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97 protocols using vgstudio max 2

1

Petrographic Analysis of Thin-Sections

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The thin-sections were produced following standard petrographic methods (Klein and Sander)50 and then studied and photographed with a Leica® DM 750 P compound polarizing microscope equipped with a digital Leica® ICC50HD camera. Histological terminology follows Francillon-Vieillot et al.51 . Some samples were micro-CT-scanned with a v|tome|xs by GE phoenix|x-ray at the Steinmann Institut für Geologie, Mineralogie und Paläontologie (StIPB) in Bonn (Germany). Image visualization was performed using VGStudio MAX 2.0 software (Volume Graphics GmbH) and Adobe Photoshop. Cross-sections of samples were transformed into black (bone) and white (cavities and vascular spaces) images to measure bone compactness with a custom-designed pixel-counting computer program developed by P. Göddertz (StIPB).
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2

In Vivo Micro-CT Imaging of Murine Liver Vasculature

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Micro-CT (LCT-200 scanner, Hitachi Aloka Medical Ltd., Tokyo, Japan) was performed at 55 kVp, with an anode current of 500 µA and a shutter speed of 500 ms. Scans were completed over 360 degrees of rotation of the x-ray tube with 450 projections to reduce signal-to-noise. Reconstructions were performed using a cone-beam filtered back projection algorithm. The axial field of view was set to 4.6 cm with an inplane spatial resolution of 91 µm. The mouse was anesthetized with isoflurane/oxygen general anesthesia systems. The tail veins of the mice were cannulated, and approximately 0.3 mL of the contrast agent Fenestra Liver Contrast (Advanced Research Technologies Inc., Saint-Laurent, QC, CA) was injected (0.013 mL of Fenestra LC per gram of body weight of animal). The total scan time for each micro-CT scan was approximately 5-10 minutes. Image acquisition started 5 and 30 minutes after contrast agent injection. Micro-CT image data were acquired reconstructed and analyzed using VGS tudio MAX 2.0 software (Volume Graphics GmbH, Heidelberg, Germany).
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3

Quantifying Corrosion Dynamics via XCT

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The 32-bit XCT datasets were reconstructed with VGSTUDIO MAX (VGStudio MAX 2.0.5, Volume Graphics, Germany) and rigidly registered with a correlative metric in Avizo (Avizo 9.7, ThermoFisher Scientific, US) using the first preloaded image as a reference. Each image was cropped to include only the specimen structures in the field of view (∼6 mm3) and converted to binary images using Otsu's method [62 (link)].
The corrosion rate (CR) was estimated using the binary 3D tomograms and applying equation (1): CR=ΔVAΔt where A was the initial total surface area of the fibres exposed to corrosion (533 ± 58 mm2) computed from the 3D XCT images before corrosion using the BoneJ [63 (link)] module of Fiji [58 (link)], Δt the corrosion time (i.e. 2, 8 or 14 days) and ΔV the reduction in volume, equal to the difference between the initial pre-corroded volume the remaining volume. Corrosion maps were obtained by subtracting the registered binary image of pre-corroded samples by the corresponding binary image of corroded samples before loading, resulting in the quantification of lost material volume.
Morphometric properties, such as pore size and solid volume fraction, were computed and corresponding 3D maps were produced using the pore network and volume fraction map modules of Avizo.
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4

X-ray Nanotomography of Biological Specimens

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Specimens were transferred from FAE to an ascending series of ethanol (70%-80%-90%-95%-100%), stained in iodine solution, transferred to acetone, and then dried at the critical point (Emitech K850, Quorum Technologies Ltd., Ashford, UK). One dried specimen was scanned at the MPI for the Science of Human History (Jena, Germany) with a SkyScan 2211 X-ray nanotomograph (Bruker, Knotich, Belgium) with an image spatial resolution of 0.68 μm (isotropic voxel size) using the following parameters: 70 kV, 300 μA, 3600 ms exposure time, 0.15°rotation steps, frame averaging on (3), and using no filter. Projections were reconstructed by NRecon (Bruker, Knotich, Belgium) into JPG files. The μCT-scan is stored in the collection of the Phyletisches Museum Jena. Amira 6.1.1 (Thermo Fisher Scientific, Waltham, USA) and VG studio Max 2.0.5 (Volume Graphics, Heidelberg, Germany) were used for the three-dimensional reconstruction and volume rendering.
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5

Micro-CT Imaging of Didinium Specimen

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Specimen DIP-S-0907 was scanned with a MicroXCT 400 (Carl Zeiss X-ray Microscopy Inc.) at the Institute of Zoology, Chinese Academy of Sciences. The entire animal (Fig. 1) was divided into seven scans that were combined to create a single model, and the scans were conducted with a beam strength of 60 kV, 8 W, and absorption contrast and a spatial resolution of 2.5464 μm. In addition, specimen DIP-S-0907 was imaged using propagation phase-contrast synchrotron radiation microtomography on the beamline 13W at the Shanghai Synchrotron Radiation Facility. The isotropic voxel size was 2.25 μm.
On the basis of the obtained image stacks, structures of the specimen were reconstructed and separated with Amira 5.4 (Visage Imaging). The subsequent volume rendering was performed with Avizo 9.0 (Thermo Fisher Scientific) and VG Studiomax 2.1 (Volume Graphics). The neonate C. ruffus was loaned from the Western Australian Museum (WAM R49553) and scanned with a SkyScan 1076 (Bruker MicroCT) at Adelaide Microscopy, University of Adelaide, Australia. The scan settings were 65 kV, 153 μA, no filter, and an isotropic voxel size of 8.7 μm. The reconstruction was carried out using the software NRecon (Bruker MicroCT), and the volume renderings were created in the software Avizo 9.0 (Thermo Fisher Scientific).
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6

