Animals were euthanized at 7 days after elastase infusion, and infused and non-infused aortas were collected, fixed, paraffin-embedded, and then cut to prepare 5-μm sections. We performed essentially the same immunohistochemistry procedures as included in our recent report [35 (link)]. Briefly, slides were deparaffinized and rehydrated, and then subjected to antigen retrieval in citrate buffer at 80 °C in a high-pressure cooker. A primary antibody (see antibodies listed in Table S2 ) and ImmPRESS HRP (horse radish peroxidase) Anti-Rabbit IgG Polymer Detection Kit (Vector Laboratories, MP-7451-15) were used to visualize the protein of interest. Images were taken under an EVOS microscope (Thermo Fisher Scientific).
Anti rabbit igg polymer detection kit
Manufactured by Vector Laboratories
Sourced in United States
The Anti-rabbit IgG Polymer Detection Kit is a laboratory reagent designed to facilitate the visualization and detection of rabbit immunoglobulin G (IgG) in immunohistochemical and other immunological applications. The kit includes a polymer-based detection system that enhances the signal intensity, providing improved sensitivity and clarity in the detection of target antigens.
Automatically generated - may contain errors
Lab products found in correlation
Sk 4105, by Vector Laboratories (5 mentions)
S 2012, by Vector Laboratories (4 mentions)
Dako autostainer, by Agilent Technologies (4 mentions)
Evos microscope, by Thermo Fisher Scientific (4 mentions)
Aperio scanscope slide scanner, by Leica (1 mentions)
Nanozoomer s210 digital slide scanner, by Hamamatsu Photonics (1 mentions)
Cd8 antibody, by Agilent Technologies (1 mentions)
Dab substrate chromagen, by Agilent Technologies (1 mentions)
Neutral buffered formalin, by CellPath (1 mentions)
Tris edta ph 9, by Agilent Technologies (1 mentions)
Dab map detection kit, by Roche (1 mentions)
8 protocols using anti rabbit igg polymer detection kit
1
Immunohistochemistry of Aortic Tissues
2
Immunohistochemical Analysis of Aortic Tissue
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Animals were euthanized at 7 days after elastase infusion, and infused and non-infused aortas were collected, fixed, paraffin-embedded, and then cut to prepare 5-μm sections. We performed essentially the same immunohistochemistry procedures as included in our recent report [35 (link)]. Briefly, slides were deparaffinized and rehydrated, and then subjected to antigen retrieval in citrate buffer at 80 °C in a high-pressure cooker. A primary antibody (see antibodies listed in Table S2 ) and ImmPRESS HRP (horse radish peroxidase) Anti-Rabbit IgG Polymer Detection Kit (Vector Laboratories, MP-7451-15) were used to visualize the protein of interest. Images were taken under an EVOS microscope (Thermo Fisher Scientific).
3
Immunohistochemical Analysis of Ucp1 in Adipose Tissue
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Adipose tissues were fixed in 10% formalin for 24 h at room temperature. Then, the tissues were embedded into paraffin and cut into 4 μm thick slices, deparaffinized, and rehydrated using xylene, ethanol, and water by standard methods. Immunohistochemistry was performed on a Dako Autostainer (Dako). Slides were incubated with 3% hydrogen peroxide and 2.5% normal horse serum (S-2012, Vector), followed by incubation with rabbit polyclonal anti-Ucp1 primary antibody diluted 1:200 in 2.5% normal horse serum (S-2012, Vector) for 60 min. Signals were detected with an anti-rabbit IgG Polymer Detection Kit (MP-7401, Vector). Labeling was visualized with 3,3′-diaminobenzidine (DAB) as the chromogen (SK-4105, Vector). Slides were counterstained with Harris hematoxylin (EK Industries), and whole-slide digital images were collected with an Aperio Scan Scope slide scanner (Aperio).
4
Collagen I Immunohistochemistry Protocol
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Immunohistochemistry was performed on a Dako Autostainer (Dako, Carpinteria, CA, USA). Slides were incubated with 3% hydrogen peroxide and 2.5% normal horse serum (S-2012, Vector Laboratories, Burlingame, CA, USA), followed by incubation with rabbit polyclonal anti-collagen I primary antibody diluted 1:200 in 2.5% normal horse serum for 60 min. Signals were detected with an anti-rabbit IgG Polymer Detection Kit (MP-7401, Vector Laboratories, Burlingame, CA, USA). Labeling was visualized with 3,3′-diaminobenzidine as the chromogen (SK-4105, Vector Laboratories, Burlingame, CA, USA). Slides were counterstained with Harris hematoxylin (EK Industries, Joliet, IL, USA), and whole-slide digital images were collected at 400× magnification with an Aperio Scan Scope slide scanner (Aperio, Vista, CA, USA).
