generated with a combination of spherical aberration corrected scanning transmission electron microscopy (Cs-STEM), selected area electron diffraction (SAED), energy-dispersive X-ray spectroscopy (EDS) and electron energy-loss spectroscopy (EELS) on the Fe L-edge. Bright-field (BF), high-angle annular dark-field (HAADF) and secondary electron (SE) imaging of nZVI acquired by a Hitachi HD-2700 STEM show near atomic resolution images and detailed morphological and structural information of nZVI. The STEM-EDS technique confirms that the fresh nZVI comprises of a metallic iron core encapsulated with a thin layer of iron oxides or oxyhydroxides. SAED patterns of the Fe core suggest the polycrystalline structure in the metallic core and amorphous nature of the oxide layer. Furthermore, Fe L-edge of EELS shows varied structural features from the innermost Fe core to the outer oxide shell. A qualitative analysis of the Fe L(2,3) edge fine structures reveals that the shell of nZVI consists of a mixed Fe(II)/Fe(III) phase close to the Fe (0) interface and a predominantly Fe(III) at the outer surface of nZVI.

Thermal magnetic field noise limits resolution in transmission electron microscopy.

PubMed

Uhlemann, Stephan; Müller, Heiko; Hartel, Peter; Zach, Joachim; Haider, Max

2013-07-26

The resolving power of an electron microscope is determined by the optics and the stability of the instrument. Recently, progress has been obtained towards subångström resolution at beam energies of 80 kV and below but a discrepancy between the expected and achieved instrumental information limit has been observed. Here we show that magnetic field noise from thermally driven currents in the conductive parts of the instrument is the root cause for this hitherto unexplained decoherence phenomenon. We demonstrate that the deleterious effect depends on temperature and at least weakly on the type of material.

Helium Ion Secondary Electron Mode Microscopy For Interconnect Material Imaging

NASA Astrophysics Data System (ADS)

Ogawa, Shinichi; Thompson, William; Stern, Lewis; Scipioni, Larry; Notte, John; Farkas, Lou; Barriss, Louise

2010-04-01

The recently developed helium ion microscope (HIM) is now capable of 0.35 nm secondary electron (SE) mode image resolution. When low-k dielectrics or copper interconnects in ultra large scale integrated circuits (ULSI) interconnect structures were imaged in this mode, it was found that unique pattern dimension and fidelity information at sub-nanometer resolution was available for the first time. This paper will discuss the helium ion microscope architecture and the SE imaging techniques that make the HIM observation method of particular value to the low-k dielectric and dual damascene copper interconnect technologies.

Soot Nanostructure: Using Fringe Analysis Software on High Resolution Transmission Electron Microscopy of Carbon Soot

NASA Technical Reports Server (NTRS)

King, James D.

2004-01-01

Using high resolution transmission electron images of carbon nanotubes and carbon particles, we are able to use image analysis program to determine several carbon fringe properties, including length, separation, curvature and orientation. Results are shown in the form of histograms for each of those quantities. The combination of those measurements can give a better indication of the graphic structure within nanotubes and particles of carbon and can distinguish carbons based upon fringe properties. Carbon with longer, straighter and closer spaced fringes are considered graphite, while amorphous carbon contain shorter, less structured fringes.

Graphitic carbon in the Allende meteorite - A microstructural study

NASA Technical Reports Server (NTRS)

Smith, P. P. K.; Buseck, P. R.

1981-01-01

High-resolution transmission electron microscopy shows that carbon in the Allende carbonaceous chondrite meteorite is predominantly a poorly crystalline graphite. Such material is of interest as an important carrier of the isotopically anomalous noble gases found in carbonaceous chondrites.

Cryo-scanning transmission electron tomography of vitrified cells.

PubMed

Wolf, Sharon Grayer; Houben, Lothar; Elbaum, Michael

2014-04-01

Cryo-electron tomography (CET) of fully hydrated, vitrified biological specimens has emerged as a vital tool for biological research. For cellular studies, the conventional imaging modality of transmission electron microscopy places stringent constraints on sample thickness because of its dependence on phase coherence for contrast generation. Here we demonstrate the feasibility of using scanning transmission electron microscopy for cryo-tomography of unstained vitrified specimens (CSTET). We compare CSTET and CET for the imaging of whole bacteria and human tissue culture cells, finding favorable contrast and detail in the CSTET reconstructions. Particularly at high sample tilts, the CSTET signals contain more informative data than energy-filtered CET phase contrast images, resulting in improved depth resolution. Careful control over dose delivery permits relatively high cumulative exposures before the onset of observable beam damage. The increase in acceptable specimen thickness broadens the applicability of electron cryo-tomography.

