Detection Method:Sandwich
Test principle:This assay employs a two-site sandwich ELISA to quantitate UCP1 in samples. An antibody specific for UCP1 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and anyUCP1 present is bound by the immobilized antibody. After removing any unbound substances, a biotin-conjugated antibody specific for UCP1 is added to the wells. After washing, Streptavidin conjugated Horseradish Peroxidase (HRP) is added to the wells. Following a wash to remove any unbound avidin-enzyme reagent, a substrate solution is added to the wells and color develops in proportion to the amount of UCP1 bound in the initial step. The color development is stopped and the intensity of the color is measured.
Product Overview:UCP1 is activated in the brown fat cell by fatty acids and inhibited by nucleotides. Sympathetic nervous system terminals release Norepinephrine onto a Beta-3 adrenergic receptor on the plasma membrane. This activates adenylyl cyclase, which catalyses the conversion of ATP to cyclic AMP. cAMP activates protein kinase A, causing its active C subunits to be freed from its regulatory R subunits. Active protein kinase A, in turn, phosphorylates triacylglycerol lipase, thereby activating it. The lipase converts triacylglycerols into free fatty acids, which activate UCP1, overriding the inhibition caused by purine nucleodides. At the termination of thermogenesis, the mitochondria oxidize away the residual fatty acids, UCP1 inactivates and the cell resumes its normal energy-conserving mode.
