- 预期应用
- ELISA法定量测定大鼠血清、血浆、组织匀浆、细胞培养物上清或其它相关液体中MIF含量。
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- 实验原理
- 本试剂盒应用双抗体夹心酶标免疫分析法测定标本中MIF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MIF抗原、生物素化的抗大鼠MIF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MIF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。
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- Intended use
- This immunoassay kit allows for the specific measurement of rat MIF concentrations in tissue homogenates, cell culture supernates，serum, and plasma.
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- Introduction
- One of the first cytokines described, MIF (macrophage migration inhibitory factor) was originally identified in studies of delayed hypersensitivity reactions where it was shown to inhibit macrophage migration. It is an important mediator of the innate immune response with potential roles in the pathophysiology of inflammatory, autoimmune, and neoplastic disorders. The rat MIF gene encodes a 115 amino acid, 12.5 kDa secreted protein. Crystallographic studies suggest that MIF exists as a homotrimer, although some reports show that it may exist as a dimer or monomer as well. Although MIF exhibits no homology with other known cytokines, it shares structural homology with several bacterial enzymes. MIF exhibits intrinsic catalytic activity and has been shown to act as a tautomerase and an oxidoreductase, although its physiological importance as an enzyme remains unclear.
MIF is well known as a mediator of acute and chronic inflammatory responses and is released into
circulation following endotoxin treatment or by pro-inflammatory cytokines including TNF-α and IFN-γ. Working in a paracrine or autocrine manner, MIF has potent stimulatory effects on cell growth and survival and enhances the production of an array of inflammatory cytokines. MIF is also known for its inhibitory effects on immunosuppressive glucocorticoids. Neutralizing MIF antibodies or knockout of the MIF gene inhibits mouse models of endotoxemia or septic shock, while treatment with the recombinant protein exacerbates toxicity. Part of the mechanism underlying MIF-induced pro-inflammatory activities is an upregulation of toll-like receptor 4 (TLR4). In rats, MIF levels are known to increase in patients with sepsis or septic shock. MIF is also elevated in the lungs of patients with acute respiratory distress syndrome (ARDS), a hyper-inflammatory response that can accompany sepsis or lung injury. Blockade of MIF activity suppresses inflammation associated with mouse models of rheumatoid arthritis (RA), and elevated levels of MIF are found in the synovial fluid of RA patients. In addition, a polymorphism that results in decreased MIF promoter activity correlates with low disease severity in RA patients. Eosinophils are a potential source for MIF found elevated in bronchoalveolar lavage fluid from asthmatic patients. In addition, elevated levels are associated with atherosclerotic lesions, suggesting that MIF may function in chronic inflammatory responses that accompany arterial wall injury.
Test principle
This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for MIF has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any MIF present is bound by the immobilized antibody. An enzyme-linked monoclonal antibody specific for MIF 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 MIF bound in the initial step. The color development is stopped and the intensity of the color is measured.
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