Serial sectioning methods for 3D investigations in materials science.

PubMed

Zankel, Armin; Wagner, Julian; Poelt, Peter

2014-07-01

A variety of methods for the investigation and 3D representation of the inner structure of materials has been developed. In this paper, techniques based on slice and view using scanning microscopy for imaging are presented and compared. Three different methods of serial sectioning combined with either scanning electron or scanning ion microscopy or atomic force microscopy (AFM) were placed under scrutiny: serial block-face scanning electron microscopy, which facilitates an ultramicrotome built into the chamber of a variable pressure scanning electron microscope; three-dimensional (3D) AFM, which combines an (cryo-) ultramicrotome with an atomic force microscope, and 3D FIB, which delivers results by slicing with a focused ion beam. These three methods complement one another in many respects, e.g., in the type of materials that can be investigated, the resolution that can be obtained and the information that can be extracted from 3D reconstructions. A detailed review is given about preparation, the slice and view process itself, and the limitations of the methods and possible artifacts. Applications for each technique are also provided. Copyright © 2014 Elsevier Ltd. All rights reserved.

Organic Dye Degradation Under Solar Irradiation by Hydrothermally Synthesized ZnS Nanospheres

NASA Astrophysics Data System (ADS)

Samanta, Dhrubajyoti; Chanu, T. Inakhunbi; Basnet, Parita; Chatterjee, Somenath

2018-02-01

The green synthesis of ZnS nanospheres using Citrus limetta (sweet lime) juice as a capping agent through a conventional hydrothermal method was studied. The particle size, morphology, chemical composition, band gap, and optical properties of the synthesized ZnS nanospheres were characterized using x-ray diffraction spectroscopy, field emission scanning electron microscopy, high-resolution transmission electron microscopy, and ultraviolet-visible spectroscopy. The photocatalytic activity of the ZnS nanospheres was evaluated by degradation of rhodamine B (RhB) and methyl orange (MO) under solar irradiation. Upon 150 min of solar irradiation, the extent of degradation was 94% and 77% for RhB and MO, respectively.

Doping effect of polyaniline/MWCNT composites on capacitance and cyclic stability of supercapacitors.

PubMed

Karthikeyan, G; Sahoo, S; Nayak, G C; Das, C K

2012-03-01

Polyaniline doped by Zn2+ ions was synthesized as nanocomposites with multiwalled carbon nanotubes (MWCNT) by in-situ oxidative polymerization and investigated as electrode material for supercapacitors. The uniform coating of polyaniline on MWCNT was characterized by field emission scanning electron microscopy (FESEM) and high resolution transmission electron microscopy (HRTEM). The effect of Zn2+ ions on nanocomposites were characterized by Fourier transform infrared (FTIR) spectroscopy. The electrochemical performances were investigated by cyclic voltammetry (CV), constant current charging/discharging cyclic test (CC) and electrochemical impedance spectroscopy (EIS) using a three-electrode system. The doped polyaniline composites show higher specific capacitance and better cyclic stability.

In search of the elusive IrB 2: Can mechanochemistry help?

DOE PAGES

Xie, Zhilin; Blair, Richard G.; Orlovskaya, Nina; ...

2015-10-20

We produced hexagonal ReB 2-type IrB 2 diboride and orthorhombic IrB monoboride phases, that were previously unknown and saw them produced by mechanochemical syntheses. High energy ball milling of elemental Ir and B powder for 30 h, followed by annealing of the powder at 1050 °C for 48 h, resulted in the formation of the desired phases. Both traditional laboratory and high resolution synchrotron X-ray diffraction (XRD) analyses were used for phase identification of the synthesized powder. Additionally, scanning electron microscopy and transmission electron microscopy were employed, along with XRD, to further characterize the microstructure of the phases produced.

In search of the elusive IrB 2: Can mechanochemistry help?

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xie, Zhilin; Blair, Richard G.; Orlovskaya, Nina

We produced hexagonal ReB 2-type IrB 2 diboride and orthorhombic IrB monoboride phases, that were previously unknown and saw them produced by mechanochemical syntheses. High energy ball milling of elemental Ir and B powder for 30 h, followed by annealing of the powder at 1050 °C for 48 h, resulted in the formation of the desired phases. Both traditional laboratory and high resolution synchrotron X-ray diffraction (XRD) analyses were used for phase identification of the synthesized powder. Additionally, scanning electron microscopy and transmission electron microscopy were employed, along with XRD, to further characterize the microstructure of the phases produced.

