本试剂盒应用双抗体夹心酶标免疫分析法测定标本中TIMP-2水平。用纯化的抗体包被微孔板，制成固相抗体，往包被单抗的微孔中依次加入TIMP-2抗原、生物素化的抗小鼠TIMP-2抗体、HRP标记的亲和素，经过彻底洗涤后用底物TMB显色。TMB在过氧化物酶的催化下转化成蓝色，并在酸的作用下转化成最终的黄色。颜色的深浅和样品中的TIMP-2呈正相关。用酶标仪在450nm波长下测定吸光度（OD值），计算样品浓度。
Intended use
This immunoassay kit allows for the specific measurement of Mouse tissue inhibitors of metalloproteinase 2,TIMP-2 concentrations in cell culture supernates, serum and plasma.
Introduction
Matrix metalloproteinases (MMPs), also called matrixins, constitute a family of zinc and calcium
dependent endopeptidases that function in the breakdown of extracellular matrix (ECM). They play an important role in many normal physiological processes such as embryonic development, morphogenesis, reproduction and tissue remodeling . They also participate in many pathological processes such as arthritis, cancer and cardiovascular disease. While the amounts of newly synthesized MMPs are regulated mainly at the levels of transcription, the proteolytic activities of existing MMPs are controlled through both the activation of proenzymes or zymogens and the inhibition of active enzymes by endogenous inhibitors,α2-macroglobulins and tissue inhibitors of metalloproteinases (TIMPs).
Among the four known members of the TIMP family, TIMP-2 has features both common to all the TIMPs and unique to itself . The common features include 12 cysteine residues that form 6 disulfide bonds, three of which are in the N- and C-terminal domain, respectively. The N-terminal domain is responsible for tight but non-covalent binding to the active MMPs in a 1:1 stoichiometry. The unique features of TIMP-2 include the binding of its C-terminal domain to the hemopexin-like domain of pro-MMP-2. This interaction is essential in the cell surface activation of pro-MMP-2 by active MMP-14 (MT1-MMP) . TIMP-2 also has functions that are independent of MMP inhibition. For example, it suppresses EGF-mediated mitogenic signaling by short-circuiting EGF R activation.
Test principle
This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for TIMP-2 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any TIMP-2 present is bound by the immobilized antibody. An enzyme-linked polyclonal antibody specific for TIMP-2 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 TIMP-2 bound in the initial step. The color development is stopped and the intensity of the color is measured.
