Tissue sections of 5 µm were stained with haematoxylin/eosin or analysed immunohistochemically according to the protocol (Mouse- and Rabbit-specific HRP/DAB Detection IHC Kit, Abcam). Shortly, sections were de-paraffinised, rehydrated and upon blocking of unspecific binding, incubated with primary antibodies for 1 h at room temperature. Detection of target antigens was performed via streptavidin-peroxidase-mediated colour reaction of AEC or DAB. Following primary antibodies were used: Ki-67 (Dako, clone M7240, 1:80), calcitonin (Novus, clone SP17, 1:10) synaptophysin (Pierce Thermo Scientific, 1:80), chromogranin A (Pierce Thermo Scientific, 1:500) and carcinoembryonic antigen (Dako, cloneII-7, 1:1000). Images were taken on an Olympus BX53.
Mouse and rabbit specific hrp dab detection ihc kit
Manufactured by Abcam
Sourced in United Kingdom
The Mouse and Rabbit Specific HRP/DAB Detection IHC kit is a laboratory tool designed for immunohistochemistry (IHC) applications. It provides a horseradish peroxidase (HRP) and 3,3'-diaminobenzidine (DAB) detection system for the visualization of mouse and rabbit primary antibodies in tissue samples.
Automatically generated - may contain errors
Lab products found in correlation
Ab18197, by Abcam (2 mentions)
Ab272504, by Abcam (1 mentions)
Ab119339, by Abcam (1 mentions)
Ab46154, by Abcam (1 mentions)
Ab34710, by Abcam (1 mentions)
Eclipse te2000 s microscope, by Nikon (1 mentions)
Antigen retrieval solution, by Agilent Technologies (1 mentions)
Sz61tr tp051000 microscope, by Olympus (1 mentions)
Citrate buffer ph 6, by Abcam (1 mentions)
Igg isotype control, by Abcam (1 mentions)
Epcam, by Cell Signaling Technology (1 mentions)
Hmga2, by R&D Systems (1 mentions)
10 protocols using mouse and rabbit specific hrp dab detection ihc kit
1
Immunohistochemical Analysis of Neuroendocrine Markers
2
SARS-CoV-2 Spike Protein IHC in Placenta
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Paraffin sections were taken in 0.5 μm. Placental paraffin sections were deparaffinized under 60 °C for 20 min. After a series of washes with PBS 0.05% Tween 20 pH 7.4, slides were incubated in hydrogen peroxide for 10 min. Antigen retrieval was performed by boiling with a domestic microwave (850 W) in 1X citrate buffer pH 6.0 for 10 min. After blocking, slides were incubated at room temperature for 45 min with Anti- SARS CoV-2 spike glycoprotein (rabbit polyclonal, ab272504 Abcam) antibody at 1:400 dilution in PBS. For visualization, sections were labeled with Mouse and Rabbit Specific HRP/DAB detection IHC kit (ab64264, Abcam) and counterstained with hematoxylin. SARS-CoV-2 (B303) infected Vero E6 cells (the Ct value for Nucleocapsid-1 (N1) gene was 18,6 by qPCR) were used as a positive control and placental tissues from uninfected women were used as a negative control. Imaging was performed by microscopy.
3
Quantitative Immunohistochemistry Analysis
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Some histological sections were used for immunohistochemistry of PECAM-1, VEGF, and PCNA35 (link), 45 , 49 (link). For PECAM-1, sections were incubated with an anti-CD31 mouse monoclonal antibody (ab119339, abcam, Cambridge, UK) diluted at 1:100 in 3% BSA-PBS and incubated overnight at 4 °C. For VEGF, sections were incubated with an anti-VEGF rabbit polyclonal antibody (ab46154, abcam, Cambridge, UK) diluted at 1:100 in 1% BSA-PBS for 45 minutes. For PCNA, sections were incubated with an anti-PCNA rabbit polyclonal antibody (ab18197, abcam, Cambridge, UK) diluted at 1:4,000 in PBS for 2 hours.
