Description
Background: Endothelial NOS (eNOS), also known as nitric oxide synthase 3 (NOS3) or constitutive NOS (cNOS), is an enzyme that in humans is encoded by the NOS3 gene located in the 7q35-7q36 region of chromosome 7. Nitric oxide is a reactive free radical which acts as a biologic mediator in several processes, including neurotransmission and antimicrobial and antitumoral activities. Nitric oxide is synthesized from L-arginine by nitric oxide synthases. Variations in this gene are associated with susceptibility to coronary spasm. Alternative splicing and the use of alternative promoters results in multiple transcript variants.
Principle of the Assay: The Picokine™ Human NOS3 Pre-Coated ELISA (Enzyme-Linked Immunosorbent Assay) kit is a solid-phase immunoassay specially designed to measure Human NOS3 with a 96-well strip plate that is pre-coated with antibody specific for NOS3. The detection antibody is a biotinylated antibody specific for NOS3. The capture antibody is a monoclonal antibody from mouse and the detection antibody is a biotinylated polyclonal antibody from goat. The kit includes Human NOS3 protein as standards. To measure Human NOS3, add standards and samples to the wells, then add the biotinylated detection antibody. Wash the wells with PBS or TBS buffer, and add Avidin-Biotin-Peroxidase Complex (ABC-HRP). Wash away the unbounded ABC-HRP with PBS or TBS buffer and add TMB. TMB is an HRP substrate and will be catalyzed to produce a blue color product, which changes into yellow after adding the acidic stop solution. The absorbance of the yellow product at 450nm is linearly proportional to Human NOS3 in the sample. Read the absorbance of the yellow product in each well using a plate reader, and benchmark the sample wells' readings against the standard curve to determine the concentration of Human NOS3 in the sample