Hematoxylin and eosin stained paraffin slide preparation and scanning with a NanoZoomer Digital Pathology System (Hamamatsu) instrument were previously described (Weisner et al., 2019 (link)). For cerebellum, two sections from each cerebellar vermis and hemisphere region were chosen to measure the thickness of the external granular layer (EGL) and granule cell layer (GL) in each lobule and fissure using the “Ruler” function in NanoZoomer Digital Pathology Software, NDP.view 2. To quantitate the number of Bergmann glia (BG), two 50 μm lines were randomly drawn parallel to each fissure on glial fibrillary acidic protein (GFAP) stained cerebellar confocal images, and the number of intersects of each 50 μm line and GFAP+ BG processes was quantitated. A TUNEL assay was performed using the DeadEnd Colorimetric TUNEL System (Promega, G7360) to measure the number of dead cells, and stained paraffin slides were scanned by a NanoZoomer Digital Pathology System (Hamamatsu).
Nanozoomer digital pathology system
Manufactured by Hamamatsu Photonics
Sourced in Japan, United Kingdom, Germany
The NanoZoomer Digital Pathology System is a high-performance whole slide imaging platform designed for digitizing glass slides. It captures high-resolution images of tissue samples, enabling efficient digital storage, analysis, and remote collaboration in pathology workflows.
Automatically generated - may contain errors
Lab products found in correlation
Deadend fluorometric tunel system, by Promega (4 mentions)
Tissue studio, by Definiens (4 mentions)
Nanozoomer digital pathology image program, by Hamamatsu Photonics (3 mentions)
Dab chromogen, by Agilent Technologies (3 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (3 mentions)
Picrosirius red staining kit, by Polysciences (2 mentions)
St5020 autostainer, by Leica (2 mentions)
Image pro plus software, by Media Cybernetics (2 mentions)
Buffered formalin, by Merck Group (2 mentions)
Lsm t pmt confocal microscope, by Zeiss (2 mentions)
Axio imager z2 microscope, by Zeiss (1 mentions)
Discovery xt, by Roche (1 mentions)
Anti human gal 7 polyclonal antibody, by R&D Systems (1 mentions)
Fv1000mpe, by Olympus (1 mentions)
Rm2255 microtome, by Leica (1 mentions)
Aniline blue solution, by Thermo Fisher Scientific (1 mentions)
Ndp view software, by Hamamatsu Photonics (1 mentions)
At180, by Thermo Fisher Scientific (1 mentions)
At100, by Thermo Fisher Scientific (1 mentions)
Ps214, by Abcam (1 mentions)
91 protocols using nanozoomer digital pathology system
1
Cerebellar Layer Thickness and Cell Death Analysis
2
Histological and Immunohistochemical Analysis of Tuberculosis
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Segments of lung tissue isolated from mice infected with M. tuberculosis were fixed in 10% buffered formalin (Sigma-Aldrich) and embedded in paraffin. Histologic sections were stained with hematoxylin and eosin (H&E) for pathological evaluation. Images of the whole microscope slide were captured using a NanoZoomer digital pathology system (Hamamatsu Photonics). For immunohistochemistry, lung tissue from human TB patients who received surgery were collected and frozen in liquid nitrogen. Serial 5-mm sections of paraffin-embedded tissue were used for immunohistochemical staining. Briefly, tissues were fixed with acetone-chloroform for 3 min and then incubated for 2 h with anti-CD157 (ab137718; Abcam). Primary antibodies were detected using a biotinylated secondary antibody system (PolinkDS-MR kit; Golden Bridge International Co.) following the manufacturer’s instructions. Images were captured using a NanoZoomer digital pathology system (Hamamatsu Photonics).