Synthesis of nanostructured vanadium powder by high-energy ball milling: X-ray diffraction and high-resolution electron microscopy characterization

NASA Astrophysics Data System (ADS)

Krishnan, Vinoadh Kumar; Sinnaeruvadi, Kumaran

2016-10-01

Vanadium metal powders, ball milled with different surfactants viz., stearic acid, KCl and NaCl, have been studied by X-ray diffraction and transmission electron microscopy. The surfactants alter the microstructural and morphological characteristics of the powders. Ball milling with stearic acid results in solid-state amorphization, while powders milled with KCl yield vanadium-tungsten carbide nanocomposite mixtures. NaCl proved to be an excellent surfactant for obtaining nanostructured fusion-grade vanadium powders. In order to understand the reaction mechanism behind any interstitial addition in the ball-milled powders, CHNOS analysis was performed.

Metallocarbohedrenes: Transmission Electron Microscopy of Mass Gated Deposits

NASA Astrophysics Data System (ADS)

Castleman, M. E. Lyn, Jr.

2002-03-01

Titanium and zirconium Met-Car cluster ions have been detected from the direct laser vaporization of metal-graphite mixtures using time-of-flight mass spectrometry. Optimization of the production conditions enabled sufficient intensities to mass select and deposit Met-Cars on surfaces. High-resolution transmission electron microscopy images of mass gated Met-Car species reveals deposited nanocrystals 2 nm in diameter. Diffraction patterns indicate the presence of multiple species and shows that the deposits have spatial orientation. Lattice parameters have been extracted. The implication of the findings will be discussed. Support for the work has been from the AFOSR F49620-01-1-0122.

A high resolution electron microscopy investigation of curvature in carbon nanotubes

NASA Astrophysics Data System (ADS)

Weldon, D. N.; Blau, W. J.; Zandbergen, H. W.

1995-07-01

Evidence for heptagon inclusion in multi-walled carbon nanotubes was sought in arc-produced carbon deposits. Transmission electron microscopy revealed many curved nanotubes although their relative abundance was low. Close examination of the micrographs in the regions of expected heptagon inclusion shows that the curvature is accomplished by folding or fracture of the lattice planes. This observed phenomenon contradicts the theoretical modelling studies which predict stable structures with negative curvature accomplished by heptagon/pentagon pairs. A possible explanation for curvature in single-walled tubes is presented based on a molecular mechanics geometry optimisation study of spa inclusion in a graphite sheet.

Micelle-assisted fabrication of necklace-shaped assembly of inorganic fullerene-like molybdenum disulfide nanospheres

NASA Astrophysics Data System (ADS)

Xiong, Yujie; Xie, Yi; Li, Zhengquan; Li, Xiaoxu; Zhang, Rong

2003-11-01

The fabrication of necklace-shaped assembly of inorganic fullerene-like molybdenum disulfide nanospheres via a micelle-assisted route is reported, in which necklace-shaped assembly of amorphous MoS 3 nanospheres is driven by the aggregation transformation of surfactants at low temperatures and then is transformed to the assembly of target fullerene-like MoS 2 by annealing. This nanostructure is a type of oriented assembly of inorganic fullerene-like structures, which is confirmed by the transmission electron microscopy and high-resolution transmission electron microscopy analysis. The optical absorption property is investigated to show their inorganic fullerene-like structure and uniform shape.

Synthesis of metal nanoparticle and patterning in polymeric films induced by electron beam

NASA Astrophysics Data System (ADS)

Yamamoto, Hiroki; Kozawa, Takahiro; Tagawa, Seiichi; Marignier, Jean-Louis; Mostafavi, Mehran; Belloni, Jacqueline

2018-03-01

Using an electron beam, thin polymeric films loaded with metal nanoparticles of silver were prepared by a one-step irradiation-induced reduction of the metal ions embedded in the polymer. The metal nanoparticles were observed by either optical absorption or microscopy. The mechanism of the reduction of metal ions and of the polymer crosslinking were deduced from the average absorbance measurements. In view of realizing specific patterns of high resolution using the electron beam, electron beam produces 200 nm wide lines that can be separated by unexposed spaces of adjustable width, where precursors were dissolved. The resolution of the electron beam has been exploited to demonstrate the achievement of nanopatterning on polymer films using a direct-writing process. This method supplies interesting applications such as masks, replicas, or imprint molds of improved density and contrast.

