| Assay Method Information | |
| | Urease Inhibition Assay |
| Description: | Reaction mixtures comprising 25 μL of enzyme (jack bean urease) solution and 55 μL of buffer containing 100 mM urea were incubated with 5 μL of test compound (0.5 mM) at 30°C for 15 min in 96-well plates. Urease activity was determined by measuring the ammonia production using the indophenol method, as described by Weatherburn11. Briefly, 45 μL of each phenol reagent (1% (w/v) phenol and 0.005% (w/v) sodium nitroprussside) and 70 μL of alkali reagent (0.5% (w/v) NaOH and 0.1% active chloride, NaOCl) were added to each well. The increasing absorbanceat 630 nm was measured after 50 min, using a microplate reader (Molecular Devices, Sunnyvale, CA, USA). All reactions were performed in triplicate in a final volume of 200 μL. The results (change in absorbance per min) were processed using SoftMax Pro software (Molecular Devices). The entire assay was performed at pH 6.8. |
| Affinity data for this assay | |
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