Firstly, the sections were deparaffinized and dehydrated using xylene and were placed in running water for 10 minutes to ensure thorough removal of xylene. 1X repair solution was applied to the sections, which were then heated in a pressure cooker at 121°C for 2-3 minutes. Subsequently, a 3% hydrogen peroxide solution was incubated with the sections in a light-protected 37°C incubator for 20 minutes to eliminate endogenous peroxidase activity. Blocking solution (composed of 0.2% PBSTriton X-100, 1% bovine serum albumin and 10% normal donkey serum) was added to the samples, and they were incubated at room temperature for 45-60 minutes to reduce nonspecific binding. Additionally, the samples were incubated overnight at 4°C for antibody binding by adding a CD31 antibody (diluted to 1:200, Cell Signaling Technology, Cat.77699S). Finally, DAPI staining solution (Solarbio, C0065) was added for nuclear staining, and the results were observed using a fluorescence microscope.
Cd31 antibody
Manufactured by Cell Signaling Technology
Sourced in United States
The CD31 antibody is a laboratory reagent used to detect the presence of the CD31 protein in biological samples. CD31, also known as PECAM-1, is a cell adhesion molecule expressed on the surface of endothelial cells, platelets, and some types of leukocytes. The CD31 antibody can be used to identify and study these cell types in various research and diagnostic applications.
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Lab products found in correlation
Dapi staining solution, by Solarbio (1 mentions)
Cd68 antibody, by Cell Signaling Technology (1 mentions)
Active caspase 3 antibody, by Cell Signaling Technology (1 mentions)
Pierce ecl western blotting substrate, by Thermo Fisher Scientific (1 mentions)
Image lab software, by Bio-Rad (1 mentions)
Chemidoc mp, by Bio-Rad (1 mentions)
Sds page, by Bio-Rad (1 mentions)
3 protocols using cd31 antibody
1
Immunohistochemical Analysis of CD31 Expression
2
Immunohistochemical Analysis of Heart Tissue
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Heart slices from the four categories were also subject to IHC analysis. Briefly, the slides were incubated in 3% H2O2 solution for 10 min and antigen retrieval was performed by steam heating in 10 mM citrate buffer (pH 6.0) for 30 min. After epitope recovery, the slides were then treated with 10% of normal goat serum for 60 min, followed by incubation with active caspase 3 antibody (1:500, Cell Signaling Technology, Catlog #9664, MA, USA), CD31 antibody (1:1000, Cell Signaling Technology, Catlog #3528), and CD68 antibody (1:500, Cell Signaling Technology, Catlog #76437) incubation overnight at 4 °C. The slides were washed and incubated with secondary horseradish peroxide (HRP)-linked secondary antibody (Vector Laboratories, MN, USA) at 1:500 dilutions for 1 h. The samples were treated with the chromogen DAB for antigen detection and the final counterstaining was performed with hematoxylin.
3
Angiogenesis Protein Expression in HUVECs
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The expressions of angiogenesisrelated proteins were assessed using western blot analysis. HUVECs were harvested with lysis buffer after incubation with Cu-DIO scaffold extract and pure medium for 3 days, respectively. Protein concentrations were measured using a bicinchoninic acid (BCA) reagent. The total protein (50 μg) was separated using the SDS-polyacrylamide gel electrophoresis gel (SDS-PAGE, BIO-RAD, United States) and transferred to a polyvinylidene fluoride (PVDF) membrane. The membrane was incubated overnight at 4 o C with platelet endothelial cell adhesion molecule-1 (CD31) antibody (1 : 1000, Cell Signaling, Massachusetts, USA) and further the relevant secondary antibody (goat antimouse, 1 : 100000, Abcam, Cambridge, UK). The immunoblotting was also performed for -Actin serving as an internal loading control. The protein bands were visualized using the enhanced chemiluminescence substrate kit (Pierce ECL western blotting substrate, Thermo Scientific, USA) and captured with a BIO-RAD imaging system (ChemiDoc MP, United States). The result was further analyzed using Image Lab software (BIO-RAD).
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