| Assay Method Information | |
| | Inhibition Assay |
| Description: | The PDE9-inhibiting activity was measured by the following method. That is, to a buffer solution containing tris(hydroxymethyl)aminomethane-hydrochloric acid (40 mM, pH 8.0), magnesium chloride (5 mM), and 2-mercaptoethanol (4 mM) were added cGMP (1 μM) and 3H-cGMP (0.33 μCi/ml) to give a substrate buffer solution. A test substance solution and an enzyme solution which had been adjusted to an optimal concentration were added thereto to perform a reaction at 30° C. The enzyme reaction was stopped by the addition of Scintillation Proximity Assay (SPA) Beads (Perkin Elmer, USA) containing 5 mM 3-isobutyl-1-methylxanthine (IBMX). For the enzyme activity, the amount of 5′-GMP, which is a reaction degradation product bound to SPA beads, was measured with a TopCount microplate reader (Hewlett Packard, USA). |
| Affinity data for this assay | |
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