OverviewData/SpecificationsLiterature/SupportHow It WorksRelated ProductsOverviewOverview:Aggrecanases ADAMTS1, ADAMTS4 and ADAMTS5 are members of the ADAMTS-family of proteases (ADAMTS = A disintegrin and metalloprotease with thrombospondin motif). They aresynthesized as multidomain proteins consisting of prodomains, protease, disintegrin, thrombospondin and cys-rich domains followed by spacer regions and additional thrombospondin motifs. The prodomains are cleaved-off intracellularly and the enzymes are processed further from their C-terminal ends after secretion.Aggrecanases cleave specific peptide bonds in aggrecan, the main proteoglycan of articular cartilage. They hydrolyze also other lecticanes as versican and brevican. Convincing evidence indicates that aggrecanases are mainly responsible for excessive aggrecan degradation in osteoarthritic joints.Data/SpecificationsData/Specifications:Species: humanSample Type: cell culture supernatesSample Size: 100 uLStandard Curve Range: 0.062 - 4 nMSensitivity: 0.025 nMAssay Length:3.75 hoursLiterature/SupportLiterature/Support/Publications:Product Insert:Aggrecanase Activity Assay Insert (PDF)Articles/FAQ:ELISA Troubleshooting GuideELISA Data Reduction GuideMeasuring aggrecanase activity to identify potential therapeutics for osteoarthritis. (Blog Post)How It WorksHow It Works:The Aggrecanase Activity Assay measures activity of aggrecanases. It is used to screen and characterize aggrecanase inhibitors. This assay consists of two modules, the Aggrecanase Module and the ELISA Module. A recombinant fragment of human aggrecan interglobular domain (aggrecan-IGD) is first digested with ag- grecanase. Proteolytic cleavage of the substrate releases an aggrecan peptide with the N-terminal sequence ARGSVIL (ARGSVIL-peptide). The ARGSVIL-peptide is then quantified with two monoclonal anti-peptide antibodies.Aggrecanase module:Proteolysis of aggrecan-IGD by aggrecanase Aggrecan-IGD is incubated with standard aggrecanase and samples of unknown aggrecanase activity. In addition to aggrecanase, different concentrations of aggrecanase inhibitors can be added. The reaction is then stopped by dilution with EDTA-containing buffer.ELISA module: Aggrecan peptide ELISA ARGSVIL-peptide standard, proteolytic digestions of aggrecan-IGD with standard aggrecanase and samples are incubated in microtiter wells precoated with anti-ARGSVIL-neoepitope antibody. ARGSVIL-peptide is bound to the coated antibody, while other components are removed by washing and aspiration. The bound ARGSVIL-peptide is detected with a second peroxidase-labeled antibody. Any excess of the conjugate is removed by washing and aspiration. The amounts of peroxidase bound to different wells are determined in reactions with peroxidase substrate TMB. The reactions are stopped by addition of sulfuric acid solution and absorbance is read at 450 nm in a microtiter plate spectrophotometer.Related ProductsRelated Products:Aggrecan Antibody, C-terminal neoepitope NITEGEAggrecan Antibody, N-terminal neoepitope ARGAggrecan Antibody, N-terminal neoepitope DIPENAggrecan Antibody, N-terminal neoepitope FFGVAggrecan IGD AntibodyReverse Related Products:Aggrecanase Activity, Sensitive ELISAMMP-13 (activated) ELISA