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Monoclonal antibody against human: CD31

Monoclonal antibody against human: CD31

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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 Information

 

Product Number:                    P5000PU

 

Size:                                        100ug

 

Concentration:                       0.5 mg/ml.

 

Storage Buffer:                       Phosphate buffered saline, pH 7.2 with 0.05% sodium azide.

 

Clone:                                     BC16-6.4EF

 

Host:                                       Mouse

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.

 

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. 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.

 

 

Background:

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.

 

References:

Hsu-Lin S, Berman CL, Furie BC, August D, Furie B. A platelet membrane protein expressed during platelet activation and secretion. Studies using a monoclonal antibody specific for thrombin-activated platelets. J Biol Chem. 259(14):9121-6 (1984).

van Mourik JA, Leeksma OC, Reinders JH, de Groot PG, Zandbergen-Spaargaren J. Vascular endothelial cells synthesize a plasma membrane protein indistinguishable from the platelet membrane glycoprotein IIa. J Biol Chem. 260(20):11300-6 (1985).

Newman,P.J., Berndt,M.C., Gorski,J., White,G.C. II, Lyman,S., Paddock,C. and Muller,W.A. PECAM-1 (CD31) cloning and relation to adhesion molecules of the immunoglobulin gene superfamily. Science 247 (4947), 1219-1222 (1990).

Simmons,D.L., Walker,C., Power,C. and Pigott,R. Molecular cloning of CD31, a putative intercellular adhesion molecule closely related to carcinoembryonic antigen. J. Exp. Med. 171 (6), 2147-2152 (1990).

Stockinger,H., Gadd,S.J., Eher,R., Majdic,O., Schreiber,W., Kasinrerk,W., Strass,B., Schnabl,E. and Knapp,W. Molecular characterization and functional analysis of the leukocyte surface protein CD31 J. Immunol. 145 (11), 3889-3897 (1990).

Albelda, S.M., Oliver, PD., Romer, LH., Buck CA. EndoCAM: A novel endothelial cell–cell adhesion molecule. J. Cell Biol., 110, 1227–1237. (1990).

Albelda,S.M., Muller,W.A., Buck,C.A. and Newman,P.J. Molecular and cellular properties of PECAM-1 (endoCAM/CD31): a novel vascular cell-cell adhesion molecule. J. Cell Biol. 114 (5), 1059-1068 (1991).

Muller, W.A., Weigl, S.A., Deng, X., Phillips DM. PECAM-1 is required for transendothelial migration of leukocytes. J. Exp. Med., 178, 449–460. (1993).

 

Vaporciyan, A.A., DeLisser, H.M., Yan, H.C., Mendiguren I.I., Thom S.R., Jones M.L., Ward P.A., Albelda S.M. Platelet-endothelial cell adhesion molecule-1 (PECAM-1) is involved in neutrophil recruitment in vivo. Science, 262, 1580–1582. (1993).

 

Kirschbaum,N.E., Gumina,R.J. and Newman,P.J. Organization of the gene for human platelet/endothelial cell adhesion molecule-1 shows alternatively spliced isoforms and a functionally complex cytoplasmic domain. Blood 84 (12), 4028-4037 (1994).

 

Miettinen M, Lindenmayer AE, Chaubal A. Endothelial cell markers CD31, CD34, and BNH9 antibody to H- and Y-antigens--evaluation of their specificity and sensitivity in the diagnosis of vascular tumors and comparison with von Willebrand factor. Mod Pathol. 7(1):82-90 (1994).

 

Pantanowitz,L., Moses,A.V. and Fruh,K. CD31 immunohistochemical staining in Kaposi Sarcoma. Arch. Pathol. Lab. Med. 136 (11), 1329 (2012).

 

  


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