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

2016 ◽  
Author(s):  
Julian Ng ◽  
Alyssa Browning ◽  
Lorenz Lechner ◽  
Masako Terada ◽  
Gillian Howard ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Neural Circuit ◽  
Cell Types ◽  
Cellular Tissue ◽  
Computer Assisted ◽  
X Ray ◽  
X Ray Imaging ◽  
Resolution Imaging ◽  
Volume Acquisition ◽  
Non Destructive

AbstractLarge 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 their 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 enables us to target specific regions for EM volume acquisition.

scholarly journals Alterations in Sub-Axonal Architecture Between Normal Aging and Parkinson’s Diseased Human Brains Using Label-Free Cryogenic X-ray Nanotomography

2020 ◽  
Vol 14 ◽  
Author(s):  
Hung Tri Tran ◽  
Esther H. R. Tsai ◽  
Amanda J. Lewis ◽  
Tim Moors ◽  
J. G. J. M. Bol ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Human Brain ◽  
Brain Tissue ◽  
Multispectral Imaging ◽  
Alpha Synuclein ◽  
Label Free ◽  
Myelinated Axons ◽  
X Ray ◽  
X Ray Imaging ◽  
Non Destructive

Gaining insight to pathologically relevant processes in continuous volumes of unstained brain tissue is important for a better understanding of neurological diseases. Many pathological processes in neurodegenerative disorders affect myelinated axons, which are a critical part of the neuronal circuitry. Cryo ptychographic X-ray computed tomography in the multi-keV energy range is an emerging technology providing phase contrast at high sensitivity, allowing label-free and non-destructive three dimensional imaging of large continuous volumes of tissue, currently spanning up to 400,000 μm3. This aspect makes the technique especially attractive for imaging complex biological material, especially neuronal tissues, in combination with downstream optical or electron microscopy techniques. A further advantage is that dehydration, additional contrast staining, and destructive sectioning/milling are not required for imaging. We have developed a pipeline for cryo ptychographic X-ray tomography of relatively large, hydrated and unstained biological tissue volumes beyond what is typical for the X-ray imaging, using human brain tissue and combining the technique with complementary methods. We present four imaged volumes of a Parkinson’s diseased human brain and five volumes from a non-diseased control human brain using cryo ptychographic X-ray tomography. In both cases, we distinguish neuromelanin-containing neurons, lipid and melanic pigment, blood vessels and red blood cells, and nuclei of other brain cells. In the diseased sample, we observed several swellings containing dense granular material resembling clustered vesicles between the myelin sheaths arising from the cytoplasm of the parent oligodendrocyte, rather than the axoplasm. We further investigated the pathological relevance of such swollen axons in adjacent tissue sections by immunofluorescence microscopy for phosphorylated alpha-synuclein combined with multispectral imaging. Since cryo ptychographic X-ray tomography is non-destructive, the large dataset volumes were used to guide further investigation of such swollen axons by correlative electron microscopy and immunogold labeling post X-ray imaging, a possibility demonstrated for the first time. Interestingly, we find that protein antigenicity and ultrastructure of the tissue are preserved after the X-ray measurement. As many pathological processes in neurodegeneration affect myelinated axons, our work sets an unprecedented foundation for studies addressing axonal integrity and disease-related changes in unstained brain tissues.

Defects of Platelet Function Recognized by X-Ray Imaging and Scanning Electron Microscopy

1985 ◽  
Vol 43 ◽  
pp. 610-613
Author(s):  
M.G. Baldini ◽  
S. Morinaga ◽  
D. Minasian ◽  
R. Feder ◽  
D. Sayre ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Cell Biology ◽  
Imaging Technique ◽  
Human Platelets ◽  
X Rays ◽  
X Ray ◽  
X Ray Imaging ◽  
Complex Phenomena ◽  
Scanning Electron

Contact X-ray imaging is presently developing as an important imaging technique in cell biology. Our recent studies on human platelets have demonstrated that the cytoskeleton of these cells contains photondense structures which can preferentially be imaged by soft X-ray imaging. Our present research has dealt with platelet activation, i.e., the complex phenomena which precede platelet appregation and are associated with profound changes in platelet cytoskeleton. Human platelets suspended in plasma were used. Whole cell mounts were fixed and dehydrated, then exposed to a stationary source of soft X-rays as previously described. Developed replicas and respective grids were studied by scanning electron microscopy (SEM).

