Array Tomography: A Novel High-Resolution Immunofluorescence Technique

2015 ◽  
pp. 377-388 ◽  
Author(s):  
Mariano Soiza-Reilly
Keyword(s):  
High Resolution ◽  
Immunofluorescence Technique ◽  
Array Tomography

scholarly journals A workflow for streamlined acquisition and correlation of serial regions of interest in array tomography

BMC Biology ◽  
2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Sergio Gabarre ◽  
Frank Vernaillen ◽  
Pieter Baatsen ◽  
Katlijn Vints ◽  
Christopher Cawthorne ◽  
...  
Keyword(s):  
High Resolution ◽  
Light And Electron Microscopy ◽  
User Interaction ◽  
Regions Of Interest ◽  
Imaging Method ◽  
Array Tomography ◽  
Reference Space ◽  
Cell Processes ◽  
Electron Microscopes ◽  
Specific Tissue

Abstract Background Array tomography (AT) is a high-resolution imaging method to resolve fine details at the organelle level and has the advantage that it can provide 3D volumes to show the tissue context. AT can be carried out in a correlative way, combing light and electron microscopy (LM, EM) techniques. However, the correlation between modalities can be a challenge and delineating specific regions of interest in consecutive sections can be time-consuming. Integrated light and electron microscopes (iLEMs) offer the possibility to provide well-correlated images and may pose an ideal solution for correlative AT. Here, we report a workflow to automate navigation between regions of interest. Results We use a targeted approach that allows imaging specific tissue features, like organelles, cell processes, and nuclei at different scales to enable fast, directly correlated in situ AT using an integrated light and electron microscope (iLEM-AT). Our workflow is based on the detection of section boundaries on an initial transmitted light acquisition that serves as a reference space to compensate for changes in shape between sections, and we apply a stepwise refinement of localizations as the magnification increases from LM to EM. With minimal user interaction, this enables autonomous and speedy acquisition of regions containing cells and cellular organelles of interest correlated across different magnifications for LM and EM modalities, providing a more efficient way to obtain 3D images. We provide a proof of concept of our approach and the developed software tools using both Golgi neuronal impregnation staining and fluorescently labeled protein condensates in cells. Conclusions Our method facilitates tracing and reconstructing cellular structures over multiple sections, is targeted at high resolution ILEMs, and can be integrated into existing devices, both commercial and custom-built systems.

scholarly journals Array Tomography: High-Resolution Three-Dimensional Immunofluorescence

2010 ◽  
Vol 2010 (11) ◽  
pp. pdb.top89-pdb.top89 ◽  
Author(s):  
K. D. Micheva ◽  
N. O'Rourke ◽  
B. Busse ◽  
S. J. Smith
Keyword(s):  
High Resolution ◽  
Three Dimensional ◽  
Array Tomography

scholarly journals Niwaki Instead of Random Forests: Targeted Serial Sectioning Scanning Electron Microscopy With Reimaging Capabilities for Exploring Central Nervous System Cell Biology and Pathology

2021 ◽  
Vol 15 ◽  
Author(s):  
Martina Schifferer ◽  
Nicolas Snaidero ◽  
Minou Djannatian ◽  
Martin Kerschensteiner ◽  
Thomas Misgeld
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
High Resolution ◽  
Cell Biology ◽  
Ion Beam ◽  
Serial Sectioning ◽  
Actual Structure ◽  
Array Tomography ◽  
Block Face ◽  
Scanning Electron

Ultrastructural analysis of discrete neurobiological structures by volume scanning electron microscopy (SEM) often constitutes a “needle-in-the-haystack” problem and therefore relies on sophisticated search strategies. The appropriate SEM approach for a given relocation task not only depends on the desired final image quality but also on the complexity and required accuracy of the screening process. Block-face SEM techniques like Focused Ion Beam or serial block-face SEM are “one-shot” imaging runs by nature and, thus, require precise relocation prior to acquisition. In contrast, “multi-shot” approaches conserve the sectioned tissue through the collection of serial sections onto solid support and allow reimaging. These tissue libraries generated by Array Tomography or Automated Tape Collecting Ultramicrotomy can be screened at low resolution to target high resolution SEM. This is particularly useful if a structure of interest is rare or has been predetermined by correlated light microscopy, which can assign molecular, dynamic and functional information to an ultrastructure. As such approaches require bridging mm to nm scales, they rely on tissue trimming at different stages of sample processing. Relocation is facilitated by endogenous or exogenous landmarks that are visible by several imaging modalities, combined with appropriate registration strategies that allow overlaying images of various sources. Here, we discuss the opportunities of using multi-shot serial sectioning SEM approaches, as well as suitable trimming and registration techniques, to slim down the high-resolution imaging volume to the actual structure of interest and hence facilitate ambitious targeted volume SEM projects.

