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Assay Method Information

Assay Name:  Enzyme Assay
Description:  The functional activity of compounds inhibiting the DAAO enzyme was determined by utilizing the co-product of the catalysis of D-Serine, H2O2 which can be quantitatively measured using the Amplex Red (Invitrogen) detection. Amplex Red reagent is a colorless substrate that reacts with hydrogen peroxide (H2O2) with a 1:1 stoichiometry in the presence of hydrogen peroxide to produce highly fluorescent resorufin (excitation/emission maxima=570/585 nm). The changes in fluorescence were monitored by a fluorescence plate reader, Envision (Perkin Elmer) and increases in DAAO activity were readily detected upon addition of D-Serine and suppression of this response observed with the application of test compounds.Human DAAO enzyme was supplied by the Takeda Pharmaceutical Company (Osaka) and each batch was tested and used at concentrations giving comparable levels of activity. The Km of D-Serine was measured for each enzyme batch to maintain consistency; this Km was used in subsequent assays.
Affinity data for this assay
 

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Last update November 1, 2007
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