Low-energy electron holographic imaging of individual tobacco mosaic virions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Longchamp, Jean-Nicolas, E-mail: longchamp@physik.uzh.ch; Latychevskaia, Tatiana; Escher, Conrad

2015-09-28

Modern structural biology relies on Nuclear Magnetic Resonance (NMR), X-ray crystallography, and cryo-electron microscopy for gaining information on biomolecules at nanometer, sub-nanometer, or atomic resolution. All these methods, however, require averaging over a vast ensemble of entities, and hence knowledge on the conformational landscape of an individual particle is lost. Unfortunately, there are now strong indications that even X-ray free electron lasers will not be able to image individual molecules but will require nanocrystal samples. Here, we show that non-destructive structural biology of single particles has now become possible by means of low-energy electron holography. As an example, individual tobaccomore » mosaic virions deposited on ultraclean freestanding graphene are imaged at 1 nm resolution revealing structural details arising from the helical arrangement of the outer protein shell of the virus. Since low-energy electron holography is a lens-less technique and since electrons with a deBroglie wavelength of approximately 1 Å do not impose radiation damage to biomolecules, the method has the potential for Angstrom resolution imaging of single biomolecules.« less

Instrumental requirements for the detection of electron beam-induced object excitations at the single atom level in high-resolution transmission electron microscopy.

PubMed

Kisielowski, C; Specht, P; Gygax, S M; Barton, B; Calderon, H A; Kang, J H; Cieslinski, R

2015-01-01

This contribution touches on essential requirements for instrument stability and resolution that allows operating advanced electron microscopes at the edge to technological capabilities. They enable the detection of single atoms and their dynamic behavior on a length scale of picometers in real time. It is understood that the observed atom dynamic is intimately linked to the relaxation and thermalization of electron beam-induced sample excitation. Resulting contrast fluctuations are beam current dependent and largely contribute to a contrast mismatch between experiments and theory if not considered. If explored, they open the possibility to study functional behavior of nanocrystals and single molecules at the atomic level in real time. Copyright © 2014 Elsevier Ltd. All rights reserved.

Removing Contamination-Induced Reconstruction Artifacts from Cryo-electron Tomograms

PubMed Central

Fernandez, Jose-Jesus; Laugks, Ulrike; Schaffer, Miroslava; Bäuerlein, Felix J.B.; Khoshouei, Maryam; Baumeister, Wolfgang; Lucic, Vladan

2016-01-01

Imaging of fully hydrated, vitrified biological samples by electron tomography yields structural information about cellular protein complexes in situ. Here we present a computational procedure that removes artifacts of three-dimensional reconstruction caused by contamination present in samples during imaging by electron microscopy. Applying the procedure to phantom data and electron tomograms of cellular samples significantly improved the resolution and the interpretability of tomograms. Artifacts caused by surface contamination associated with thinning by focused ion beam, as well as those arising from gold fiducial markers and from common, lower contrast contamination, could be removed. Our procedure is widely applicable and is especially suited for applications that strive to reach a higher resolution and involve the use of recently developed, state-of-the-art instrumentation. PMID:26743046

Quantifying the cellular uptake of semiconductor quantum dot nanoparticles by analytical electron microscopy.

PubMed

Hondow, Nicole; Brown, M Rowan; Starborg, Tobias; Monteith, Alexander G; Brydson, Rik; Summers, Huw D; Rees, Paul; Brown, Andy

2016-02-01

Semiconductor quantum dot nanoparticles are in demand as optical biomarkers yet the cellular uptake process is not fully understood; quantification of numbers and the fate of internalized particles are still to be achieved. We have focussed on the characterization of cellular uptake of quantum dots using a combination of analytical electron microscopies because of the spatial resolution available to examine uptake at the nanoparticle level, using both imaging to locate particles and spectroscopy to confirm identity. In this study, commercially available quantum dots, CdSe/ZnS core/shell particles coated in peptides to target cellular uptake by endocytosis, have been investigated in terms of the agglomeration state in typical cell culture media, the traverse of particle agglomerates across U-2 OS cell membranes during endocytosis, the merging of endosomal vesicles during incubation of cells and in the correlation of imaging flow cytometry and transmission electron microscopy to measure the final nanoparticle dose internalized by the U-2 OS cells. We show that a combination of analytical transmission electron microscopy and serial block face scanning electron microscopy can provide a comprehensive description of the internalization of an initial exposure dose of nanoparticles by an endocytically active cell population and how the internalized, membrane bound nanoparticle load is processed by the cells. We present a stochastic model of an endosome merging process and show that this provides a data-driven modelling framework for the prediction of cellular uptake of engineered nanoparticles in general. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