3
Histological and Immunostaining Analysis of Testes
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Testes were fixed in Bouin's solution for hematoxylin–eosin (HE) or periodic acid–Schiff (PAS) staining for 5 h (juvenile testes) or overnight at RT, or in 4% paraformaldehyde (for immunostaining) at RT. Testes were then embedded in paraffin and cut in 3 μm slices. HE- and PAS-stained sections were scanned using the automated tissue slide scanning tool of a Hamamatsu NanoZoomer Digital Pathology system. TUNEL assays were performed with the DeadEnd Fluorometric TUNEL System (Promega), according to the manufacturer's protocol. For immunostaining, sections were deparaffinized, washed and permeabilized in 0.1% Triton X-100/PBS. Sections were then blocked in blocking buffer (10% donkey serum, 0.05% Tween-20, 0.05% Triton X-100 in PBS) for 30 min at RT and incubated with the primary antibodies (anti-SKAP, 1:100; anti-N-ter SKAP, 1:2000; anti-TRA98, 1:100 and anti-SOX9, 1:100 in blocking buffer) at RT overnight. Secondary antibodies were also diluted in blocking buffer and added at 37 °C for 1 h. Images were acquired with a sCMOS ORCA Flash 4.2 camera (Hamamatsu), an Axioimager Z2 microscope (Carl Zeiss), a 63× PL APO 1.4 lens, DAPI (FS49), FITC (FS38 HE), Texas Red (FS45), Cy3 (FS43) and Cy5 (FS50) fluorescence filter sets.
4
Semiquantitative USP19 Immunohistochemistry in Renal Cancer
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Immunohistochemistry staining for USP19 was performed on the paraffin-embedded kidney clear cell carcinoma tissue array (KD481, KD601; BioMax), followed by secondary antibody and DAB (3, 3’-diaminobenzadine) disclosure. Nuclei were counterstained with hematoxylin. Images were captured using an NanoZoomer Digital Pathology system (Hamamatsu Photonics, Hamamatsu, Japan). USP19 expression levels were semiquantitatively assessed in tissue samples. Both the extent and intensity of USP19 immunostaining were taken into consideration when analyzing the data. The intensity of staining was scored from 0 to 3 and the extent of staining was from 0 to 100%. The final quantitation of each staining was obtained by multiplying the two scores. The slides were analyzed by two independent pathologists.
5
Quantifying Thrombus Composition and NETosis
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Whole microscopic digital images of the slides were captured using the NanoZoomer Digital Pathology System (Hamamatsu Photonics, Hamamatsu, Japan). Examples of quantification are shown in
Supplementary Fig. S1 (available in the online version). The thrombus size was evaluated based on the cross-sectional area, and each component of the specimen was semi-automatically quantified using the entire specimen with the Fiji-ImageJ software package.
24 (link)
Compared with the area of the entire specimen, the proportional areas were evaluated for red blood cells (RBCs) using H&E staining, fibrin using phosphotungstic acid–hematoxylin staining, and platelets using immunohistochemical staining for CD42b. The densities of white blood cells and CD163-positive cells were evaluated based on H&E staining and immunohistochemical staining.
NETosis is the process of extracellular trap formation by thread-like structures of decondensed DNA that are decorated with proteins from cytoplasmic granules.
25 (link)
The extent of NETosis in thrombi was evaluated using the density of H3Cit-positive cells in all patients because H3Cit is a marker for immune cells that are about to release extracellular traps.
26 (link)
The density of H3Cit-positive cells was assessed based on immunohistochemical staining as described above.
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Whole microscopic digital images of the slides were captured using the NanoZoomer Digital Pathology System (Hamamatsu Photonics, Hamamatsu, Japan). Examples of quantification are shown in
24 (link)
Compared with the area of the entire specimen, the proportional areas were evaluated for red blood cells (RBCs) using H&E staining, fibrin using phosphotungstic acid–hematoxylin staining, and platelets using immunohistochemical staining for CD42b. The densities of white blood cells and CD163-positive cells were evaluated based on H&E staining and immunohistochemical staining.
NETosis is the process of extracellular trap formation by thread-like structures of decondensed DNA that are decorated with proteins from cytoplasmic granules.
25 (link)
The extent of NETosis in thrombi was evaluated using the density of H3Cit-positive cells in all patients because H3Cit is a marker for immune cells that are about to release extracellular traps.
26 (link)
The density of H3Cit-positive cells was assessed based on immunohistochemical staining as described above.
