Heparan degrading enzyme detection
Heparan sulfate is a complex and ubiquitous polysaccharide that is a component of mammalian cell membranes. It has been identified in mammalian liver, spleen, skin, placenta, and platelets as well as in cancerous tissues such as melanomas, lymphomas, sarcomas, fibrosarcomas, and colon cancer. Heparan sulfate has additionally been found to play an important defensive role in tumor cell invasion. The activity of heparan degrading enzyme (heparanase), an enzyme that cleaves heparan sulfate, is reported to be considerably higher in invasive cancer than normal cells. Thus, the correlation of heparan sulfate degrading enzyme activity with cancer malignancy is of much research interest.
Heparan sulfate is a complex and ubiquitous polysaccharide that is a component of mammalian cell membranes. It has been identified in mammalian liver, spleen, skin, placenta, and platelets as well as in cancerous tissues such as melanomas, lymphomas, sarcomas, fibrosarcomas, and colon cancer. Heparan sulfate has additionally been found to play an important defensive role in tumor cell invasion. The activity of heparan degrading enzyme (heparanase), an enzyme that cleaves heparan sulfate, is reported to be considerably higher in invasive cancer than normal cells. Thus, the correlation of heparan sulfate degrading enzyme activity with cancer malignancy is of much research interest.
This Heparan Degrading Enzyme Assay Kit provides a 96-well non-radioactive assay format for the measurement of heparan sulfate degradation in cultured cells, tissues, or serum, as well as for the screening of heparan sulfate degradation enzyme inhibitors. This kit is based on the ability of heparin-like molecules to bind bFGF (basic fibroblast growth factor).
When heparan sulfate is degraded, it loses its ability to bind bFGF. Thus, the enzymatic activity of a sample can be quantitated by comparing the amount of bFGF-bound undegraded heparan sulfate with control heparanase-free total-bound heparan sulfate. A unique feature of the Heparan Degrading Enzyme ELISA is the use of CBD-FGF, a fusion protein composed of the human fibronectin cell-binding domain and human fibroblast growth factor. CBD-FGF is immobilized on the surface of microtiter ELISA plates by an anti-fibronectin antibody that binds the CBD epitope. Termed DOC (Domain Oriented Capture), this solid phase attachment method enables the formation of a natural 3D bFGF structure for enhanced accessibility to and binding of heparan sulfate. Detection occurs via a colorimetric assay method that uses biotinylated heparan sulfate as the substrate.
Overview
- Safe: No radioactive isotopes required
- 96-well format allows simultaneous assay of many samples
- Any tissue, cell, or serum, regardless of mammalian species, can be used
- Enables interaction studies of bFGF to heparin-like substances and other ligands
- Unique solid phase attachment method creates a natural 3D bFGF structure for enhanced accessibility and binding to heparan sulfate
More Information
Applications
- Quantitative measurement of heparan sulfate degradation in mammals
- Screening of heparan sulfate degradation enzyme inhibitors
- Heparan degrading enzyme ELISA
Product citations
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Hashi, H., Hatai, M., Kimizuka, F., Kato, I. & Yaoi, Y. Angiogenic activity of a fusion protein of the cell-binding domain of fibronectin and basic fibroblast growth factor. Cell Struct. Funct. 19, 37–47 (1994).
He, X. et al. Hypoxia increases heparanase-dependent tumor cell invasion, which can be inhibited by antiheparanase antibodies. Cancer Res. 64, 3928–33 (2004).
Katayama, M. et al. Isolation and characterization of two monoclonal antibodies that recognize remote epitopes on the cell-binding domain of human fibronectin. Exp. Cell Res. 185, 229–36 (1989).
Nakajima, M. et al. Suramin. A potent inhibitor of melanoma heparanase and invasion. J. Biol. Chem. 266, 9661–6 (1991).
Nakajima, M., Irimura, T., Di Ferrante, D., Di Ferrante, N. & Nicolson, G. L. Heparan sulfate degradation: relation to tumor invasive and metastatic properties of mouse B16 melanoma sublines. Science 220, 611–3 (1983).
Nakajima, M., Irimura, T., Di Ferrante, N. & Nicolson, G. L. Metastatic melanoma cell heparanase. Characterization of heparan sulfate degradation fragments produced by B16 melanoma endoglucuronidase. J. Biol. Chem. 259, 2283–90 (1984).
Nicolson, G. L. et al. Cancer cell heparanase activity associated with invasion and metastasis. Adv. Enzyme Regul. 38, 19–32 (1998).
Ricoveri, W. & Cappelletti, R. Heparan sulfate endoglycosidase and metastatic potential in murine fibrosarcoma and melanoma. Cancer Res. 46, 3855–61 (1986).
Addtional product information
Please see the product's Certificate of Analysis for information about storage conditions, product components, and technical specifications. Please see the Kit Components List to determine kit components. Certificates of Analysis and Kit Components Lists are located under the Documents tab.
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