Diatom Valve Three-Dimensional Representation: A New Imaging Method Based on Combined Microscopies

PubMed Central

Ferrara, Maria Antonietta; De Tommasi, Edoardo; Coppola, Giuseppe; De Stefano, Luca; Rea, Ilaria; Dardano, Principia

2016-01-01

The frustule of diatoms, unicellular microalgae, shows very interesting photonic features, generally related to its complicated and quasi-periodic micro- and nano-structure. In order to simulate light propagation inside and through this natural structure, it is important to develop three-dimensional (3D) models for synthetic replica with high spatial resolution. In this paper, we present a new method that generates images of microscopic diatoms with high definition, by merging scanning electron microscopy and digital holography microscopy or atomic force microscopy data. Starting from two digital images, both acquired separately with standard characterization procedures, a high spatial resolution (Δz = λ/20, Δx = Δy ≅ 100 nm, at least) 3D model of the object has been generated. Then, the two sets of data have been processed by matrix formalism, using an original mathematical algorithm implemented on a commercially available software. The developed methodology could be also of broad interest in the design and fabrication of micro-opto-electro-mechanical systems. PMID:27690008

How good can cryo-EM become?

DOE Office of Scientific and Technical Information (OSTI.GOV)

Glaeser, Robert M.

The suddenness with which single-particle cryo-electron microscopy (cryo-EM) has emerged as a method for determining high-resolution structures of biological macromolecules invites the questions, how much better can this technology get, and how fast is that likely to happen? While we can rightly celebrate the maturation of cryo-EM as a high-resolution structure-determination tool, I believe there still are many developments to look forward to.

High-Resolution Detector For X-Ray Diffraction

NASA Technical Reports Server (NTRS)

Carter, Daniel C.; Withrow, William K.; Pusey, Marc L.; Yost, Vaughn H.

1988-01-01

Proposed x-ray-sensitive imaging detector offers superior spatial resolution, counting-rate capacity, and dynamic range. Instrument based on laser-stimulated luminescence and reusable x-ray-sensitive film. Detector scans x-ray film line by line. Extracts latent image in film and simultaneously erases film for reuse. Used primarily for protein crystallography. Principle adapted to imaging detectors for electron microscopy and fluorescence spectroscopy and general use in astronomy, engineering, and medicine.

Atmospheric pressure scanning transmission electron microscopy.

PubMed

de Jonge, Niels; Bigelow, Wilbur C; Veith, Gabriel M

2010-03-10

Scanning transmission electron microscope (STEM) images of gold nanoparticles at atmospheric pressure have been recorded through a 0.36 mm thick mixture of CO, O2, and He. This was accomplished using a reaction cell consisting of two electron-transparent silicon nitride membranes. Gold nanoparticles of a full width at half-maximum diameter of 1.0 nm were visible above the background noise, and the achieved edge resolution was 0.4 nm in accordance with calculations of the beam broadening.

Atomic Resolution Cryo-EM Structure of β-Galactosidase.

PubMed

Bartesaghi, Alberto; Aguerrebere, Cecilia; Falconieri, Veronica; Banerjee, Soojay; Earl, Lesley A; Zhu, Xing; Grigorieff, Nikolaus; Milne, Jacqueline L S; Sapiro, Guillermo; Wu, Xiongwu; Subramaniam, Sriram

2018-05-10

The advent of direct electron detectors has enabled the routine use of single-particle cryo-electron microscopy (EM) approaches to determine structures of a variety of protein complexes at near-atomic resolution. Here, we report the development of methods to account for local variations in defocus and beam-induced drift, and the implementation of a data-driven dose compensation scheme that significantly improves the extraction of high-resolution information recorded during exposure of the specimen to the electron beam. These advances enable determination of a cryo-EM density map for β-galactosidase bound to the inhibitor phenylethyl β-D-thiogalactopyranoside where the ordered regions are resolved at a level of detail seen in X-ray maps at ∼ 1.5 Å resolution. Using this density map in conjunction with constrained molecular dynamics simulations provides a measure of the local flexibility of the non-covalently bound inhibitor and offers further opportunities for structure-guided inhibitor design. Published by Elsevier Ltd.

A High Resolution Look at Black Sand Particles from Sand Dunes of Saudi Arabia Using Electron Microscopy

NASA Astrophysics Data System (ADS)

Hussain, M. M.; Aburizaiza, O. S.; Siddique, A.; Hershey, D. L.; Guerrieri, D. A.; Qurashi, J.; Abbass, M.; Blake, D. R.; Khwaja, H. A.

