Analysis Affinity of M6495 for Human ADAMTS-5 by KinExA (CAT#: STEM-MB-0039-CJ)

Introduction

Healthy cartilage homeostasis depends on intact collagen scaffolds and high polymerin content. ADAMTS-5 is a spondin-motif-5 decomposin and metalloproteinase that is closely associated with arthritic diseases in the cleavage of polymeric proteins. Inhibition of ADAMTS-5 may prevent the development of osteoarthritis (OA). M6495 is a highly specific ADAMTS-5 inhibitor that has been shown to inhibit polymeric protein transformation in cartilage explants. In addition, M6495 interferes with signaling cross-talk between cartilage and synovium by reducing the release of GAG, type II collagen (C2M) and type III collagen (C3M). At present, M6495 is used in the clinical development of OA.

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Principle Applications Procedure Materials Notes Related Products

Principle

KinExA is a two-stage analysis system. In the first stage, a number of solutions are prepared, where one partner remains constant (constant binding partner, or CBP) and the other (titrant) is variable, usually serially diluted. As the titrant is added, the free CBP decreases and is analysed by a sophisticated and precise microfluorescence measurement device. The signal generated can be mathematically related to the affinity (KD) of the two molecules for each other, as well as the kinetic parameters of binding (kon) and dissociation (koff).

Applications

Immunology/Inflammation, Toxicology; Pharmacology

Procedure

1. preparation of the functionalized beads which will capture the analyte for measuremen.
2. preparation of a series of solutions consisting of a constant initial concentration of one component of the binary reaction and serial dilutions of the other reactant. The component that is kept constant is the constant binding partner (CBP) , and is the one which will be analyzed.
3. each reaction mixture is sampled and the fluorescence of free CBP bound to the capture beads is obtained for subsequent numerical analysis.

Materials

• Sample Type: Antibodies, Cells, Small Molecules, Proteins, DNA, Lipids, Viruses, Serum, & More
• Equipment: Kinetic Exclusion Assay (KinExA)

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