Neutrophil elastase (NE), also known as polymorphonuclear leukocyte elastase, is a major protease in the primary granules of neutrophils, is involved in microbicidal activity. It is located primarily in the azurophil granules of polymorphonuclear leukocytes. NE is an important factor promoting inflammation, has bactericidal effects, and shortens the inflammatory process. NE also regulates tumor growth by promoting metastasis and tumor microenvironment remodeling. However, NE plays a role in killing tumors under certain conditions and promotes other diseases such as pulmonary ventilation dysfunction. Additionally, it plays a complex role in various physiological processes and mediates several diseases. Myeloperoxidase (MPO) has been identified as an interactor of NE, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human NE and recombinant rat MPO. Briefly, biotin-linked NE were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to MPO-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST 3 times and incubation with Streptavidin-HRP for 30min, then wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50 µl stop solution to the wells and read at 450 nm immediately. The binding activity of recombinant human NE and recombinant rat MPO was shown in Figure 1, the EC50 for this effect is 0.66 ug/mL.