The dorsal skin and one ear per mouse were fixed in 10% (v/v) neutral buffered formalin for 24 h at 4°C. Tissue samples were embedded in paraffin and sectioned (4 μm thickness). To evaluate tissue architecture, sections were stained with a hematoxylin and eosin (H&E) solution (Sigma-Aldrich, St. Louis, MO, USA) and mounted under coverslips using a Dako mounting medium (DakoCytomation, Glostrup, Denmark). Epidermal and ear thicknesses were quantified by microscopic examination. Briefly, 10 randomly selected areas were observed in H&E-stained preparations (from 3 sections per each animal), under the microscope (Olympus Microscope System CKX53; Olympus, Tokyo, Japan). Representative images were acquired, and thickness measurements were performed using the ImageJ 1.50i software (National Institutes of Health, Bethesda, MD, USA). To measure the degree of mast cell infiltration, sections were stained with toluidine blue (TB), and the number of mast cells was quantified in four fields of view, under the microscope (Olympus Microscope System CKX53; Olympus, Tokyo, Japan). All measurements/quantifications were performed in a blinded manner.
Microscope system ckx53
Manufactured by Olympus
Sourced in Japan
The Olympus Microscope System CKX53 is a versatile and reliable microscope designed for various laboratory applications. It features a compact and ergonomic design, allowing for efficient use in limited space. The system provides high-quality optical performance and can be equipped with a range of accessories to suit different needs.
Automatically generated - may contain errors
Lab products found in correlation
H e solution, by Merck Group (3 mentions)
Dako mounting medium, by Agilent Technologies (2 mentions)
Freezing microtome, by Leica (1 mentions)
Mouse anti cytokeratin 14, by Abcam (1 mentions)
Vectashield mounting medium containing dapi, by Vector Laboratories (1 mentions)
Rabbit anti aβ, by Abcam (1 mentions)
7 protocols using microscope system ckx53
1
Histological Analysis of Mouse Skin and Ear
2
Quantitative Histological Analysis of Tissue Architecture
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To evaluate tissue architecture, sections were stained with hematoxylin and eosin (H&E) solution (Sigma-Aldrich) and mounted under cover slips using Dako mounting medium (Dako Cytomation, Glostrup, Denmark). Epidermal thickness was quantified by microscopic examination. Briefly, 10 randomly selected areas were observed in H&E-stained sections (three sections per animal) under a microscope (Olympus Microscope System CKX53; Tokyo, Japan). To measure the degree of mast cell infiltration, sections were stained with toluidine blue (TB), and mast cells were counted in four fields of view under a microscope (Olympus Microscope System CKX53). Collagen density was measured using the Masson’s Trichrome (MT) Stain Kit (Vitro Vivo Biotech, Rockville, MD, USA), according to the manufacturer’s protocol. All measurements and quantitative analyses were performed according to a previously published method [31 (link)].
3
Iba-1 Immunohistochemistry in Brain Tissue
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Brains were sectioned via a freezing microtome (Leica Instruments GmbH, Nussloch, Germany). Brain tissue fixation on cover slips and further steps were carried out as our previously described [29 (link)]. Detection of the Iba-1 was performed using a microscope (Olympus Microscope System CKX53; Olympus, Tokyo, Japan).
4
Histopathological Analysis of Liver Tissue
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To perform histopathology, the liver tissue was fixed in 10% neutral-buffered formalin. Tissue samples were then embedded into paraffin and sectioned into 4 μm thick slices, followed by staining with hematoxylin and eosin (H&E) solution (Sigma-Aldrich, St. Louis, MO, USA). The stained sections were examined under a light microscope (Olympus Microscope System CKX53; Olympus, Tokyo, Japan).
5
Immunofluorescence Staining of Dorsal Skin
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Dorsal skin sections were rinsed briefly in PBS and treated with 0.5% BSA for 30 min. The dorsal skin tissue on the slide was incubated with rabbit anti-filaggrin (1:500 dilution; GeneTex Inc., Irvine, CA, USA), mouse anti-involucrin (1:500 dilution; Invitrogen, Carlsbad, CA, USA), rabbit anti-loricrin (1:500 dilution; Abcam, Cambridge, UK), and mouse anti-cytokeratin 14 (1:500 dilution; Abcam) overnight at 4 °C in the presence of 0.3% Triton X-100 and normal goat serum. The slides were then incubated for 2 h with an Alexa Fluor-conjugated secondary antibody (diluted 1:500). The dorsal skin tissue on the slide was washed in PBS and mounted using Vectashield mounting medium, containing 4′,6-diamidino-2-phenylindole. Images were obtained using a fluorescence microscope (Olympus Microscope System CKX53). A threshold for positive staining was determined for each image that included processes but excluded background staining. Dorsal skin tissue regions were analyzed quantitatively using ImageJ 1.50i (National Institutes of Health).
6
Hematoxylin-Eosin Staining of 3HSE Skin Sections
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Skin sections from the 3HSE were prepared for hematoxylin and eosin staining. Sections (5 μm thick) of 10% neutral formalin solution-fixed paraffin-embedded tissues were cut on saline-coated glass slides and then deparaffinized 3 times with xylene and dehydrated through a graded alcohol bath. The deparaffinized sections were stained with hematoxylin for 5 min. The slides were then washed and stained with EosinY. Finally, they were dehydrated and washed. Representative images were taken using a fluorescence microscope (Olympus Microscope System CKX53).
7
Immunostaining of Amyloid-Beta in Brain Sections
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The brain sections were rinsed briefly in PBS and treated with 0.5% BSA for 30 min. The sections were incubated with rabbit anti-Aβ (1:500 dilution; Abcam, Cambridge, UK) overnight at 4 °C in the presence of 0.3% Triton X-100 and normal goat serum. They were then incubated for 2 h with an Alexa Fluor conjugated secondary antibody (diluted 1:500). Finally, sections were washed in PBS and mounted using Vectashield mounting medium containing DAPI (Vector Labs, Burlingame, USA). The images were taken using a fluorescence microscope (Olympus Microscope System CKX53; Olympus, Tokyo, Japan). A threshold for positive staining was determined for each image which included all cell bodies and processes but excluded any background staining.
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