|
预期应用
ELISA法定量测定人血清、血浆或其它相关液体中HO-1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HO-1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HO-1抗原、生物素化的抗人HO-1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HO-1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 HO-1 concentrations in serum, and plasma.
Introduction
Heme Oxygenase-1 (HO-1) also known as Hsp32, is the inducible isoform of heme oxygenase that catalyzes the NADPH, O2 and cytochrome P450 reductase dependent oxidation of heme to carbon monoxide, ferrous iron and biliverdin which is rapidly reduced to bilirubin. These products of the HO reaction have important physiological effects: carbon monoxide is a potent vasodilator and has been implicated to be a physiological regulator of cGMP and vascular tone; biliverdin and its product bilirubin are potent antioxidants; “free” iron increases oxidative stress and regulates the expression of many mRNAs (e.g., DCT-1, ferritin and transferring receptor) by affecting the conformation of iron regulatory protein (IRP)-1 and its binding to iron regulatory elements (IREs) in the 5’- or 3’- UTRs of the mRNAs. To date, three identified heme oxygenase isoforms are part of the HO system that catalyze heme into biliverdin and carbon monoxide. These are inducible HO-1 or Hsp32, constitutive HO-2 that is abundant in the brain and testis, and HO-3 which is related to HO-2 but is the product of a different gene. The HO system is the rate-limiting step in heme degradation and HO activity decreases the levels of heme which is a well known potent catalyst of lipid peroxidation and oxygen radical formation. The expression of HO-1 is highly responsive to all types of stimuli that cause oxidative stress and it is up regulated during exposure to oxidants, UV-A irradiation and a series of agents including cytokines, hormones, heme and heavy metals. HO-1 is a vital component of neuronal defense mechanisms and oxidative stress has been postulated to be the underlying basis for neuronal cell death in neurodegenerative diseases such as Alzheimer’s disease (AD) and Parkinson’s disease. The expression of HO-1 is normally very low in the brain but increases markedly after heat shock, ischemia or glutathione depletion. Spatial distribution of HO-1 expression in AD brain is essentially identical to that of the pathogenic conformational changes of tau protein that is the major component of the intraneuronal lesion of AD, neurofibrillary tangles.
Test principle
This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for HO-1 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any HO-1 present is bound by the immobilized antibody. An enzyme-linked monoclonal antibody specific for HO-1 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 HO-1 bound in the initial step. The color development is stopped and the intensity of the color is measured. |
