Silibinin (purity ≥ 99.0%) and hematoxylin and eosin (H&E) were procured from Beijing Solarbio Science & Technology Co., Ltd. (Beijing, China). Other solvents and reagents were obtained as analytical grade and used without further purification. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was purchased from Sigma (Tokyo, Japan). Hoechst 33342/propidium iodide (PI) double staining kit was bought from Genview (Beijing, China), and a 5-ethynyl-20-deoxyuridine (EdU) assay kit was acquired from Ribobio (Guangzhou, China). Antibodies including Anti-CASP3, Anti-MMP3, Anti-SRC, Anti-MAPK10, Anti-CDK6, Anti-PPARα and Anti-JAK were obtained from Cell Signaling Technology (Shanghai, China).
Hematoxylin and eosin h e
Manufactured by Solarbio
Sourced in China
Hematoxylin and eosin (H&E) is a widely used staining technique in histology and pathology. Hematoxylin stains cell nuclei blue, while eosin stains cytoplasm and other structures pink. This combination allows for the visualization of tissue morphology and cellular details.
Automatically generated - may contain errors
Lab products found in correlation
Bca kit, by Thermo Fisher Scientific (3 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Paraformaldehyde (pfa), by Solarbio (2 mentions)
Anti casp3, by Cell Signaling Technology (1 mentions)
Anti src, by Cell Signaling Technology (1 mentions)
Anti pparα, by Cell Signaling Technology (1 mentions)
3 4 5 dimethylthiazol 2 yl 2 5 diphenyltetrazolium bromide mtt, by Merck Group (1 mentions)
5 ethynyl 20 deoxyuridine edu assay kit, by RiboBio (1 mentions)
Silibinin, by Solarbio (1 mentions)
Anti mmp3, by Cell Signaling Technology (1 mentions)
Anti jak, by Cell Signaling Technology (1 mentions)
Anti cdk6, by Cell Signaling Technology (1 mentions)
3 4 5 dimethyl 2 thiazolyl 2 5 diphenyl 2h tetrazolium bromide mtt, by Merck Group (1 mentions)
Masson trichrome staining, by Solarbio (1 mentions)
Rhodamine phalloidine, by Merck Group (1 mentions)
N methyl pyrrolidone (nmp), by Merck Group (1 mentions)
4 6 diamino 2 phenylindole dapi, by Merck Group (1 mentions)
N 3 dimethylaminopropyl n ethylcarbodiimide hydrochloride edc, by Merck Group (1 mentions)
Citric acid buffer, by Solarbio (1 mentions)
Horseradish peroxidase conjugated secondary antibody, by Abcam (1 mentions)
25 protocols using hematoxylin and eosin h e
1
Silibinin-Mediated Cellular Mechanisms
2
Histological Analysis of Bladder and Nerve
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To further evaluate the bladder and nerve, the rats were sacrificed at 1 week, 4 week and 6 week (three time points) after surgery. The bladder and injury sites of the spinal sacral nerve were harvested, and the bladders were weighed (Mettler Toledo, Switzerland). As previously described, the nerves and bladders were fixed with 4% paraformaldehyde at 4 °C for 24 h. Thereafter, all the tissues were embedded in OCT Tissue Tek (Sakura, USA) and serially sectioned at 6-μm thickness. Subsequently, the bladder slices were subjected to hematoxylin–eosin (H&E) and Masson’s trichrome staining, and the nerve slices were stained with hematoxylin–eosin (H&E). After staining, light microscopic examination was conducted in tissue sections to evaluate the structure of the bladder and nerve (Olympus, Japan). The thickness of the bladder wall was measured, and the collagen tissue percentage was calculated as the ratio collagen/smooth muscle. The nerves were stained with hematoxylin and eosin (H&E) (Solarbio Life Sciences, Beijing, China). All the calculations and measurements were conducted using an image analysis system (ImageJ 1.50).
3
Micromorphometric Analysis of Testis
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One week after the first intraperitoneal injection of miR-199-5p antagomir or agomir, the testis was collected and fixed in Bouin’s fixative (Phygene, Fuzhou, China) overnight, dehydrated in an increasing ethanol gradient, cleared in xylene, embedded in paraffin, sectioned into 5 μm slices using a Leica RM2016 rotary microtome (Leica Biosystems GmbH, Nussloch, Germany), dewaxed with xylene, rehydrated with ethanol, and stained with hematoxylin and eosin (H&E) (Solarbio, Beijing, China), as described everywhere. The testis sections were examined and photographed using an Olympus microscope CX41 (Olympus Co., Tokyo, Japan).
4
Femur Histology and Immunohistochemistry
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Femurs were cleaned off tissue, fixed in 4% paraformaldehyde at 4°C for 24 h, and then decalcified in 10% ethylenediaminetetraacetic acid (pH 7.2) at room temperature for 14 days. The segmental bone was embedded in paraffin and cut into 3.5 μm thick sections. Then, sections were stained with hematoxylin and eosin (H&E) (Solarbio Biotechnology, Beijing, China) for histological analysis. The sections were then deparaffinized and briefly washed with PBS for immunohistochemistry. They were then incubated in 3% H2O2 to block endogenous peroxidase activity. Bax and beclin-1 primary antibodies were incubated with bone sections overnight at 4°C. The sections were cleaned with PBS and incubated with goat anti-rabbit horseradish peroxidase-conjugated secondary antibodies for 30 min. The immunostained sections were then dyed with DAB. A series of five random fields was photographed per section. The areas of positive (immunoreactive) cells and total cells in each section were counted and quantitatively characterized using Image-Pro Plus 6.0.
