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Display image description
Single computed slice through a tomographic reconstruction of a Node of Ranvier from rat peripheral nerve, prepared by a combination of chemical fixation, high pressure freezing and freeze substitution
Full resolution image description
Zip file contains the volume (yinghpf_good.img/hdr) in Analyze 7.5 format.
File format
1792, 2671, 250
Volume scale
0.0014, 0.0014, 0.0014
Animation description
Annotated movie through the computed slices of a tomographic reconstruction of a Node of Ranvier from rat peripheral nerve, along with surface renderings of structures segmented from the reconstruction. See the Segmentation section for a details about the structures and the segmentation process.

Image 2D
Display image description
Single image taken on an intermediate voltage electron microscope at zero degrees tilt from a single axis tilt series through the Node of Ranvier from rat peripheral nerve prepared by chemical fixation followed by high pressure freezing and free substitution. Image is shown in negative contrast. Bright spots are colloidal gold particles applied to the section surface to serve as fiducial markers for subsequent alignment.
Full resolution image description
Original tilt images in suprim format (*.f). Also contains fid, preali, rawtlt, st, suprim files needed for alignment. According to the scanning dimensions given in the paper, the images (hpf*.f) submitted represent cropped and aligned images used for the reconstruction. The raw tilt images are contained in the .st file as a multifile image stack.

Display image description
Manual segmentation of paranodal loops, microtubules, neurofilaments, sepjate junctions and cross bridges using Xvoxtrace v 2.9. Different parts of the myelin sheath were segmented as different objects. Objects were surfaced using both Synu and Amira
Segmentation file description
Tar archive containing the trace file hpfnode_backup.trace and the segmented objects in synu (*.synu) and Amira (*.inventor) format.

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CCDB:3696*  Cite 
Project: P1704
Project name
Analysis of the Cross Bridges in the Paranodal Region of the Node of Ranvier
Electron tomography was used to examine cytoskeletal-cytoskeletal, membrane-cytoskeletal, and heterologous cell connections in the paranodal region of the node of Ranvier in peripheral nerves.
Funding agency
National Institutes of Health
Mark Ellisman
Gina Sosinsky
Guy Perkins
Sassan Ghassemzadeh
Alex Perez
Ying Jones
Start date
End date
Experiment ID
High resolution tomogram of paranodal area, particularly septate junction
Structure, localization of components
Gina Sosinsky
Ying Jones
Microscopy product
Microscopy product ID
JEOL 4000
Microscopy type
Product type
Image basename
Scientific name
rattus norvegicus
Sprague Dawley
1 months
Age class
young adult
Tissue section
Anatomical location
Spinal Root
Leica Ultracut UCT
0.5 µm
Specimen description
spinal root
peripheral nervous system
Node of Ranvier
Cell type
Schwann cell
Imaging parameters
Electron microscopy product
Recording medium
Accelerating voltage
400 kv
Specimen preparation
Protocol used
Rats were first anaesthetized by intraperitoneal injection of Nembutal (35 mg per kg body weight), and whole blood was flushed out with oxygenated rat Ringers solution at 37 C. They were then fixed by intracardial perfusion with an aldehyde mixture composed of 2 percent paraformaldehyde and 2.5 percent glutaraldehyde in 0.15 M cacodylate buffer (pH 7.4) at 37 C. The spinal root was dissected out, immersed in the same fixative and transferred to the high-pressure freezing (HPF) unit. These samples were trimmed and blotted (to remove excess water) before placing in the carrier. Samples were loaded into the carriers of the Bal-Tec HPM010 HPF unit (Bal-Tec, Liechtenstein) and operated according to manufacturers instructions. Carriers with a depth of 0.2 mm were used with thin bundles of spinal root to provide uniform ultrastructural preservation. The inert liquid, hexadecene, was added to the carrier to fill in the well, excluding air from surrounding the sample.Freeze Substitution: After the samples were high-pressure frozen, they were freeze-substituted in dry acetone containing 0.1 percent tannic acid for 24 h at negative 90 degrees C in a Leica EMAFS freeze substitution unit (FSU, Leica Microsystems, Bannockburn, IL), then in acetone containing 2 percent OsO4 and 0.1 percent of uranyl acetate for the remaining time of the procedure. Both tannic acid and uranyl acetate were added not only to help stain the material but also to aid in membrane preservation. During this procedure, the water in the sample is completely extracted and replaced by the fixative, i.e., freeze-substituted. The temperature for substitution was maintained at minus 90 C for 3 days to ensure that the biological structures were cryo-immobilized during this substitution. The temperature was raised gradually at a rate of 2 degrees C per hour to minus 60 degrees C and held there for 8 h because OsO4 does not start to cross-link until around minus 70 degrees C. The samples were then warmed to minus 20 degrees C at a rate of 5 degrees C per hour and held at this temperature for 12 h. The carriers were then taken from the FSU, the tissue removed from the carriers and quickly washed several times in dry acetone at room temperature (RT) to remove the OsO4 and uranyl acetate. Acetone above 70 percent can extract lipids at RT. Hence, the exposure time to it was minimized. The tissues were then incubated in a 50:50 mixture of acetone/Durcupan for 4 h at RT followed by 100 percent Durcupan resin of 24 h infiltration with three changes. Resin polymerization was accomplished in a vacuum oven at 65 to 70 degrees C for 48 h. The resin-embedded samples were removed from the oven and sections were cut using a Leica ultramicrotome. Sections were then stained 30 min in 2 percent aqueous uranyl acetate, followed by 15 min in lead salts. Fiducial cues consisting of 15 and 20 nm colloidal gold particles were deposited on opposite sides of the section.
Imaging product type
Single tilt
Min range
-60 degrees
Max range
60 degrees
Tilt increment
2 degrees