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Assay Method Information

Assay Name:  Fluorimetric Assay
Description:  The activities of recombinant hMAO-A and hMAO-B were determined using p-tyramine as common substrate and calculated as 0.18 +/- 0.01 nmol/mg/min (n = 3) and 0.12 +/- 0.02 nmol/mg/min (n = 3), respectively. The interactions of the synthesized compounds with hMAO isoforms were determined by a fluorimetric method described previously. The production of H2O2 catalyzed by MAO isoforms was detected using a non-fluorescent Amplex-Red reagent which reacts with H2O2 in the presence of horseradish peroxidase to produce the fluorescent product resorufin. The reaction was started by the addition of 200 uM Amplex Red reagent, 1 U/mL horseradish peroxidase, and p-tyramine (concentration range 0.1 to 1 mM). Control experiments were carried out using reference inhibitors (Selegiline and Moclobemide). The possible capacity of compounds to modify the fluorescence generated in the reaction mixture due to non-enzymatic inhibition was determined by adding these compounds to solutions containing only the Amplex Red reagent in sodium phosphate buffer.
Affinity data for this assay
 

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Last update November 1, 2007
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