scholarly journals A Computational Synaptic Antibody Characterization and Screening Framework for Array Tomography

2018 ◽  
Author(s):  
Anish K. Simhal ◽  
Belvin Gong ◽  
James S. Trimmer ◽  
Richard J. Weinberg ◽  
Stephen J. Smith ◽  
...  
Keyword(s):  
High Resolution ◽  
Sensitivity And Specificity ◽  
Screening Tool ◽  
Molecular Composition ◽  
Target Specificity ◽  
Efficient Tool ◽  
Antibody Screening ◽  
Array Tomography ◽  
Synapse Density ◽  
Application Specific

ABSTRACTApplication-specific validation of antibodies is a critical prerequisite for their successful use. Here we introduce an automated framework for characterization and screening of antibodies against synaptic molecules for high-resolution immunofluorescence array tomography (AT). The proposed Synaptic Antibody Screening Tool (SACT), is designed to provide an automatic, robust, flexible, and efficient tool for antibody characterization at scale. By allowing the user to define the molecular composition and size of synapses expected to contain the antigen, the method detects and characterizes puncta and synapses, and outputs automatically computed characteristics such as synapse density and target specificity ratio, which reflect the sensitivity and specificity of immunolabeling with a given antibody. These measurements provide an objective way to characterize and compare the performance of different antibodies against the same target, and can be used to objectively select the antibodies best suited for AT and potentially for other immunolabeling applications.

Observations of the spatial structure of interstellar hydrogen

1967 ◽  
Vol 31 ◽  
pp. 45-46
Author(s):  
Carl Heiles
Keyword(s):  
High Resolution ◽  
Spatial Structure ◽  
Velocity Dispersion ◽  
Temperature Variations

High-resolution 21-cm line observations in a region aroundlII= 120°,b11= +15°, have revealed four types of structure in the interstellar hydrogen: a smooth background, large sheets of density 2 atoms cm-3, clouds occurring mostly in groups, and ‘Cloudlets’ of a few solar masses and a few parsecs in size; the velocity dispersion in the Cloudlets is only 1 km/sec. Strong temperature variations in the gas are in evidence.

Combining integrated systems-biology approaches with intervention-based experimental design provides a higher-resolution path forward for microbiome research

2019 ◽  
Vol 42 ◽  
Author(s):  
J. Alfredo Blakeley-Ruiz ◽  
Carlee S. McClintock ◽  
Ralph Lydic ◽  
Helen A. Baghdoyan ◽  
James J. Choo ◽  
...  
Keyword(s):  
Systems Biology ◽  
High Resolution ◽  
Experimental Design ◽  
Integrated Systems ◽  
Microbiome Research ◽  
Constructive Criticism

Abstract The Hooks et al. review of microbiota-gut-brain (MGB) literature provides a constructive criticism of the general approaches encompassing MGB research. This commentary extends their review by: (a) highlighting capabilities of advanced systems-biology “-omics” techniques for microbiome research and (b) recommending that combining these high-resolution techniques with intervention-based experimental design may be the path forward for future MGB research.

Very High Resolution Analysis of the Dynamics of a Coronal Plasmoid

1994 ◽  
Vol 144 ◽  
pp. 593-596
Author(s):  
O. Bouchard ◽  
S. Koutchmy ◽  
L. November ◽  
J.-C. Vial ◽  
J. B. Zirker
Keyword(s):  
High Resolution ◽  
Solar Eclipse ◽  
Total Solar Eclipse ◽  
Field Of View ◽  
Small Field ◽  
High Resolution Analysis ◽  
Ccd Observations ◽  
Resolution Analysis ◽  
Very High ◽  
Small Field Of View

AbstractWe present the results of the analysis of a movie taken over a small field of view in the intermediate corona at a spatial resolution of 0.5“, a temporal resolution of 1 s and a spectral passband of 7 nm. These CCD observations were made at the prime focus of the 3.6 m aperture CFHT telescope during the 1991 total solar eclipse.