2013-12-01

Particulate air pollution is a problem of health concern. The microscopic make-up of different varieties of sand particles found and collected at a sand dune site in Badr, Saudi Arabia has been determined. Primary emphasis is given to the use of multiple high resolution electron microscopy (viz., Scanning Electron Microscopy with Energy Dispersive X-ray spectrometry (SEM/EDS) and Laser Scanning Microscopy (LSM)) to study the morphologies, emission source types, size, and elemental composition of the particles, and to evaluate the presence of ';coatings or contaminants' adsorbed or carried on by the black sand particles. White sand contains natural coarse particles associated with wind-blown releases from crustal surfaces, weathering of an igneous/metamorphic rock source, and volcanic activities. Silicates (alumino-silicates) and quartz (clear, milky, rose) dominate white sand and rest appears to contain calcite, olivine, feldspar, and magnetite. Black sand particles exhibit very different morphologies and microstructures (surface roughness) compared with white sand and volcanic ash. Morphological analyses have shown that the black sand contain ultrafine particles. Black sand is strongly magnetic, which indicates the mineral magnetite (strongly magnetic) or elemental iron. Iron, C, O, Ti, Si, V, and S particles dominate the black sand. Natural and anthropogenic sources have been implicated for the observed particles. Analysis revealed that the surface of white sand particles is mainly covered with the fine particles. It is known that emissions from combustion contain carbon soot and other contaminants that are easily absorbed by soil particles during a long-range transport.

Nanoscopic analysis of oxygen segregation at tilt boundaries in silicon ingots using atom probe tomography combined with TEM and ab initio calculations.

PubMed

Ohno, Y; Inoue, K; Fujiwara, K; Kutsukake, K; Deura, M; Yonenaga, I; Ebisawa, N; Shimizu, Y; Inoue, K; Nagai, Y; Yoshida, H; Takeda, S; Tanaka, S; Kohyama, M

2017-12-01

We have developed an analytical method to determine the segregation levels on the same tilt boundaries (TBs) at the same nanoscopic location by a joint use of atom probe tomography and scanning transmission electron microscopy, and discussed the mechanism of oxygen segregation at TBs in silicon ingots in terms of bond distortions around the TBs. The three-dimensional distribution of oxygen atoms was determined at the typical small- and large-angle TBs by atom probe tomography with a low impurity detection limit (0.01 at.% on a TB plane) simultaneously with high spatial resolution (about 0.4 nm). The three-dimensional distribution was correlated with the atomic stress around the TBs; the stress at large-angle TBs was estimated by ab initio calculations based on atomic resolution scanning transmission electron microscopy data and that at small-angle TBs were calculated with the elastic theory based on dark-field transmission electron microscopy data. Oxygen atoms would segregate at bond-centred sites under tensile stress above about 2 GPa, so as to attain a more stable bonding network by reducing the local stress. The number of oxygen atoms segregating in a unit TB area N GB (in atoms nm -2 ) was determined to be proportional to both the number of the atomic sites under tensile stress in a unit TB area n bc and the average concentration of oxygen atoms around the TB [O i ] (in at.%) with N GB ∼ 50 n bc [O i ]. © 2017 The Authors Journal of Microscopy © 2017 Royal Microscopical Society.

Rare-earth-doped nanophosphors for multicolor cathodoluminescence nanobioimaging using scanning transmission electron microscopy.

PubMed

Furukawa, Taichi; Fukushima, Shoichiro; Niioka, Hirohiko; Yamamoto, Naoki; Miyake, Jun; Araki, Tsutomu; Hashimoto, Mamoru

2015-05-01

We describe rare-earth-doped nanophosphors (RE-NPs) for biological imaging using cathodoluminescence(CL) microscopy based on scanning transmission electron microscopy (STEM). We report the first demonstration of multicolor CL nanobioimaging using STEM with nanophosphors. The CL spectra of the synthesized nanophosphors (Y2O3∶Eu, Y2O3∶Tb) were sufficiently narrow to be distinguished. From CL images of RE-NPs on an elastic carbon-coated copper grid, the spatial resolution was beyond the diffraction limit of light.Y2O3∶Tb and Y2O3∶Eu RE-NPs showed a remarkable resistance against electron beam exposure even at high acceleration voltage (80 kV) and retained a CL intensity of more than 97% compared with the initial intensity for 1 min. In biological CL imaging with STEM, heavy-metal-stained cell sections containing the RE-NPs were prepared,and both the CL images of RE-NPs and cellular structures, such as mitochondria, were clearly observed from STEM images with high contrast. The cellular CL imaging using RE-NPs also had high spatial resolution even though heavy-metal-stained cells are normally regarded as highly scattering media. Moreover, since theRE-NPs exhibit photoluminescence (PL) excited by UV light, they are useful for multimodal correlative imaging using CL and PL.

