Cayman’s α-KG Detection Assay provides fluorometric- or colorimetric-based methods for quantifying α-KG in biological samples such as serum, plasma, urine, and tissue. It can also be utilized to determine intracellular and extracellular α-KG concentrations in cell culture samples. In this assay, alanine transaminase (ALT) deaminates α-KG to form pyruvate.1,2 Pyruvate is then oxidized by pyruvate oxidase to yield acetyl phosphate, hydrogen peroxide (H2O2), and carbon dioxide.3 As H2O2 is produced, the cycling mechanism of horseradish peroxidase (HRP) results in the concurrent reduction of H2O2 to H2O and the oxidation of 10-acetyl-3,7-dihydroxyphenoxazine (ADHP) to produce the highly fluorescent compound resorufin.4 Resorufin fluorescence is analyzed with an excitation wavelength between 530-540 nm and an emission wavelength between 585-595 nm. Alternatively, the absorbance of resorufin can be measured at 570 nm.Needed but not supplied: Please download the kit booklet to verify if UltraPure Water (Milli-Q or equivalent) or any other components are needed for this assay.WARNING This product is not for human or veterinary use.
