Matrix metalloproteinase 9 (MMP-9) has been implicated in various pathophysiologic conditions such as cancer invasion and and bone re~orption.~ To study the release and function of MMP-9 in body fluid and tissue, we established a one-step sandwich enzyme immunoassay (EIA) system for MMP-9 and applied it to measure MMP-9 in human plasma from normal subjects and patients with disease.ProMMP-9 was purified from the conditioned medium of HT1080 cells stimulated with tumor necrosis factor-alpha as previously described? Monoclonal antibodies for MMP-9 were prepared against the purified proMMP-9. A monoclonal antibody for TIMP-1 from bovine dental pulp is capable of cross-reacting with human TIMP-1.6 A one-step sandwich EIA system for MMP-9 was developed using a combination of monoclonal antibodies (clones 73-18B3 and 73-74E11) as a solid phase and a peroxidase conjugate, respectively.The sensitivity of this EIA for standard proMMP-9 was 2.4 pglassay (0.24 ng/mL), and linearity was obtained between 0.24 and 250 ng/mL (2.4-2,500 pg/ assay) (FIG. 1). The intra-assay coefficients of variation (CVs) of normal plasma having MMP-9 at a concentration of 35,104, and 109 ng/mL were 1.8-3.9% (n = 10). Inter-assay CVs were 1 . 8 4 2 % (n = 8) at plasma MMP-9 levels of 38, 47, and 114 ng/mL. Recovery of standard proMMP-9 added to human plasma having an MMP-9 concentration of 35,45, and 100 ng/mL over a range of 12.5-200 ng/mL was 103 C 7.6% (mean rf: SD). This EIA system recognized proMMP-9, the intermediate form with M , = 83 kDa and their complexes formed with TIMP-I.The concentration of MMP-9 in normal human sera by our ElA was 346 c 197 ng/mL. MMP-9 levels in human sera increased with time after venipuncture. Therefore, our one-step sandwich EIA technique was applied to evaluate MMP-9
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.