ab76533 staining CD31 in human kidney tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and antigen retrieval was by heat mediation in a EDTA buffer. Samples were incubated with primary antibody at a dilution of 1/100. A goat anti-rabbit IgG H&L (HRP) ab97051 was used as the secondary antibody at a dilution of 1/500.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab76533).

Anti-CD31 antibody [EPR3094] (ab76533) at 1/10000 dilution + THP-1 (Human monocytic leukemia cell line) whole cell lysate at 20 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 83 kDa

Blocking and diluting buffer: 5% NFDM /TBST 

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab76533).

Immunohistochemical analysis of endothelial colony forming progenitor cell plugs, staining CD31 (green) with ab76533.

Following antigen retrieval and blocking, sections were incubated with primary antibody (1/1000) overnight at 4°C. A Cy5®-conjugated anti-rabbit IgG (2 µg/ml) was used as the secondary antibody.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab76533).

 

All lanes : Anti-CD31 antibody [EPR3094] (ab76533) at 1/20000 dilution

Lane 1 : THP-1 cell lysate
Lane 2 : Jurkat cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : HRP labelled goat anti-rabbit at 1/2000 dilution

Predicted band size: 83 kDa

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab76533).

Clone EPR3094 (ab207090) has been successfully conjugated by Abcam. This image was generated using Anti-CD31 antibody [EPR3094] (Alexa Fluor® 647). Please refer to ab218582 for protocol details.

IHC image of CD31 staining in a section of formalin-fixed paraffin-embedded normal human kidney*.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6) in a Biocare Medical NxGen pressure cooker using retrieval settings of 110^oC for 8 minutes. Non-specific protein-protein interactions were then blocked in TBS containing 0.025% (v/v) Triton X-100, 0.3M (w/v) glycine and 1% (w/v) BSA for 1h at room temperature.  The section was then incubated overnight at +4°C in TBS containing 0.025% (v/v) Triton X-100 and 1% (w/v) BSA with ab218582 at 1/100 dilution (shown in red) and counterstained using ab195887, Mouse monoclonal to alpha Tubulin (Alexa Fluor^® 488), at 1/250 dilution (shown in green). Nuclear DNA was labelled with DAPI (shown in blue). The section was then mounted using Fluoromount^®.

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

For other IHC staining systems (automated and non-automated), customers should optimize variable parameters such as antigen retrieval conditions, antibody concentrations and incubation times.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre.

Immunohistochemical analysis of paraffin embedded human kidney tissue using ab76533 at a 1/250 dilution. Note positive staining of endothelial cells.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab76533).

Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin embedded human muscle tissue using ab76533 at a 1/250 dilution. Note positive staining of endothelial cells.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab76533).

Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.

ab76533 staining CD31 in human muscle tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with formaldehyde and a heat mediated antigen retrieval step was performed using citrate buffer pH 6.0. Samples were then blocked with 3% serum for 30 minutes at 20°C followed by incubation with the primary antibody at a 1/200 dilution for 12 hours at20°C. A Cy3®-conjugated goat anti-rabbit polyclonal was used as secondary antibody at a 1/200 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab76533).

Equilibrium disassociation constant (K_D) measurement to determine antibody affinity to the target antigen.

Click here to learn more about K_D

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab76533).