Oil red O staining was performed to determine the number of neutral lipids deposited in the liver, the method was described as Yin et al. (2021) [12 (link)]. Liver samples were firstly fixed with 4% paraformaldehyde and then frozen with liquid nitrogen. Secondly, the sectioned slides (6–10 μm thickness) were stained with oil red O solution (3 g L−1, Servicebio, Wuhan, China) for 8–10 min. Then, the nuclei were counterstained with hematoxylin solution (Servicebio, Wuhan, China) for 3–5 min. After staining, the slides were washed and a cover glass was fixed to the slide with glycerol jelly mounting medium (Servicebio, Wuhan, China). The lipid droplets were observed using a light microscope (Nikon Eclipse Ni-U, Tokyo, Japan) at 100× magnification.
Hematoxylin solution
Manufactured by Wuhan Servicebio Technology
Sourced in China
Hematoxylin solution is a laboratory reagent used in histology and cytology for the staining of cell nuclei. It is a natural dye extracted from the heartwood of the logwood tree. The solution contains hematoxylin, a chromogenic compound that binds to the DNA and RNA in cell nuclei, staining them blue or purple.
Automatically generated - may contain errors
Lab products found in correlation
Eosin solution, by Wuhan Servicebio Technology (4 mentions)
Hematoxylin differentiation solution, by Wuhan Servicebio Technology (2 mentions)
Bx53f, by Olympus (2 mentions)
Glycerol jelly mounting medium, by Wuhan Servicebio Technology (1 mentions)
Oil red o solution, by Wuhan Servicebio Technology (1 mentions)
Eclipse ni u, by Nikon (1 mentions)
Optical microscope, by Olympus (1 mentions)
Eosin staining solution, by Wuhan Servicebio Technology (1 mentions)
Chemiluminescent horseradish peroxidase hrp substrate, by Merck Group (1 mentions)
Nitrocellulose membrane, by Merck Group (1 mentions)
Gefitinib, by Selleck Chemicals (1 mentions)
Sapk jnk, by Cell Signaling Technology (1 mentions)
Cyclin d1, by Cell Signaling Technology (1 mentions)
Ptyr705 stat3, by Cell Signaling Technology (1 mentions)
Ptyr701 stat1, by Cell Signaling Technology (1 mentions)
Pser9 gsk3β, by Cell Signaling Technology (1 mentions)
Pthr308 akt, by Cell Signaling Technology (1 mentions)
Antibodies against α tubulin, by Santa Cruz Biotechnology (1 mentions)
P jak1 tyr1022 1023, by Cell Signaling Technology (1 mentions)
P jak2 tyr1007 1008, by Cell Signaling Technology (1 mentions)
17 protocols using hematoxylin solution
1
Oil Red O Staining of Liver Lipids
2
Hematoxylin and Eosin Staining Protocol
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HE staining was conducted according to the routine protocol [17 (link)]. In brief, sections were stained with hematoxylin solution (Servicebio, China) for 5 min followed by five dips in 1% acid ethanol (1% HCl in 70% ethanol), and then rinsed in distilled water. Subsequently, the sections were stained with eosin solution for 3 min followed by dehydration with graded alcohol and clearing in xylene. The stained slices were photographed and examined using an optical microscope (Olympus, Japan).
3
Histological Analysis of BMAL1 Monkey Tissues
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For histological analysis, skin and muscle tissues from BMAL1+/+ and BMAL1–/– cynomolgus monkeys were fixed with 4% PFA for 2 days, embedded in paraffin and sectioned at 5-μm intervals. The sections were deparaffinized and rehydrated by xylene and graded series of ethanol (100%, 100%, 95%, 80%, 70%, and 50%), and then washed by distilled water followed by staining with hematoxylin solution (Servicebio, China). After 5-min incubation of hematoxylin, the sections were washed with running water, differentiated with 1% acid alcohol, and washed with running tap water again until the nuclei were stained to a suitable intensity. Next, the sections were counterstained in the eosin (Solarbio), followed by dehydration in a grader series of ethanol (50%, 70%, 80%, 90%, 95%, 100%, and 100%), and immersion in xylene. Finally, the sections were mounted on the coverslip with neutral balsam for long-term preservation.
