For histological analysis, tissues were sliced at 5 μm thickness sections. Modified Masson's Trichrome Stain Kit (Scy Teklaboratories, USA) was used to determine interstitial fibrosis, which can distinguish blue fibrosis and red muscle. All sections were scanned with digital microscopy (Olympus, Japan) equipped with a 20X objective lens for 15 to 20 images per sample. All images were acquired under same conditions. The automatic exposure and white balance were turned off. Images were analyzed with Image J software (version 1.48v; National Institutes of Health). The area of fibers in deconvoluted color images was measured with ‘Threshold’ tool. Blood vessels, perivascular tissue and epicardium were excluded when measuring fibrotic area for fibrosis quantification. For transmission electron microscopy observation, rat hearts were fixed in 2% glutaraldehyde and immersed in 2% osmium tetroxide and 1% aqueous uranyl acetate for 1 h. The sample was washed by a series of ethanol solution and incubated into propylene oxide and EMbed 812 mixtures for 1 h, followed by polymerization at 70 °C. After sliced at 80 nm sections, 5% uranyl acetate and Reynold's lead citrate were used to stain. Sections were observed by a 40–120 kV transmission electron microscope (Hitachi H600 Electron Microscope, Hitachi, Japan). At least 10 fields of each sample were analyzed.
Digital microscopy
Manufactured by Olympus
Sourced in Japan
Digital microscopy is a type of microscopy that utilizes digital imaging sensors and software to capture, process, and display microscopic images. The core function of digital microscopy is to provide a digital representation of specimens under magnification, enabling users to view, analyze, and document microscopic details.
Automatically generated - may contain errors
6 protocols using digital microscopy
1
Histological and Ultrastructural Analysis of Cardiac Fibrosis
2
Quantifying Cell Proliferation with EdU
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Cell proliferation was detected using an EdU kit (RiboBio, Guangzhou,
China) according to the manufacturer’s instructions. Images were
analyzed using digital microscopy (Olympus), and cells were counted
using Image J software.
China) according to the manufacturer’s instructions. Images were
analyzed using digital microscopy (Olympus), and cells were counted
using Image J software.
3
Senescence-Associated β-Galactosidase Assay
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Detection of senescence was carried out using a senescence-associated
β-galactosidase staining kit (Beyotime Biotechnology, Haimen, China).
Cells were cultured with the mixture overnight at 37°C in a 24-well
plate and analyzed using digital microscopy (Olympus, Tokyo,
Japan).
β-galactosidase staining kit (Beyotime Biotechnology, Haimen, China).
Cells were cultured with the mixture overnight at 37°C in a 24-well
plate and analyzed using digital microscopy (Olympus, Tokyo,
Japan).
4
Evaluating SLNCR1 Impact on Melanoma Migration
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To evaluate the effect of SLNCR1 on melanoma cell migration, a wound healing assay was performed Approximately 5 × 10^4 cells were seeded in a 6-well plate. After 48 h, the cells were wounded to create a straight scratch. The wells were then washed and treated with fresh medium containing 2% FBS. Digital microscopy (Olympus) was used to capture images of each wound at 0, 24, and 48 h. The wound space was calculated by subtracting the scratch widths at 24 or 48 h from the width at 0 h using the MRI tool in Image J.
5
Apoptosis Quantification in Human SG
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Apoptosis in human SG tissues was identified by TUNEL staining using an in-situ Cell Death Detection Kit (Roche, Germany). Numbers of apoptotic cells were quantified by digital microscopy (Olympus, Japan), and apoptotic indices were calculated using three random fields per sample.
6
Transwell Assay for Cell Migration
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Transwell permeable support (8-μm, Corning, Tewkesbury, USA) was used for cell migration. Cells, transfected for 48 h or non-transfected, were suspended in serum-free medium into the upper chamber, culture medium containing 10% FBS was added to the lower chamber. After 24 h of incubation, the cells stayed on the upper surface of the membrane (non-invasive or non-migrating) were removed. Cells on the bottom surface of the membrane (migratory) were xed with 4% paraformaldehyde for 10 min, and stained with 0.4% crystal violet solution. The migratory cells were imaged using a digital microscopy (Olympus, Tokyo, Japan).
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