All sections were incubated with a Mouse and Rabbit Specific HRP/DAB Detection IHC kit (abcam, Cambridge, UK) before counterstained with hematoxylin. Images were captured with a microscope imaging system. Images of the sections were quantified using ImageJ software (1.48 v, Wayne Rasband, National Institutes of Health, USA) with color threshold selection. The common values used for all samples in terms of Hue, Saturation, and Brightness are in the range between 50–235, 0–255, and 0–155, respectively. Once the area of the intensity with common values was selected, ImageJ calculated the area of intensity over background in percentage.
All sections were incubated with a Mouse and Rabbit Specific HRP/DAB Detection IHC kit (abcam, Cambridge, UK) before counterstained with hematoxylin. Images were captured with a microscope imaging system. Images of the sections were quantified using ImageJ software (1.48 v, Wayne Rasband, National Institutes of Health, USA) with color threshold selection. The common values used for all samples in terms of Hue, Saturation, and Brightness are in the range between 50–235, 0–255, and 0–155, respectively. Once the area of the intensity with common values was selected, ImageJ calculated the area of intensity over background in percentage.
4
Porcine Intervertebral Disc Histology
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Porcine cervical discs were fixed in 10% neutral buffered formaldehyde solution for 2 days, and subsequently immersed in a decalcification solution (0.5 M aluminium chloride, 8.5% HCl, and 5% formic acid) for 3 days of decalcification. Following decalcification, samples were prepared for histological inspection and the section specimen was then stained with hematoxylin and eosin (H&E) and picrosirius red.
For immunostaining for type I collagen, the sections were immersed in a solution of methanol with 3% H2O2 for 10 min to quench endogenous peroxidase activity and then incubated in a citrate buffer (pH 6.0) antigen retrieval solutions for 30 min at 95 °C. Following overnight incubation of the slides in type I collagen antibody (5 μg/mL overnight at 4 °C; ab34710, Abcam, Cambridge, UK), the sections were labeled with streptavidin-biotin. The presence of antigens was detected using 3,3′-diaminobenzidine tetrahydrochloride substrate (Mouse and Rabbit Specific HRP/DAB Detection IHC kit (ab64264, Abcam, Cambridge, UK)). Sections were further counterstained with hematoxylin.
For immunostaining for type I collagen, the sections were immersed in a solution of methanol with 3% H2O2 for 10 min to quench endogenous peroxidase activity and then incubated in a citrate buffer (pH 6.0) antigen retrieval solutions for 30 min at 95 °C. Following overnight incubation of the slides in type I collagen antibody (5 μg/mL overnight at 4 °C; ab34710, Abcam, Cambridge, UK), the sections were labeled with streptavidin-biotin. The presence of antigens was detected using 3,3′-diaminobenzidine tetrahydrochloride substrate (Mouse and Rabbit Specific HRP/DAB Detection IHC kit (ab64264, Abcam, Cambridge, UK)). Sections were further counterstained with hematoxylin.
5
Immunohistochemical Analysis of PCNA Expression
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Some histological sections were used for immunohistochemistry [7 (link)]. Sections were incubated with an anti-PCNA rabbit polyclonal antibody
(ab18197, abcam, Cambridge, UK ) diluted at 1/4,000 in PBS for 2 h, washed in PBS, and
incubated with a Mouse and Rabbit Specific HRP/DAB Detection IHC kit (abcam, Cambridge, UK
). Sections were counterstained with hematoxylin and then dehydrated with ethanol. Control
specimens were also stained, but PBS was substituted for the primary antibody. All section
images were captured with a microscope.
(ab18197, abcam, Cambridge, UK ) diluted at 1/4,000 in PBS for 2 h, washed in PBS, and
incubated with a Mouse and Rabbit Specific HRP/DAB Detection IHC kit (abcam, Cambridge, UK
). Sections were counterstained with hematoxylin and then dehydrated with ethanol. Control
specimens were also stained, but PBS was substituted for the primary antibody. All section
images were captured with a microscope.
6
Comprehensive Immunohistochemical Analysis
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VECs were stained using anti-von Willebrand Factor (vWF) antibody in 1 : 100 dilution. TF was stained using anti-mouse TF antibody in 1 : 100 dilution. PTX in the lungs was stained using paclitaxel antibody. The Mouse and Rabbit Specific HRP/DAB detection IHC kit (Abcam, ab64264) was used according to manufacturer's instructions. H&E staining was used for histological diagnosis.
