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The MaxDiscovery™ Glyceraldehyde 3-Phosphate Dehydrogenase (GAPDH) ELISA Kit is an enzyme immunoassay that analyzes the quantity of GAPDH in cells, tissues, serum or urine. Glyceraldehyde 3-Phosphate Dehydrogenase (GAPDH) is well known as one of the key enzymes involved in glycolysis. GAPDH initiates the second stage of glycolysis, catalyzing the reaction that converts glyceraldehyde 3-phosphate (GAP) into 1,3 bisphosphoglycerate (1,3 BPG). GAPDH oxidizes and phosphorylates GAP to produce 1,3 BPG, which is then used as an intermediate in the synthesis of ATP.
While the glycolytic function of GAPDH is widely known, recent evidence suggests that GAPDH is a highly versatile molecule that plays several diverse roles in living systems. Mammalian GAPDH is involved in a great number of intracellular processes such as membrane fusion, microtubule bundling, phosphotransferase activity, nuclear RNA export, DNA replication and DNA repair. There have also been many findings that GAPDH plays a role in different pathologies including prostate cancer progression, programmed neuronal cell death and age-related neuronal diseases, i.e. Alzheimer’s and Huntington’s disease.
The GAPDH gene is constitutively expressed at high levels in almost all tissues. However, the molecular mechanism that sustains high-level expression of this housekeeping enzyme is still unclear. GAPDH is almost always a tetramer and is localized to the cytoplasm in healthy cells. Translocation of GAPDH into the nucleus is seen during its role in the early stages of apoptosis and oxidative stress. Because of its high-level and constitutive expression, GAPDH is widely used as a loading control for Northern/Western blots and for protein normalization.
Like most ELISA assays, the MaxDiscovery™ GAPDH ELISA Test relies on a Horseradish Perioxidase (HRP) conjugated antibody and the TMB (3,3´,5,5´-tetramethylbenzidine) substrate. TMB is a chromogen that yields a blue color when oxidized with hydrogen peroxide (catalyzed by HRP) that has major absorbances at 370 nm and 652 nm. The color then changes to yellow with the addition of acid with maximum absorbance at 450 nm. The relative amount of GAPDH protein in the cells will be directly proportional to the amount of signal that is obtained at 450 nm. |
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