4
Histological Analysis of Basal Ganglia Hematoma
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Tissues from the hematoma area of the right basal ganglia area are collected for pathological examination. Part of brain tissues were fixed with 10% formaldehyde (Solarbio, Beijing, China) for 10 min and embedded in paraffin. Slides, 4 μm thick, were cut, deparaffinated with xylene, and rehydrated with descending concentration of ethanol (100%, 96%, 80%, 70%, and 50% for 2 min, respectively). Afterwards, slides were stained with hematoxylin solution (Servicebio, Wuhan, China) for 5 min and in hydrochloric acid-ethanol mixtures for 3s, followed by washing for 15 min. Slides were stained with 0.5% eosin solution (Servicebio) for 1 min. Slides were treated with 80%, 95% and 100% alcohol for 2 min respectively, and dehydrated with xylene for 2 min. Pathological examination was performed under a microscope (Nikon, Japan). Three rats were examined per group. Five visual fields were randomly selected from the hematoma area of the right basal ganglia area for each slide.
5
Paraffin-Embedded Tissue Staining
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Paraffin sections were dewaxed with xylene, dehydrated with gradient alcohol, and then placed in distilled water. The samples were stained with hematoxylin solution (Servicebio, Wuhan, China) for 5 min. After washing with distilled water for 3 times, using 1% hydrochloric acid alcohol for color separation for 20 s. followed by incubation in 0.1M PBS pH7.4 for 3 min, The slices were washed through a graded alcohol concentrations for 2 min sequentially and stained with eosin (Servicebio, Wuhan, China) for 30 s. Finally, the samples were detained in 95% alcohol for 1 min, dehydrated in anhydrous ethanol and xylene.
6
Signaling Pathway Antibody Panel for JAK-STAT Analysis
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Antibodies against p-Tyr1022/1023-JAK1 (#3331), p-Tyr1007/1008-JAK2 (#3776), p-Tyr980/981-JAK3 (#5031), p-Tyr1054/1055-TYK2 (#9321), p-Tyr701-STAT1 (#9167), p-Tyr705-STAT3 (#9145), p-Tyr694-STAT5 (#9359), JAK1 (#3332S), JAK2 (#3230), JAK3 (#8863), TYK2 (#9312), STAT1 (#14994), STAT3 (#12640), STAT5 (#25656), c-Myc (#13987), cyclin D1 (#AF0931), Bcl-xL (#2764), p-Ser536-NF-κB (#3033), p-Thr308-AKT (#9275), p-Ser9-GSK-3β (#5558), p-Thr183/Tyr185-SAPK/JNK (#4668), NF-κB (#8242), AKT (#4691), GSK-3β (#9832), and SAPK/JNK (#9252) were purchased from Cell Signaling Technology (Danvers, MA, USA). Antibodies against α-tubulin were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Protease inhibitor (B14001) and phosphatase inhibitor (B15001) were purchased from Bimake (Houston, TX, USA). Recombinant human STAT3 protein (ab268982) was obtained from Abcam (Cambridge, Britain). Recombinant human interleukin-6 (IL-6) protein (200-06) was purchased from PeproTech (Rocky Hill, CT, USA). Nitrocellulose membranes and chemiluminescent horseradish peroxidase (HRP) substrate were obtained from Millipore (Billerica, MA, USA). Gefitinib was purchased from Selleckchem (Houston, TX, USA). DAB color solution, hematoxylin solution, and eosin staining solution were purchased from Servicebio (Wuhan, China).
