内皮素是一种由21个氨基酸组成的活性多肽，为目前已知缩血管活性最强的物质。在生理状态下，由于内皮细胞合成内皮素很少且清除速度快，故血浆中内皮素浓度很低。 但在内皮细胞损伤、组织缺血缺氧等情况下，内皮细胞大量合成内皮素并释放入血，成为参与疾病病理生理过程的重要体液因素。
本试剂盒应用双抗体夹心酶标免疫分析法测定标本中内皮素水平。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内皮素、生物素化的抗人内皮素抗体、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内皮素呈正相关。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 Canine Endothelin-1(ET-1) concentrations in serum and plasma.
Introduction
Endothelin-1 (ET-1), a peptide of 21 amino acid residues, is the most potent vasoconstrictive substance known. Originally isolated from porcine aortic endothelial cells, ET-1 is now known to be one of a family of three mammalian vasoactive peptides that also includes Endothelin-2 (ET-2) and Endothelin-3 (ET-3). These related peptides differ from ET-1 at two and six amino acid residue positions, respectively. A fourth peptide, vasoactive intestinal contractor (VIC), is sometimes classified as rat ET-2. All members of the endothelin family contain two essential disulfide bridges and six conserved amino acid residues at the C-terminus. Additionally, all of the endothelin family members are synthesized initially as prepropolypeptides of approximately 200 amino acid residues encoded by separate genes. These are proteolytically cleaved to produce biologically-inactive propolypeptides of approximately 40 amino acid residues termed “big endothelins”. Big ET-1 is cleaved by the proteolytic action of a membrane-bound metalloprotease [endothelin-converting enzyme (ECE-1)] producing the 21 amino acid residue active peptide. The endothelins are produced by a variety of tissues in vivo, including lung and kidney , brain,pituitary and peripheral endocrine tissues and placenta . There are also reports of ET-1 production in vitroby pancreatic cancer cells,mast cells and a variety of endothelial, epithelial and smooth muscle cells .
The best known action of ET-1 is as a vasoconstrictor. Injection of a single dose of ET-1 produces aninitial decrease in systemic blood pressure followed by a prolonged increase in blood pressure, lastingfor 1 to 3 hours. Endothelins also stimulate contraction of most smooth muscles and stimulate secretion by tissues including kidney, liver and adrenals.Endothelins also act in the brain, stimulating secretion by hypothalamic and pituitary cells.Two receptors for endothelins have been characterized and designated ETA and ETB. Although these receptors are structurally and functionally distinct, they share some similarities. Each contains seven stretches of 20-27 hydrophobic amino acid residues, which are likely to represent transmembrane domains. Both have an N-terminal signal sequence and a long N-terminal extracellular domain. The ETA receptor shows a higher affinity for ET-1 than for ET-2 and lowest affinity for ET-3. The ETB receptor shows approximately equal affinity for each of the three endothelins .
Test principle
This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for ET-1 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any ET-1 present is bound by the immobilized antibody. An enzyme-linked monoclonal antibody specific for ET-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 ET-1 bound in the initial step. The color development is stopped and the intensity of the color is measured.