Microstructural Analysis of Murine Bone Specimens

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Femurs and tibias were scanned after fixation in paraformaldehyde. The μCT scans were performed using a nanotom® m (phoenix|x-ray, GE Sensing & Inspection Technologies GmbH, Wunstorf, Germany) equipped with a 180 kV/15 W nanofocus X-ray source. A tungsten transmission target, an accelerating voltage of 70 kV, and a beam current of 260 μA were used. To increase mean energy of the photon spectrum and consequently reduce beam hardening artefacts, a 0.5 mm aluminium filter was inserted between source and specimen. A region of air was designated in all scans as suggested by the software (GE Sensing & Inspection Technologies GmbH) to standardize the grey-scale for data interpretation. 1440 equiangular projection images were acquired over 360° with an exposure time of 1 second. The radiographs were reconstructed using a cone beam filtered back-projection algorithm with the manufacturer's software phoenix datos|x 2.0.1 RTM (GE Sensing & Inspection Technologies GmbH). Whole bones were scanned and reconstructed with a voxel size of 18.5 μm. Datasets were visualised using VG Studio MAX 2.1 (Volume Graphics GmbH, Heidelberg, Germany) and analysed using ImageJ with the BoneJ and 3D Shape plugins [20 (link)–22 (link)].
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7

Volumetric Visualization of 3D CT Scans

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CT reconstruction was done using TomoPy, an open-source package from Argonne National Labs (http://tomopy.readthedocs.io). Some of the image processing was conducted in Fiji/ImageJ2 (https://fiji.sc/). Software used for volumetric visualizations in this manuscript include Avizo version 9.4 (Thermo Fisher Scientific, Waltham, MA) and VGStudio Max 2.1 (Volume Graphics, Heidelberg, Germany). Videos 1 and 4 were generated in Avizo. Videos 2, 3, 5 and 6 were generated in VGStudio Max 2.1 using an intensity histogram adjusted to better discern otherwise faint or overlapping structures.
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8

Micro-CT Analysis of Bone Samples

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Samples were scanned in air using the XT H 225 micro-CT machine (Nikon Metrology Inc., Brighton, MI, USA) at an isotropic voxel size of 6.7 μm. The scan settings were 90 kV, 90 μA, 720 projections, 2 frames per projection, and an integration time of 708 ms. 3D reconstructions were done using the software CT Pro 3D (Nikon metrology, Inc., Brighton, MI, USA). BMP datasets for each scan were made using VG Studio MAX 2.1 (Volume Graphics GmbH, Heidelberg, Germany). Morphometric analysis was completed with the SkyScan CT-Analyser (CTAn) software (Bruker micro-CT, Belgium) according to Bruker micro-CT’s Method Note 2. Method Note 8 was used to select cortical and trabecular regions of interest in an automated manner. All 3D models were created with the CT-Volume (CTVol) software (Bruker micro-CT, Belgium). All micro-CT images presented are at the same magnification.
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9

3D Microstructure Reconstruction from MicroCT

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2D longitudinal projection images were directly obtained from the synchrotron microCT scanning system and used for reconstruction of the 3D volume. We have applied VGStudio Max 2.1 (Volume Graphics, Heidelberg, Germany) to the microCT data for 3D visualizations. It handles large data in raw, tiff and other formats.
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10

Mandibular Tooth Enamel Mineralization Analysis

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Mandibles were dissected from 3-week-old (when teeth may be not affected by mechanical stresses) WT and cKO mice (n = 3 per genotype), symmetrically cut in half, fixed in 4% paraformaldehyde (PFA; pH 7.2), cleaned of soft tissue, washed in phosphate-buffered saline (PBS) for at least 24 h, transferred into 70% ethanol, dehydrated in an ascending series of alcohol, and then dried naturally in air. Morphological analysis and measurement of enamel mineralization of the mandibular teeth were performed using a µCT system (µCT 50; Scanco Medical AG, Bassersdorf, Switzerland), with a nominal isotropic resolution of 5 µm inside a 19-mm diameter scanning vial under conditions of 70 kVp, 114 mA, 8 W, 500 ms integration time and 500 projections per 180°. Threshold values in the µCT evaluation program (version 6.5-2, Scanco, Wayne, PA, USA) were set such that the mineralized enamel appeared as a white high-density solid next to dentin as a transparent object. Three-dimensional images were reconstructed using VGStudio MAX 2.1 software (Volume Graphics, Heidelberg, Germany).
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