Practical electron tomography

1995 ◽  
Vol 53 ◽  
pp. 742-743
Author(s):  
C.L. Woodcock
Keyword(s):  
Electron Tomography ◽  
Tomographic Reconstruction ◽  
Three Dimensional ◽  
3D Shape ◽  
Computational Resources ◽  
Shape Of Particles

Despite the potential of the technique, electron tomography has yet to be widely used by biologists. This is in part related to the rather daunting list of equipment and expertise that are required. Thanks to continuing advances in theory and instrumentation, tomography is now more feasible for the non-specialist. One barrier that has essentially disappeared is the expense of computational resources. In view of this progress, it is time to give more attention to practical issues that need to be considered when embarking on a tomographic project. The following recommendations and comments are derived from experience gained during two long-term collaborative projects.Tomographic reconstruction results in a three dimensional description of an individual EM specimen, most commonly a section, and is therefore applicable to problems in which ultrastructural details within the thickness of the specimen are obscured in single micrographs. Information that can be recovered using tomography includes the 3D shape of particles, and the arrangement and dispostion of overlapping fibrous and membranous structures.

scholarly journals High-resolution three-dimensional probes of biomaterials and their interfaces

2012 ◽  
Vol 370 (1963) ◽  
pp. 1337-1351 ◽  
Author(s):  
Kathryn Grandfield ◽  
Anders Palmquist ◽  
Håkan Engqvist
Keyword(s):  
High Resolution ◽  
Electron Tomography ◽  
Three Dimensional ◽  
Controlled Drug Release ◽  
Biological Tissues ◽  
Dimensional Structure ◽  
Bone Interface ◽  
Biomaterial Interfaces ◽  
Titania Coating

Interfacial relationships between biomaterials and tissues strongly influence the success of implant materials and their long-term functionality. Owing to the inhomogeneity of biological tissues at an interface, in particular bone tissue, two-dimensional images often lack detail on the interfacial morphological complexity. Furthermore, the increasing use of nanotechnology in the design and production of biomaterials demands characterization techniques on a similar length scale. Electron tomography (ET) can meet these challenges by enabling high-resolution three-dimensional imaging of biomaterial interfaces. In this article, we review the fundamentals of ET and highlight its recent applications in probing the three-dimensional structure of bioceramics and their interfaces, with particular focus on the hydroxyapatite–bone interface, titanium dioxide–bone interface and a mesoporous titania coating for controlled drug release.

scholarly journals A Survey of the Use of Iterative Reconstruction Algorithms in Electron Microscopy

2017 ◽  
Vol 2017 ◽  
pp. 1-17 ◽  
Author(s):  
C. O. S. Sorzano ◽  
J. Vargas ◽  
J. Otón ◽  
J. M. de la Rosa-Trevín ◽  
J. L. Vilas ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Electron Tomography ◽  
Tomographic Reconstruction ◽  
Three Dimensional ◽  
Iterative Algorithms ◽  
Projection Methods ◽  
Particle Analysis ◽  
Reconstruction Algorithms ◽  
Large Families ◽  
Key Steps

One of the key steps in Electron Microscopy is the tomographic reconstruction of a three-dimensional (3D) map of the specimen being studied from a set of two-dimensional (2D) projections acquired at the microscope. This tomographic reconstruction may be performed with different reconstruction algorithms that can be grouped into several large families: direct Fourier inversion methods, back-projection methods, Radon methods, or iterative algorithms. In this review, we focus on the latter family of algorithms, explaining the mathematical rationale behind the different algorithms in this family as they have been introduced in the field of Electron Microscopy. We cover their use in Single Particle Analysis (SPA) as well as in Electron Tomography (ET).

scholarly journals Compressed sensing improved iterative reconstruction-reprojection algorithm for electron tomography