5
Immunohistochemical Analysis of UCP1 in Adipose Tissues
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Adipose tissues were fixed in 10% formalin for 24 hours at room temperature. Then the tissues were embedded into paraffin and cut at 4 μm thick, de-paraffinized, and rehydrated through xylene, ethanol, and water by standard methods. For antigen retrieval, slides were submerged in 0.01 M sodium citrate (pH 6.0) and heated to 96 °C for 20 minutes in a laboratory microwave (PELCO). Immunohistochemistry was performed on a Dako Autostainer (Dako, Carpinteria, CA). Slides were incubated with 3% hydrogen peroxide and 2.5% normal horse serum (S-2012, Vector), followed by incubation with rabbit polyclonal anti-UCP1 primary antibody diluted 1:200 in 2.5% normal horse serum (Vector, S-2012) for 60 minutes. Signal was detected with an anti-rabbit IgG Polymer Detection Kit (MP-7401, Vector) Labeling and was visualized with 3, 3′-diaminobenzidine (DAB) as the chromogen (SK-4105, Vector). Slides were counterstained with Harris hematoxylin (EK Industries, Joliet, IL) and whole slide digital images were collected at 20X magnification with an Aperio Scan Scope slide scanner (Aperio, Vista, CA). Images shown were representative result of at least three biological replicates. Scanned images of H&E staining were analyzed by Photoshop CS3 to calculate cell numbers. Averaged adipocyte size was calculated as the image area divided by cell numbers.
6
Immunohistochemical Detection of UCP1
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Immunohistochemistry was performed on a Dako Autostainer (Agilent, Santa Clara, CA, USA). Slides were incubated with 3% hydrogen peroxide and 2.5% normal horse serum (S-2012;Vector Laboratories, Burlingame, CA, USA) followed by incubation with rabbit polyclonal anti-UCP1 primary antibody diluted 1:200 in 2.5% normal horse serum (S-2012; Vector Laboratories) for 60 min. Signals were detected with an anti-rabbit IgG Polymer Detection Kit (MP-7401; Vector Laboratories). Labeling was visualized with 3,3′-diaminobenzidine as the chromogen (SK-4105; Vector Laboratories). Slides were counterstained with Harris hematoxylin (EK Industries, Joliet, IL, USA), and whole-slide digital images were collected at × 20 magnification with an Aperio Scan Scope Slide Scanner (Leica Biosystems).
7
Immunohistochemical Analysis of UCP1 in Adipose Tissues
8
Quantifying Tumor mCherry Expression in Mice
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Tumor from mice were harvested and placed into 10% neutral buffered formalin (Cellpath). mCherry staining and quantification were performed by UCL Pathology. In brief, deparaffinized hydrated tissue sections underwent antigen unmasking in Tris–EDTA pH 9 (Dako) at high pressure in a pressure cooker for 8 min. After washing and quenching, sections were blocked in 2.5% Horse Serum (Vector ImmPRESS Kit) for 20 min at room temperature. Incubation with primary antibody anti-mCherry (Abcam, ab167453, 1 µg/ml) was for 60 min at room temperature, secondary antibody (anti-Rabbit IgG Polymer Detection Kit, MP-7401, Vector Laboratories) for 30 min at room temperature and DAB + substrate/chromagen (Dako) for 5 min at room temperature prior to counterstaining and mounting.
Slides were scanned in the Hamamatsu NanoZoomer S210 Digital slide scanner. The image analysis has been performed on the whole section with the positive cell counting algorithm from QuPath image analysis software.
Fixation, embedding and CD8 staining were performed at the UCL Institute of Neurology, using the Ventana Discovery XT instrument and Ventana DAB Map detection Kit (760-124). For pre-treatment, Ventana CC1 (950-124) was used. The CD8 antibody (Dako M7103) was used at 1:100 dilution for 1 h before Rabbit anti-murine secondary antibody (Dako E0354) was used.
Slides were scanned in the Hamamatsu NanoZoomer S210 Digital slide scanner. The image analysis has been performed on the whole section with the positive cell counting algorithm from QuPath image analysis software.
Fixation, embedding and CD8 staining were performed at the UCL Institute of Neurology, using the Ventana Discovery XT instrument and Ventana DAB Map detection Kit (760-124). For pre-treatment, Ventana CC1 (950-124) was used. The CD8 antibody (Dako M7103) was used at 1:100 dilution for 1 h before Rabbit anti-murine secondary antibody (Dako E0354) was used.
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