X-Ray Nanotomography (XRMT) Tool for Non-Destructive High-Resolution Imaging of ICs

2001 ◽  
Author(s):  
Wenbing Yun ◽  
Steve Wang ◽  
David Scott ◽  
Kenneth W. Nill ◽  
Waleed S. Haddad
Keyword(s):  
High Resolution ◽  
Process Development ◽  
3D Visualization ◽  
Tomographic Reconstruction ◽  
Reconstruction Algorithms ◽  
X Ray ◽  
Visualization Software ◽  
Volumetric Reconstruction ◽  
Resolution Imaging ◽  
Non Destructive

Abstract A high-resolution table-sized x-ray nanotomography (XRMT) tool has been constructed that shows the promise of nondestructively imaging the internal structure of a full IC stack with a spatial resolution better than 100 nm. Such a tool can be used to detect, localize, and characterize buried defects in the IC. By collecting a set of X-ray projections through the full IC (which may include tens of micrometers of silicon substrate and several layers of Cu interconnects) and applying tomographic reconstruction algorithms to these projections, a 3D volumetric reconstruction can be obtained, and analyzed for defects using 3D visualization software. XRMT is a powerful technique that will find use in failure analysis and IC process development, and may facilitate or supplant investigations using SEM, TEM, and FIB tools, which generally require destructive sample preparation and a vacuum environment.

Three Dimensional X-Ray Computed Tomography in Materials Science

MRS Bulletin ◽  
1988 ◽  
Vol 13 (1) ◽  
pp. 13-18 ◽  
Author(s):  
J.H. Kinney ◽  
Q.C. Johnson ◽  
U. Bonse ◽  
M.C. Nichols ◽  
R.A. Saroyan ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Sample Preparation ◽  
Visible Light ◽  
Three Dimensional ◽  
Materials Characterization ◽  
Imaging Technology ◽  
X Ray ◽  
X Ray Imaging ◽  
Visible Light Imaging

Imaging is the cornerstone of materials characterization. Until the middle of the present century, visible light imaging provided much of the information about materials. Though visible light imaging still plays an extremely important role in characterization, relatively low spatial resolution and lack of chemical sensitivity and specificity limit its usefulness.The discovery of x-rays and electrons led to a major advance in imaging technology. X-ray diffraction and electron microscopy allowed us to characterize the atomic structure of materials. Many materials vital to our high technology economy and defense owe their existence to the understanding of materials structure brought about with these high-resolution methods.Electron microscopy is an essential tool for materials characterization. Unfortunately, electron imaging is always destructive due to the sample preparation that must be done prior to imaging. Furthermore, electron microscopy only provides information about the surface of a sample. Three dimensional information, of great interest in characterizing many new materials, can be obtained only by time consuming sectioning of an object.The development of intense synchrotron light sources in addition to the improvements in solid state imaging technology is revolutionizing materials characterization. High resolution x-ray imaging is a potentially valuable tool for materials characterization. The large depth of x-ray penetration, as well as the sensitivity of absorption crosssections to atomic chemistry, allows x-ray imaging to characterize the chemistry of internal structures in macroscopic objects with little sample preparation. X-ray imaging complements other imaging modalities, such as electron microscopy, in that it can be performed nondestructively on metals and insulators alike.

Rapid, Non-destructive Inspection and Classification of Inhalation Blisters Using Low-Energy X-ray Imaging

2018 ◽  
Vol 13 (3) ◽  
pp. 270-282 ◽  
Author(s):  
Nagaraja Rao ◽  
Brian Ament ◽  
Richard Parmee ◽  
Jonathan Cameron ◽  
Martin Mayo
Keyword(s):  
Low Energy ◽  
X Ray ◽  
X Ray Imaging ◽  
Non Destructive

On-Site Non-Destructive Inspection of Bridges Using the 950 keV X-Band Electron Linac X-ray Source

2017 ◽  
Vol 12 (3) ◽  
pp. 578-584 ◽  
Author(s):  
Mitsuru Uesaka ◽  
◽  
Yuki Mitsuya ◽  
Eiko Hashimoto ◽  
Katsuhiro Dobashi ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Bridge Inspection ◽  
Electron Linac ◽  
X Ray ◽  
Band Electron ◽  
Reconstruction Methods ◽  
X Ray Imaging ◽  
X Band ◽  
Near Future ◽  
Non Destructive

Regular diagnosis of the structural health of infrastructure, such as bridges, is indispensable to ensure safety and reliable operation of the society. Non-destructive tests based on X-ray imaging are powerful tools to inspect the inside of a concrete structure in detail. Establishing a diagnostic method of bridges based on X-ray visualization is required to examine the internal conditions and helps in the rationalization of maintenances. We demonstrated our 950 keV X-band electron linac based X-ray source for on-site actual bridge inspection and visualized the inner structure of a bottom floor slab. For the more precise inspection of the conditions of wires and rods, we applied three-dimensional image reconstruction methods for bridge mock-up samples. Partial angle computed tomography and tomosynthesis gave cross section images of the samples with 1 mm resolution. We are planning to investigate another part of the bridge, with the 950 keV X-ray system in the near future.

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