Intended use
This immunoassay kit allows for the specific measurement of total porcine active and pro-Matrix Metalloproteinase 9 (total MMP-9) concentrations in cell culture supernates, serum, and plasma.
Introduction
Matrix metalloproteinases (MMPs) are a family of zinc-dependent endopeptidases that degrade extracellular matrix proteins. They are secreted as zymogens (pro-MMPs) that are activated by a variety of proteinases or by reaction with organic mercurials. They are inhibited by specific tissue inhibitors of metalloproteinases (TIMPs) and by α2-macroglobulin. The regulation of MMP activity is important in tissue remodeling, inflammation, tumor growth and metastasis.
Porcine MMP-9 (also known as gelatinase B) is secreted as a 92 kDa zymogen. Cleavage of pro-MMP-9 at or near residue 87 results in the active enzyme with a mass of approximately 82 kDa. MMP-9 has three fibronectin type II domains, a hemopexin-like domain and a proline-rich type V collagen-homologous domain. Pro-MMP-9 can be activated by MMP-3 or by certain bacterial proteinases. MMP-9 is inhibited byα2-macroglobulin or by TIMP-1, which binds to pro-MMP-9 as well as to active MMP-9.
Pro-MMP-9 is secreted by monocytes, macrophages, neutrophils, keratinocytes, fibroblasts, osteoclasts, chondrocytes, skeletal muscle satellite cells, endothelial cells, and various tumor cells. Pro-MMP-9 expression is upregulated by TGF-β1, IL-1β, TGF-α, PDGF-AB, TNF-α and IL-1α. Substrates for MMP-9 include denatured collagen type I (gelatin), native collagens type IV, V, VII, X and XI, fibrinogen, vitronectin, IL-1β and entactin, a molecule that bridges laminin and type IV collagen.
MMP-9 is involved in inflammation, tissue remodeling, wound healing, mobilization of matrix-bound growth factors and processing of cytokines. Its expression correlates with the desmoplasia (abnormal collagen deposition) that accompanies pancreatic cancer, with the metastasis to lymph nodes by porcine breast carcinoma cells and with the invasion of regional vessels in giant cell tumors of bones. MMP-9 may be elevated in gingival crevicular fluid and saliva in patients with gingivitis and periodontal diseases.
Test principle
This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for MMP-9 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any MMP-9 present is bound by the immobilized antibody. An enzyme-linked monoclonal antibody specific for MMP-9 is added to the wells. Following a wash to remove any unbound antibody-enzyme reagent, a substrate solution is added to the wells and color develops in proportion to the amount of total MMP-9 bound in the initial step. The color development is stopped and the intensity of the color is measured.