5
Multifunctional Nanocomposite Synthesis
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PLGA (molecular weight=100,000, LA/GA=75/25) was obtained from ChangchunSinoBiomaterials Co., Ltd. GO was purchased from Chengdu Organic Chemicals Co. Ltd, China (thickness: 0.55–1.2nm diameter:0.5–3μm) (MFG Code 021519). L-lysine was purchased from Beijing Chemical Reagent Co., Ltd, China (Product number 62016734). HAuCl4·3H2O (CAS number 16961-25-4), Cetylpyridinium chloride (CPC) (CAS number 6004-24-6) and dopamine hydrochloride (CAS number 62-31-7) was purchased from Aladdin Reagents Co., Ltd, China. N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDC) (CAS number 25952-53-8), N-Methylpyrrolidone (NMP) (CAS number 872-50-4), 3-(4,5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide (MTT) (CAS number 298-93-1), 4,6-diamino-2-phenylindole (DAPI) (CAS number 28718-90-3) and rhodamine-phalloidine (Product number P1951) were purchased from Sigma-Aldrich (USA). The reagents for cell experiments were purchased from Sigma-Aldrich (USA). Hematoxylin and eosin (H&E) (Product number G1120), Masson trichrome staining (Product number G1340) and Sirius Red kits (Product number S8060) were purchased from Solarbio Co., Ltd, China.
6
Liver Histopathology after UMSC Transplant
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Mice were sacrificed at the designated time points (12 h, 1 day, 2 days, 5 days, 7 days, 14 days, 21 days, and 28 days) after UMSC transplantation. Liver lobes were excised from the same site and fixed with 4% paraformaldehyde (Solarbio, Beijing, China) for 24 h for histological and immunological analyses. Paraffin-embedded liver tissue was sliced into 5-µm sections and stained with hematoxylin and eosin (HE; Solarbio). Pathological changes in liver tissue were assessed under an optical microscope. For immunohistochemical analysis, the tissue sections were heated in citric acid buffer (0.02 mol/L, pH=5.8) (Solarbio) for antigen retrieval. Bovine serum albumin (5%; Sigma-Aldrich) in phosphate-buffered saline (Solarbio) was used to block non-specific binding. Then, according to the instructions of the reagent manufacturer, the sections were incubated with an anti-MyD88 antibody (Abcam, Cambridge, UK) overnight at 4°C. The sections were then incubated with a horseradish peroxidase-conjugated secondary antibody (Abcam) at 37°C for 1 h and evaluated under an optical microscope. The optical density value was calculated by ImagePro Plus software (Media Cybernetics, Inc., Rockville, MD, USA).
7
Comprehensive Colitis Evaluation: Standardized Scoring and Histopathology
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The macroscopic grading of DSS-induced colitis is based on a standard scoring system called the disease activity index (DAI) [24 (link)]. The details are shown in Table 1 . The body weight, stool consistency, and rectal bleeding of each mouse were recorded in fact daily. All mice were anesthetized and sacrificed by cervical dislocation 24 h after the last drug treatment. Their colons were excised, and the length was measured promptly. The distal colonic tissues were fixed in 4% paraformaldehyde solution (Servicebio, China), embedded in paraffin (Solarbio, China), sectioned, and then stained with hematoxylin and eosin (HE) (Solarbio, China) for optical microscopic (CKX53, Olympus, Japan) observation. Histopathological scores were determined according to the previous criteria [25 (link)] shown in Table 2 .
8
Histological Analysis of Aortic Atherosclerosis
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The aortic roots along with the basal portion of the heart were fixed with a 4% paraformaldehyde, followed by embedding in an OCT compound (SAKURA, Torrance, CA, USA). To assess the atherosclerotic lesion in aortic roots, serial cross-sections (7 μm thick) of aortic roots were prepared from the site where the three aortic valves appeared. The frozen sections were then stained with hematoxylin and eosin (H&E) (Solarbio, Beijing, China). Immunohistochemistry was performed on the aortic root for the expression of IL-6. Briefly, 7 μm thick sections were stained with primary antibody overnight at 4 °C, then the secondary antibody conjugated with horseradish peroxidase was added. The DAB substrate solution was used to induce the formation of colored precipitate at the tissue antigen-binding sites, and then the hematoxylin was added.
9
Histological Analysis of Mouse Kidneys
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The mice were anesthetized by inhalation of carbon dioxide and then euthanized by cervical dislocation. The kidneys were separated and fixed in 4% paraformaldehyde (Solarbio Life Sciences, Beijing, China), then embedded in paraffin and cut into sections with 4 µM thickness. The sections were stained with hematoxylin and eosin (H&E) (Solarbio) after paraffinization and rehydration, then mounted with neutral gum [22 (link)].
10
Histological Analysis of Tumor Burden
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The tumors harvested from the tumor‐bearing mice were fixed with 4% paraformaldehyde, and then the tumor sections were embedded in paraffin. To examine the tumor burden, hematoxylin and eosin (HE) (Solarbio) was used to stain some of the tumor sections, whereas some of the sections were subjected to TUNEL staining to detect apoptosis. For the TUNEL‐staining assay, proteinase K was used to treat all sections for 15 min at room temperature. The sections were further stained with a TdT‐mediated dUTP Nickel End Labelling Kit (Beyotime).
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