2020 ◽  
Vol 21 (S6) ◽  
Author(s):  
Lun Li ◽  
Renmin Han ◽  
Zhaotian Zhang ◽  
Tiande Guo ◽  
Zhiyong Liu ◽  
...  
Keyword(s):  
Compressed Sensing ◽  
Iterative Reconstruction ◽  
Electron Tomography ◽  
Tomographic Reconstruction ◽  
Three Dimensional ◽  
Structural Content ◽  
Execution Speed ◽  
Effectiveness And Efficiency ◽  
Real World Datasets ◽  
Dimensional Object

Abstract Background Electron tomography (ET) is an important technique for the study of complex biological structures and their functions. Electron tomography reconstructs the interior of a three-dimensional object from its projections at different orientations. However, due to the instrument limitation, the angular tilt range of the projections is limited within +70∘ to −70∘. The missing angle range is known as the missing wedge and will cause artifacts. Results In this paper, we proposed a novel algorithm, compressed sensing improved iterative reconstruction-reprojection (CSIIRR), which follows the schedule of improved iterative reconstruction-reprojection but further considers the sparsity of the biological ultra-structural content in specimen. The proposed algorithm keeps both the merits of the improved iterative reconstruction-reprojection (IIRR) and compressed sensing, resulting in an estimation of the electron tomography with faster execution speed and better reconstruction result. A comprehensive experiment has been carried out, in which CSIIRR was challenged on both simulated and real-world datasets as well as compared with a number of classical methods. The experimental results prove the effectiveness and efficiency of CSIIRR, and further show its advantages over the other methods. Conclusions The proposed algorithm has an obvious advance in the suppression of missing wedge effects and the restoration of missing information, which provides an option to the structural biologist for clear and accurate tomographic reconstruction.

Application of electron tomography to elucidate the 3D architecture of diverse asymmetric biological specimens

1993 ◽  
Vol 51 ◽  
pp. 494-495
Author(s):  
Bruce F. McEwen
Keyword(s):  
Electron Tomography ◽  
Tomographic Reconstruction ◽  
Three Dimensional ◽  
Single Copy ◽  
Angular Range ◽  
Electron Dose ◽  
Total Electron ◽  
Reconstruction Methods ◽  
3D Architecture ◽  
3D Information

Electron tomography refers to the use of tomographic reconstruction methods to obtain three-dimensional (3D) information from transmission electron microscopy (TEM). This is accomplished by tilting the sample over a wide angular range with 180° being ideal, but 120° to 140° more typical due to limitations imposed by the geometry of most TEM specimens. If symmetry is present in the sample then the full angular range is not required. Furthermore, if the specimen is present as identical units, either in crystalline arrays or as single particles, the different tilts can be collected from different specimens and hence spare the total electron dose on any given particle. However, many important biological specimens are large, asymmetric objects which, due to their size, are not found as identical individual units. For such cases, the only recourse is to collect all the views from a single copy of the object, generally using a single axis tilt geometry.

scholarly journals Practical Electron Tomography

1996 ◽  
Vol 4 (1) ◽  
pp. 30-38
Author(s):  
C.L. Woodcock
Keyword(s):  
Electron Tomography ◽  
Computational Resources

Despite the potential of the technique, electron tomography has yet to be widely used by biologists. This is in part related to the rather daunting list of equipment and expertise that are required. Thanks to continuing advances in theory and instrumentation, tomography is now more feasible for the non-specialist. One barrier that has essentially disappeared is the expense of computational resources. In view of this progress, it is time to give more attention to practical issues that need to be considered when embarking on a tomographic project. The following recommendations and comments are derived from experience gained during two long-term collaborative projects.

scholarly journals Parakeet: a digital twin software pipeline to assess the impact of experimental parameters on tomographic reconstructions for cryo-electron tomography

Open Biology ◽  
2021 ◽  
Vol 11 (10) ◽  
Author(s):  
James M. Parkhurst ◽  
Maud Dumoux ◽  
Mark Basham ◽  
Daniel Clare ◽  
C. Alistair Siebert ◽  
...  
Keyword(s):  
Data Acquisition ◽  
Biological Samples ◽  
Electron Tomography ◽  
Tomographic Reconstruction ◽  
Three Dimensional ◽  
Software Pipeline ◽  
Digital Twin ◽  
Cryo Electron Tomography ◽  
The Impact