7
Caspase-3 Immunohistochemistry in Rat Brain
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The paraffin-embedded slides were dewaxed with xylene and rehydrated with descending concentrations of ethanol. A 10 mmol/L sodium citrate solution was prepared for antigen retrieval for 6 min and then placed at room temperature for 1 h. Slides were washed with PBST for 5 min and incubated with 0.3% H2O2/methanol solution for 30 min at 4°C to inactivate endogenous peroxidase activity, and then blocked using 3% bovine serum albumin (Ausbian, Australia). Subsequently, slides were incubated with a primary antibody against Caspase-3 (Abcam, UK; 1:100) overnight at 4°C and then incubated with horseradish peroxidase-linked streptavidin (Thermo Fisher) for 80 min. After washing with PBS, color was developed by a mixture of 85% double distilled water +5% 3,3’-diaminobenzidine (DAB), 5% DAB substrate +5% DAB chromogen for 3 min (DAB substrate kit; Thermo Fisher). Next, slides were stained with hematoxylin solution (Servicebio) for 5 min and coverslipped with resin. Three rats were examined per group, and five visual fields were randomly selected from the hematoma area of the right basal ganglia area for each slide. Positive Caspase-3 staining was observed under a light microscopy, and the brown chromogen at the edge of the hematoxylin-stained cell nuclei and in the cytoplasm or plasma membrane of the cells was considered as positive staining.
8
Histological Analysis of Prefrontal Cortex and Hippocampus
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We fixed the prefrontal cortex and hippocampal tissues in 4% paraformaldehyde for 1 day, dehydrated them in a graded series of alcohols, subsequently embedded them in paraffin, and cut them into 5 mm-thick sections. The tissues were then stained with Hematoxylin and Eosin (H&E) according to routine protocols. Briefly, after dewaxing and rehydration, tissue sections were stained with hematoxylin solution (Servicebio, China) and rinsed. They were then stained with eosin solution (Servicebio, China), dehydrated with graded alcohol, and removed in xylene. Sections were assessed using an upright optical microscope (Nikon Eclipse E100, Tokyo, Japan).
9
Histological Analysis of Hippocampal Neurons
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Six Mice from each group without undergoing behavior test were anesthetized with chloral hydrate and then sacrificed. To evaluate histological damage, mice were perfused with saline and paraformaldehyde. Their brains were removed, fixed in paraformaldehyde for 24 h, dehydrated in alcohol, and embedded in wax. The wax was trimmed and sectioned into 4 μm slices for staining with HE and Nissl. Then, the sections were dewaxed and dehydrated using xylene and ethanol solutions before being rinsed with tap water. For HE staining, they were stained with hematoxylin solution (Servicebio, G1003) and treated with differentiation and bluing solutions before being fixed with ethanol and stained with Eosin dye. The sections were then dehydrated and placed in xylene before being sealed with neutral gum. For Nissl Staining, they were stained with Nissl dye (Servicebio, G1036) and treated with a differentiation solution before being rinsed and sealed with neutral gum. The pathological changes in the hippocampus were observed under a light microscope. The Nissl-stained positive neurons in the hippocampal dentate gyrus (DG) region were counted under a light microscope. Besides, each section were visually counted in a blinded manner. The results show the different number of surviving neurons in CIH group compared to the CON group in same regions (Chu et al., 2019 (link); Ke et al., 2020 (link)).
10
Histological Analysis of Adipose Tissue
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The fresh dissected AT was fixed with a fixator for more than 24 h and subjected to alcohol gradient dehydration. Then, the wax-soaked AT was embedded and placed in the paraffin slicer for sectioning to a 4 μm slice. The paraffin sections were dewaxed and dyed with hematoxylin solution (Servicebio, Wuhan, China) for 3–5 min. Hematoxylin differentiation solution (Servicebio) was differentiated for 2–5 s, and the Hematoxylin Scott Tap Bluing (Servicebio) was treated for 2–5 s. The slices were dyed in eosin solution (Servicebio) for 5 min and then placed in gradient alcohol and xylene for 5 min, respectively, and sealed with neutral gum. Lastly, the tissue sections were observed under the Olympus BX51 microscope (Olympus) and the images were collected for analysis.
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