7
Cardiac Cleaved Caspase-3 Quantification
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Immunohistochemistry was performed to assess cardiac levels of cleaved caspase-3 according to the manufacturer's instructions (Mouse and Rabbit Specific HRP/DAB Detection IHC Kit, ab64264, Abcam). Briefly, the paraffin sections of hearts were incubated with primary antibodies against cleaved caspase-3 (1:250, Cat. No. 9664, Cell Signaling Technology) at 4 °C overnight. After washing, sections were stained with Biotinylated Goat Anti-Polyvalent for 10 min at room temperature. After washing, the sections were incubated with Streptavidin Peroxidase and DAB Substrate. Stained sections were imaged with a Nikon Eclipse TE2000-S microscope and five images were captured under high power fields (100× magnifications) randomly.
8
Immunohistochemical Analysis of AdipoR1 in PVAT
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PVAT samples were fixed in 10% formalin, dehydrated, and embedded in paraffin wax. The PVAT sections were cut on a rotary microtome and used for IHC analysis using a mouse- and rabbit-specific HRP/DAB IHC detection kit (Abcam, Waltham, MA, USA). The slides were first treated with antigen retrieval solution (Dako, Glostrup, Denmark), followed by hydrogen peroxide to inhibit endogenous peroxidase. The samples were then incubated with a protein-blocking serum to prevent excessive background staining. To analyze the distribution of protein, the slides were incubated with a mouse monoclonal antibody against adipoR1 at a 1:500 dilution (Santa Cruz Biotechnology, Dallas, TX, USA). The slides were then incubated with micropolymer secondary antibody, followed by DAB substrate. Subsequently, the slides were counterstained with hematoxylin and sequentially dehydrated. The negative control slides were stained, without incubation, with adipoR1 primary antibody. The sections were photographed using an Olympus SZ61TR-TP051000 microscope (Olympus, Tokyo, Japan), and the images were analyzed using ImageJ software version 8 (ImageJ Software, National Institute of Health, Bethesda, MD, USA).
9
Immunohistochemical Analysis of Tissue Samples
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Upon receipt of fresh tissue, a piece was removed and immediately fixed in formalin. The fixed tissue was processed and embedded in paraffin. For histology staining, 5 µm sections were mounted onto glass slides. Rehydration and antigen retrieval were performed using citrate buffer, pH 6.0 (Abcam, Cambridge, UK). Slides were stained with Anti-pan Cytokeratin AE1/AE3 + 5D3 (Abcam, Cambridge, UK) or IgG Isotype control (Abcam, Cambridge, UK) for 2 hours at room temperature. Staining was detected using Mouse and Rabbit Specific HRP/DAB IHC Detection kit (Abcam, Cambridge, UK). Images were taken on an Olympus IX70 microscope with a Jenoptik (Jena, Germany) ProgRes C14plus camera and ProgRes CapturePro software. Brightfield spheroid images were taken on the same microscope with the same camera and software.
10
Immunohistochemical Analysis of Cell Markers
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Samples were fixed in 3.7% paraformaldehyde pH 7.4 (cat. no. P2031; Biosesang, Inc.) overnight at 4˚C, dehydrated using a series of ethanol solutions of increasing concentrations (50% ethanol, 70% ethanol and 100% ethanol), and embedded in paraffin. Tissue sections 4 µm thick were incubated in Mayer's hematoxylin solution (Lillie's Modification) for 5 min and eosin Y solution (modified alcoholic) for 3 min at 25˚C using hematoxylin and eosin (H&E) staining kit (cat. no. ab245880; Abcam), and mouse and rabbit specific HRP/DAB IHC detection kit (cat. no. ab236466; Abcam, the avidin-biotin-peroxidase complex (ABC) method according to the manufacturer's protocol. Cell markers, including STRO-1 (cat. no. MAB1038-SP; 1:100; R&D Systems, Inc.), high mobility group AT-hook 2 (HMGA-2; cat. no. 8179S; 1:400; Cell Signaling Technology, Inc.), epithelial cell adhesion molecule (EpCAM; cat. no. 2929S; 1:500; Cell Signaling Technology, Inc.) and fibroblast-specific protein-1 (FSP-1; 13018S; 1:400; Cell Signaling Technology, Inc.) were used as the primary antibodies.
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