FeXIV Line Emission Polarization of the July 11, 1991 Solar Corona

1994 ◽  
Vol 144 ◽  
pp. 541-547
Author(s):  
J. Sýkora ◽  
J. Rybák ◽  
P. Ambrož
Keyword(s):  
High Resolution ◽  
Solar Corona ◽  
Emission Line ◽  
Solar Eclipse ◽  
White Light ◽  
Preliminary Analysis ◽  
Line Emission ◽  
Total Solar Eclipse ◽  
Degree Of Polarization ◽  
High Resolution Images

AbstractHigh resolution images, obtained during July 11, 1991 total solar eclipse, allowed us to estimate the degree of solar corona polarization in the light of FeXIV 530.3 nm emission line and in the white light, as well. Very preliminary analysis reveals remarkable differences in the degree of polarization for both sets of data, particularly as for level of polarization and its distribution around the Sun’s limb.

Microcalorimeters for X-Ray Spectroscopy

1988 ◽  
Vol 102 ◽  
pp. 41
Author(s):  
E. Silver ◽  
C. Hailey ◽  
S. Labov ◽  
N. Madden ◽  
D. Landis ◽  
...  
Keyword(s):  
High Resolution ◽  
High Efficiency ◽  
Thermal Response ◽  
Low Noise ◽  
High Resolution Spectroscopy ◽  
Superconducting Transition ◽  
Sapphire Substrates ◽  
Laboratory Plasmas ◽  
Adiabatic Demagnetization ◽  
Theoretical Predictions

The merits of microcalorimetry below 1°K for high resolution spectroscopy has become widely recognized on theoretical grounds. By combining the high efficiency, broadband spectral sensitivity of traditional photoelectric detectors with the high resolution capabilities characteristic of dispersive spectrometers, the microcalorimeter could potentially revolutionize spectroscopic measurements of astrophysical and laboratory plasmas. In actuality, however, the performance of prototype instruments has fallen short of theoretical predictions and practical detectors are still unavailable for use as laboratory and space-based instruments. These issues are currently being addressed by the new collaborative initiative between LLNL, LBL, U.C.I., U.C.B., and U.C.D.. Microcalorimeters of various types are being developed and tested at temperatures of 1.4, 0.3, and 0.1°K. These include monolithic devices made from NTD Germanium and composite configurations using sapphire substrates with temperature sensors fabricated from NTD Germanium, evaporative films of Germanium-Gold alloy, or material with superconducting transition edges. A new approache to low noise pulse counting electronics has been developed that allows the ultimate speed of the device to be determined solely by the detector thermal response and geometry. Our laboratory studies of the thermal and resistive properties of these and other candidate materials should enable us to characterize the pulse shape and subsequently predict the ultimate performance. We are building a compact adiabatic demagnetization refrigerator for conveniently reaching 0.1°K in the laboratory and for use in future satellite-borne missions. A description of this instrument together with results from our most recent experiments will be presented.

Radiation Damage, Contrast and Statistics in High Resolution Biological Electron Microscopy

1970 ◽  
Vol 28 ◽  
pp. 260-261
Author(s):  
Robert M. Glaeser
Keyword(s):  
High Resolution ◽  
Particle Flux ◽  
Unit Area ◽  
Signal To Noise ◽  
Resolution Image ◽  
High Resolution Image ◽  
Pass Through ◽  
Counting Statistics ◽  
The Relationship ◽  
Picture Element

It is well known that a large flux of electrons must pass through a specimen in order to obtain a high resolution image while a smaller particle flux is satisfactory for a low resolution image. The minimum particle flux that is required depends upon the contrast in the image and the signal-to-noise (S/N) ratio at which the data are considered acceptable. For a given S/N associated with statistical fluxtuations, the relationship between contrast and “counting statistics” is s131_eqn1, where C = contrast; r2 is the area of a picture element corresponding to the resolution, r; N is the number of electrons incident per unit area of the specimen; f is the fraction of electrons that contribute to formation of the image, relative to the total number of electrons incident upon the object.

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