In cryo-electron tomography (cryo-ET) of biological samples, the quality of tomographic reconstructions can vary depending on the transmission electron microscope (TEM) instrument and data acquisition parameters. In this paper, we present Parakeet, a ‘digital twin’ software pipeline for the assessment of the impact of various TEM experiment parameters on the quality of three-dimensional tomographic reconstructions. The Parakeet digital twin is a digital model that can be used to optimize the performance and utilization of a physical instrument to enable in silico optimization of sample geometries, data acquisition schemes and instrument parameters. The digital twin performs virtual sample generation, TEM image simulation, and tilt series reconstruction and analysis within a convenient software framework. As well as being able to produce physically realistic simulated cryo-ET datasets to aid the development of tomographic reconstruction and subtomogram averaging programs, Parakeet aims to enable convenient assessment of the effects of different microscope parameters and data acquisition parameters on reconstruction quality. To illustrate the use of the software, we present the example of a quantitative analysis of missing wedge artefacts on simulated planar and cylindrical biological samples and discuss how data collection parameters can be modified for cylindrical samples where a full 180° tilt range might be measured.

scholarly journals Rapid Method for Electron Tomographic Reconstruction and Three-Dimensional Modeling of the Murine Synapse Using an Automated Fiducial Marker-Free System

2013 ◽  
Vol 19 (S5) ◽  
pp. 182-187 ◽  
Author(s):  
Hyun-wook Kim ◽  
Seung Hak Oh ◽  
Namkug Kim ◽  
Eiko Nakazawa ◽  
Im Joo Rhyu
Keyword(s):  
Electron Tomography ◽  
Tomographic Reconstruction ◽  
Three Dimensional ◽  
3D Models ◽  
Three Dimension ◽  
Synaptic Organization ◽  
Back Projection ◽  
Virtual Image ◽  
Free System ◽  
Three Dimensional Modeling

AbstractElectron tomography (ET) has recently afforded new insights into neuronal architecture. However, the tedious process of sample preparation, image acquisition, alignment, back projection, and additional segmentation process of ET repels beginners. We have tried Hitachi's commercial packages integrated with a Hitachi H-7650 TEM to examine the potential of using an automated fiducial-less approach for our own neuroscience research. Semi-thick sections (200–300 nm) were cut from blocks of fixed mouse (C57BL) cerebellum and prepared for ET. Sets of images were collected automatically as each section was tilted by 2° increments (±60°). “Virtual” image volumes were computationally reconstructed in three dimension (3D) with the EMIP software using either the commonly used “weighted back-projection” (WBP) method or “topography-based reconstruction” (TBR) algorithm for comparison. Computed tomograms using the TBR were more precisely reconstructed compared with the WBP method. Following reconstruction, the image volumes were imported into the 3D editing software A-View and segmented according to synaptic organization. The detailed synaptic components were revealed by very thin virtual image slices; 3D models of synapse structure could be constructed efficiently. Overall, this simplified system provided us with a graspable tool for pursuing ET studies in neuroscience.

Electron tomographic quantitation of mineral distribution in different collagen zones of calcifying tendon

1991 ◽  
Vol 49 ◽  
pp. 190-191
Author(s):  
B. F. McEwen ◽  
W. J. Landis ◽  
M. J. Song
Keyword(s):  
Electron Tomography ◽  
Tomographic Reconstruction ◽  
Three Dimensional ◽  
Organic Matrix ◽  
High Voltage Electron Microscopy ◽  
3D Images ◽  
Mineral Distribution ◽  
Voltage Electron ◽  
First Time

