预期应用
ELISA法定量测定人血清、血浆、细胞培养物上清或其它相关液体中iNOS含量。
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iNOS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iNOS、生物素化的抗人iNOS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iNOS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 Human iNOS concentrations in cell culture supernates, serum, and plasma.
Introduction
NO is produced by a group of enzymes called nitric oxide synthases (NOS). These enzymes catalyze the production of NO and L-citrulline from L-arginine, O2, and NADPH-derived electrons. Mammalian systems contain three well-characterized isoforms of the enzyme: neuronal NOS (nNOS, also called NOS1), inducible NOS (iNOS or NOS2), and endothelial NOS (eNOS or NOS3). The names reflect characteristics of the activity or the original tissues in which the enzymes were first described, but it is now known that each of these isoforms is expressed in a variety of tissues and cell types . The three main isoforms share structural similarities and have nearly identical catalytic mechanisms.
They all require a number of cofactors and prosthetic groups for activity including FAD, FMN,heme, calmodulin, and tetrahydrobiopterin. The homodimeric form is required for NO production, and the subunits have molecular masses of approximately 160 kDa (nNOS), 135 kDa (eNOS), and 130 kDa (iNOS). Three distinct domains are necessary for catalytic activity. Starting at the C-terminus there is a reductase domain, a calmodulin-binding domain,and an oxygenase domain. The reductase domain contains the FAD and FMN moieties and shares extensive amino acid (aa) homology with cytochrome P-450 reductase. This domain transfers electrons from NADPH to the oxygenase domain. The oxygenase domain actually catalyzes the conversion of arginine into citrulline and NO and contains the binding sites for heme, tetrahydrobiopterin, and arginine. Calmodulin binding is required for activity of all of the NOS isoforms.
Test principle
This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for iNOS has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any iNOS present is bound by the immobilized antibody. An enzyme-linked polyclonal antibody specific for iNOS 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 iNOS bound in the initial step. The color development is stopped and the intensity of the color is measured. | 
