Monoclonal antibody against human: CD31
Monoclonal antibody against human CD31 (The cluster of differentiation 31 (CD31) also known Platelet endothelial cell adhesion molecule (PECAM-1) or Glycoprotein IIa (GPIIa).
Product Number: P5000PU
Concentration: 0.5 mg/ml.
Storage Buffer: Phosphate buffered saline, pH 7.2 with 0.05% sodium azide.
Immunogen: Biochemically characterized crude subcellular/membrane fraction obtained from HUVEC cells.
Isotype: Mouse IgG1, Kappa
Reactivity: The antibody was tested against normal and human tumor tissue and HUVEC cells. The antibody was also tested against mouse and rat tissues. Staining of vasculature is observed in all tissues examined. The HUVEC cells showed cell surface and cytoplasmic staining. Not tested on formalin fixed, paraffin embedded (FFPE) tissue.
Applications: Indirect immunofluorescence, immunohistochemistry & western blotting. Suggested concentrations are as follows; Western blotting 2-3ug/ml, immunohistochemistry 3-5ug/ml. The BC16-6.4EF antibody may be used on endothelial cells such as HUVEC cells grown on chamber slides, cytospins and cryosections of human tissue. For staining, the cultured cells and cryosections should be fixed in 1-2% paraformaldehyde for 30 mins, permeabilized in 0.25% Triton X 100 in PBS for 30 mins and non-specific binding blocked with 1% BSA in PBS. The primary antibody may be diluted in PBS with 1% BSA and incubated on cells/tissue overnight at 4C. The antibody may also be used to inhibit endothelial cell–cell interactions.
Western blotting-: Use the BC16-6.4EF antibody at 2-3 ug/ml in Tris buffered Saline with 0.05% Tween 20 and 5% non-fat dry milk (Blotto) or similar diluents.
Immunostaining: Use the antibody at 2-3ug/ml diluted with PBS containing 1% BSA.
Human umbilical vein endothelial cells (HUVEC) stained with 5 ug/ml BC16-6.4EF anti CD31 antibody diluted in PBS with 1% BSA. FITC conjugated rabbit anti mouse Ig secondary.
Suggested positive control cells and tissues: For staining and western blotting, human umbilical endothelial cells (Thermo Fisher) are suggested as positive control cells. The antibody reacts with a band of approximate molecular size 135kDa. For immunohistochemical staining, cryosections of most human tissue with vasculature may be used.
Preparation and Storage: The monoclonal antibody is purified from tissue culture supernatant using Protein G sepharose columns. Store undiluted at 4° C or aliquot and store at -20C for long term storage. Repeated freeze thaw cycles should be avoided.
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- Caution: The product contains 0.05% sodium azide. Sodium azide produces highly toxic hydrazoic acid under acidic conditions. It is therefore recommended to dilute azide compounds in running water before discarding to prevent the accumulation of potentially explosive deposits in the plumbing.
Platelet endothelial cell adhesion molecule-1 (PECAM-1) or cluster designation 31 (CD31) is a member of the immunoglobulin (Ig) gene superfamily and a transmembrane glycoprotein. The CD31 protein is widely expressed by cells of the vascular endothelium lineage, platelets, Kupffer cells, granulocytes, megakaryocytes, monocytes, neutrophils, and some types of T-cells. CD31 functions as a cell adhesion molecule in endothelial cell homotypic interactions in the process of angiogenesis. CD31 is also involved in leukocyte migration and integrin activation. It plays a key role in the adhesion cascade between the endothelial cells and inflammatory cells, enabling leukocyte migration in inflammatory sites. CD31 is one of the best markers for benign and malignant vascular tumors. It is expressed in certain tumors including angiomas and angiosarcomas including epitheloid hemangioendothelioma, epitheloid sarcoma-like hemangioendothelioma. Its expression in Kaposi sarcoma lesions is variable. Human CD31 comprises 738 amino acids and its predicted molecular weight would therefore be approximately 81kDa. However the observed molecular weight, as reported by various research groups including our own, is higher and at around 120kDa and 140kDa. The disparity probably reflects post translational modifications including glycosylation and probably because different isoforms of the protein may exist.
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