It is a complete assay system designed to screen inhibitors of neutrophil elastase, providing enough material to perform at least 96 assays.
Neutrophil elastase, also known as leukocyte elastase (LE), is a serine protease found in polymorphonuclear neutrophils (PMNs) that functions in inflammation. It can cleave a variety of proteins, including fibrous elastin, collagen, clotting factors, TNF-α, and α1-antichymotrypsin. It has been shown to be taken up by tumor cells and accelerate tumor growth through its degradation of insulin receptor substrate-1 (IRS-1). Because of its potential roles in cancer and inflammatory diseases such as emphysema, cystic fibrosis, and atherosclerosis, it is a target for drug discovery. The kit uses the chromogenic substrate MeOSuc-Ala-Ala-Pro-Val-pNA; cleavage of p-nitroaniline (pNA) from the substrate increases absorbance at 405nm. A neutrophil elastase inhibitor, elastatinal, is included for use as a control.
Colorimetric detection, HTS
Neutrophil elastase (human), (purified)pNA substrate (MeOSuc-AAPV-pNA) (60μL; 20mM in DMSO)pNA calibration standard (p-nitroaniline) (1ml; 50μM in assay buffer)Elastatinal (inhibitor) (10μL; 20mM in DMSO)Neutrophil elastase assay buffer (20ml; 100mM HEPES, pH 7.25, 500mM NaCl, 0.05% Tween-20; liquid in a screw-cap plastic bottle)96-well microplate