Description
Introduction: Endothelin-1(ET-1), a peptide of 21 amino acid residues, is the most potent vasoconstrictive substance known. Originally isolated from 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. The biochemistry and biology of the endothelins have been the subject of several reviews.
Principle of the Assay: The microtiter plate provided in this kit has been pre-coated with an antibody specific to ET-1. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific for ET-1 and Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. Then a TMB (3,3',5,5' tetramethyl-benzidine) substrate solution is added to each well. Only those wells that contain ET-1, biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of a sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450 nm +/- 2 nm. The concentration of ET-1 in the samples is then determined by comparing the O.D. of the samples to the standard curve