A combined optical, SEM and STM study of growth spirals on the polytypic cadmium iodide crystals

NASA Astrophysics Data System (ADS)

Singh, Rajendra; Samanta, S. B.; Narlikar, A. V.; Trigunayat, G. C.

2000-05-01

Some novel results of a combined sequential study of growth spirals on the basal surface of the richly polytypic CdI 2 crystals by optical microscopy, scanning electron microscopy (SEM) and scanning tunneling microscopy (STM) are presented and discussed. Under the high resolution and magnification achieved in the scanning electron microscope, the growth steps of large heights seen in the optical micrographs are found to have a large number of additional steps of smaller heights existing between any two adjacent large height growth steps. When further seen by a scanning tunneling microscope, which provides still higher resolution, sequences of unit substeps, each of height equal to the unit cell height of the underlying polytype, are revealed to exist on the surface. Several large steps also lie between the unit steps, with heights equal to an integral multiple of either the unit cell height of the underlying polytype or the thickness of a molecular sheet I-Cd-I. It is suggested that initially a giant screw dislocation may form by brittle fracture of the crystal platelet, which may gradually decompose into numerous unit dislocations during subsequent crystal growth.

Analyzing indirect secondary electron contrast of unstained bacteriophage T4 based on SEM images and Monte Carlo simulations

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ogura, Toshihiko, E-mail: t-ogura@aist.go.jp

2009-03-06

The indirect secondary electron contrast (ISEC) condition of the scanning electron microscopy (SEM) produces high contrast detection with minimal damage of unstained biological samples mounted under a thin carbon film. The high contrast image is created by a secondary electron signal produced under the carbon film by a low acceleration voltage. Here, we show that ISEC condition is clearly able to detect unstained bacteriophage T4 under a thin carbon film (10-15 nm) by using high-resolution field emission (FE) SEM. The results show that FE-SEM provides higher resolution than thermionic emission SEM. Furthermore, we investigated the scattered electron area within themore » carbon film under ISEC conditions using Monte Carlo simulation. The simulations indicated that the image resolution difference is related to the scattering width in the carbon film and the electron beam spot size. Using ISEC conditions on unstained virus samples would produce low electronic damage, because the electron beam does not directly irradiate the sample. In addition to the routine analysis, this method can be utilized for structural analysis of various biological samples like viruses, bacteria, and protein complexes.« less

Synthesis of Silver and Gold Nanoparticles Using Antioxidants from Blackberry, Blueberry, Pomegranate, and Turmeric Extracts

EPA Science Inventory

Greener synthesis of Ag and Au nanoparticles is described using antioxidants from blackberry, blueberry, pomegranate, and turmeric extracts. The synthesized particles were characterized using X-ray diffraction (XRD), transmission electron microscopy (TEM), high resolution TEM (HR...

STED super-resolution microscopy reveals an array of MINOS clusters along human mitochondria

PubMed Central

Jans, Daniel C.; Wurm, Christian A.; Riedel, Dietmar; Wenzel, Dirk; Stagge, Franziska; Deckers, Markus; Rehling, Peter; Jakobs, Stefan

2013-01-01

The mitochondrial inner membrane organizing system (MINOS) is a conserved large hetero-oligomeric protein complex in the mitochondrial inner membrane, crucial for the maintenance of cristae morphology. MINOS has been suggested to represent the core of an extended protein network that controls mitochondrial function and structure, and has been linked to several human diseases. The spatial arrangement of MINOS within mitochondria is ill-defined, however. Using super-resolution stimulated emission depletion (STED) microscopy and immunogold electron microscopy, we determined the distribution of three known human MINOS subunits (mitofilin, MINOS1, and CHCHD3) in mammalian cells. Super-resolution microscopy revealed that all three subunits form similar clusters within mitochondria, and that MINOS is more abundant in mitochondria around the nucleus than in peripheral mitochondria. At the submitochondrial level, mitofilin, a core MINOS subunit, is preferentially localized at cristae junctions. In primary human fibroblasts, mitofilin labeling uncovered a regularly spaced pattern of clusters arranged in parallel to the cell growth surfaces. We suggest that this array of MINOS complexes might explain the observed phenomenon of largely horizontally arranged cristae junctions that connect the inner boundary membrane to lamellar cristae. The super-resolution images demonstrate an unexpectedly high level of regularity in the nanoscale distribution of the MINOS complex in human mitochondria, supporting an integrating role of MINOS in the structural organization of the organelle. PMID:23676277