6
Immunostaining of Galectin-7 in TMAs
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Representative specimens from our previous TMA analysis were immunostained for gal-7 using the Discovery XT automated immunostainer (Ventana Medical Systems, Tucson, AZ) [6 (link)]. Deparaffinized sections were incubated in Cell Conditioning 1 (pH 8.0) for antigen retrieval and then stained for 60 min with the anti-human gal-7 polyclonal antibody (R&D Systems, Minneapolis, MN) using a 1:150 dilution. The slides were counterstained with hematoxylin and bicarbonate. Each section was scanned at a high resolution using the Nanozoomer Digital Pathology System (Hamamatsu, Bridgewater, NJ). The study was approved by the research ethics committee of the research center at the Centre Hospitalier de l’Université de Montréal (approval No. SL 05.019).
7
Histological Analysis of Harvested Organs
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Chosen organs were harvested from treated mice, fixed in 10% formalin before mounting in paraffin. Sections were then stained with hematoxylin and eosin, then scanned using the NanoZoomer Digital Pathology system and NPD.scan 2.3.4 software (Hamamatsu, Hamamatsu city, Japan).
8
Histological and Apoptosis Analysis of Tumor Samples
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At the end of therapeutic period, the mice were euthanized. Their tumors were then collected for pathological studies, respectively. The tumors were fixed in 10% formalin, embedded in paraffin, sectioned, and then stained by H&E. The stained tissue sections were then observed using NanoZoomer digital pathology system (Hamamatsu, Japan) to monitor the possible histological changes. For assessment of cell apoptosis, TUNEL assays were performed by following the manufacturer’s protocol (Roche, Germany). The stained tissue sections were counterstained with 4, 6-diamidino-2-phenylindole (DAPI) (Thermo Fisher Scientific, USA) and captured by CLSM (Olympus, FV1000MPE, Japan).
9
Detailed Histological Evaluation of Ocular Tissues
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Eyeballs were fixed in dedicated eyeball fixed liquid (glacial acetic acid: formaldehyde: normal saline: ethyl alcohol=1:2:7:10) for at least 48 h. After gradient ethanol dehydration, paraffin embedding and other traditional sample disposal processes as previously described,33 (link),34 (link) 4 micron (μm) sections were cut through optic nerve in sagittal position. Then the sections were baked and dewaxed before hematoxylin and eosin (H&E) staining. After vitrification by dimethylbenzene and sealing by resinene, H&E staining sections were scanned with NanoZoomer Digital Pathology System (Hamamatsu Photonics Co., Ltd) and viewed with NDP view software. Then the thickness of cornea, retina and choroid were measured. The description of corneal thickness was from corneal epithelium layer to endothelial layer, and retinal thickness was from retinal internal limiting membranes to pigment epithelium layer. As for choroid thickness, it was from Bruch’s membrane (connected with retinal pigment epithelium layer) to sub-choroid space (attached to the sclera).
10
Histological Analysis of Intestinal Morphology
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Tissue samples fixed in a 10% formalin solution were dehydrated through a series of alcohol solutions, cleared in xylene, and embedded in paraffin wax (Microtechnical Services Ltd., Exeter, UK). Sections (3 to 5 μm thick) were prepared and stained with either modified hematoxylin and eosin (H&E) or periodic acid-Schiff (PAS) using standard protocols. After staining, the slides were scanned with the NanoZoomer digital pathology system (Hamamatsu, Welwyn Garden City, UK). Measurements of villus height and crypt depth were made using the NanoZoomer digital pathology image program (Hamamatsu). Ten well-oriented villi per tissue section of 4 birds from each diet group at each sampling time were scanned at 40× resolution for each tissue sample. Villus height was measured from the tip of the villus to the crypt opening, and the associated crypt depth was measured from the base of the crypt to the level of the crypt opening. The ratio of villus height to relative crypt depth was calculated from these measurements. Goblet cells were enumerated from ileal sections stained with PAS. In one case the histology section did not fulfill the quality criterion of 10 well-oriented villi and was omitted from the analysis.
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