The skeletal system of vertebrates is composed of hard tissue formed by the deposition of hydroxyapatite, a calcium phosphate salt, into an organic matrix that is principally collagen. Although bone and other vertebrate calcifying tissues have been well studied, the processes by which mineralization is initiated and regulated at the molecular level are incompletely understood. Recently these processes have been investigated through high resolution tomographic three-dimensional (3D) reconstruction of initial mineralization sites of calcifying tendon imaged by high-voltage electron microscopy (HVEM). Such tomographic reconstruction provided the first direct 3D images of crystallization sites and clearly established in vivo that, as indicated in earlier reports, initial crystals are thin, irregularly-shaped platelets rather than rod-shaped needles. The crystals develop in both length (along their crystallographic c-axis) and width but seem limited in thickness. Crystals in local tendon regions are approximately parallel to each other and fuse in coplanar alignments to form larger platelets within collagen. Crystal growth is thought to be initiated in hole zones of the protein and develop into adjacent overlap zones. In the present study, electron tomography has been used to quantitate the mineral distribution in collagen for the first time.

What HVEM tomographic reconstruction methods tell us about biological ultrastructure: Present and future developments

1991 ◽  
Vol 49 ◽  
pp. 436-437
Author(s):  
B. F. McEwen ◽  
J. Frank
Keyword(s):  
Electron Tomography ◽  
Tomographic Reconstruction ◽  
Three Dimensional ◽  
Structural Investigations ◽  
High Voltage Electron Microscopy ◽  
Future Developments ◽  
Voltage Electron ◽  
Reconstruction Methods ◽  
Different Depths ◽  
3D Information

For biological specimens, the primary application of high-voltage electron microscopy (HVEM) has been to enhance three-dimensional (3D) structural investigations. The chief advantage of higher accelerating voltages is that they enable imaging of thicker specimens which are correspondingly richer in 3D information. In order to realize this advantage, however, the specimen must be viewed from more than one direction so that features from different depths superimposed in a single projection may be separated. The simplest application of this approach is stereo viewing, which uses a pair of images viewed from directions separated by a relatively small angle. Due to its simpicity, stereo viewing is widely used but it has the disadvantages of being qualitative, subjective and limited in the amount of 3D information that can be revealed. These limitations can be overcome by electron tomography where several views of the specimen are collected over a wide angular range (up to 180°), entered into a computer, and a 3D image computed from these views according to well-established mathematical principles.

Cryo-Electron Tomography of Isolated Triad Junctions from Skeletal Muscle

2001 ◽  
Vol 7 (S2) ◽  
pp. 94-95 ◽  
Author(s):  
C.-E. Hsieh ◽  
M. Marko ◽  
B.K. Rath ◽  
S. Fleischer ◽  
T. Wagenknecht
Keyword(s):  
Skeletal Muscle ◽  
Plasma Membrane ◽  
Electron Tomography ◽  
Mass Density ◽  
Tomographic Reconstruction ◽  
Three Dimensional ◽  
Cryo Electron Tomography ◽  
Junctional Complexes ◽  
Triad Junction ◽  
Triad Junctions

In skeletal muscle, depolarization of the plasma membrane, which is initiated at the neuromuscular junction, is transduced to a rise in cytoplasmic calcium at specialized structures known as triad junctions (TJs). TJs occur in the myofiber’s interior at regions near the z-lines, where transversely oriented tubular invaginations of the plasma membrane (T-tubules) form junctions with two elements of the sarcoplasmic reticulum (SR). Isolation of membrane fractions that are enriched in junctional complexes and which retain function has been reported.Figure 1 shows a region of an electron micrograph containing an isolated TJ in the frozen-hydrated state. in the orientation shown, two SR-derived vesicles sandwich a flattened vesicle derived from the T-tubule. The junctional regions contain a complex distribution of density, presumably due to proteins that are known to be present in TJs. Electron tomography offers the means to determine the three-dimensional mass density from such micrographs, which would greatly aid in their interpretation. Only recently has the automated data collection technology for determining tomograms of non-stained, frozen-hydrated specimens become available. Here we describe the first tomographic reconstruction of a frozen-hydrated triad junction by automated electron tomography.

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