Toward tunable doping in graphene FETs by molecular self-assembled monolayers

NASA Astrophysics Data System (ADS)

Li, Bing; Klekachev, Alexander V.; Cantoro, Mirco; Huyghebaert, Cedric; Stesmans, André; Asselberghs, Inge; de Gendt, Stefan; de Feyter, Steven

2013-09-01

In this paper, we report the formation of self-assembled monolayers (SAMs) of oleylamine (OA) on highly oriented pyrolytic graphite (HOPG) and graphene surfaces and demonstrate the potential of using such organic SAMs to tailor the electronic properties of graphene. Molecular resolution Atomic Force Microscopy (AFM) and Scanning Tunneling Microscopy (STM) images reveal the detailed molecular ordering. The electrical measurements show that OA strongly interacts with graphene leading to n-doping effects in graphene devices. The doping levels are tunable by varying the OA deposition conditions. Importantly, neither hole nor electron mobilities are decreased by the OA modification. As a benefit from this noncovalent modification strategy, the pristine characteristics of the device are recoverable upon OA removal. From this study, one can envision the possibility to correlate the graphene-based device performance with the molecular structure and supramolecular ordering of the organic dopant.In this paper, we report the formation of self-assembled monolayers (SAMs) of oleylamine (OA) on highly oriented pyrolytic graphite (HOPG) and graphene surfaces and demonstrate the potential of using such organic SAMs to tailor the electronic properties of graphene. Molecular resolution Atomic Force Microscopy (AFM) and Scanning Tunneling Microscopy (STM) images reveal the detailed molecular ordering. The electrical measurements show that OA strongly interacts with graphene leading to n-doping effects in graphene devices. The doping levels are tunable by varying the OA deposition conditions. Importantly, neither hole nor electron mobilities are decreased by the OA modification. As a benefit from this noncovalent modification strategy, the pristine characteristics of the device are recoverable upon OA removal. From this study, one can envision the possibility to correlate the graphene-based device performance with the molecular structure and supramolecular ordering of the organic dopant. Electronic supplementary information (ESI) available: AFM images of self-assembled monolayers of OA on HOPG; AFM height image of the graphene surface on a SiC substrate; high resolution STM image of a self-assembled monolayer of OA on HOPG; transfer curves of a graphene FET with and without baking steps; transfer curves of a graphene FET under high vacuum conditions; transfer curves of a graphene FET and its Raman response before and after OA treatment; transfer curves of a graphene FET before and after rinsing with n-hexane. See DOI: 10.1039/c3nr01255g

Scattered electrons in microscopy and microanalysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ottensmeyer, F.P.

The use of scattered electrons alone for direct imaging of biological specimens makes it possible to obtain structural information at atomic and near-atomic spatial resolutions of 0.3 to 0.5 nanometer. While this is not as good as the resolution possible with x-ray crystallography, such an approach provides structural information rapidly on individual macromolecules that have not been, and possibly cannot be, crystallized. Analysis of the spectrum of energies of scattered electrons and imaging of the latter with characteristic energy bands within the spectrum produces a powerful new technique of atomic microanalysis. This technique, which has a spatial resolution of aboutmore » 0.5 nanometer and a minimum detection sensitivity of about 50 atoms of phosphorus, is especially useful for light atom analysis and appears to have applications in molecular biology, cell biology, histology, pathology, botany, and many other fields.« less

Scattered electrons in microscopy and microanalysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ottensmeyer, F.P.

The use of scattered electrons alone for direct imaging of biological specimens makes it possible to obtain structural information at atomic and near-atomic spatial resolutions of 0.3 to 0.5 nanometer. While this is not as good as the resolution possible with x-ray crystallography, such an approach provides structural information rapidly on individual macromolecules that have not been, and possibly cannot be, crystallized. Analysis of the spectrum of energies of scattered electrons and imaging of the latter with characteristic energy bands within the spectrum produce a powerful new technique of atomic microanalysis. This technique, which has a spatial resolution of aboutmore » 0.5 nanometer and a minimum detection sensitivity of about 50 atoms of phosphorus, is especially useful for light atom analysis and appears to have applications in molecular biology, cell biology, histology, pathology, botany, and many other fields.« less