Amylase Fluorometric (Green) Assay Kit, EnzChek® from Molecular Probes (Invitrogen)

The EnzChek Ultra Amylase Assay Kit (E33651) provides a solution-based assay featuring the speed, high sensitivity, and convenience required for measuring amylase activity or for screening amylase inhibitors in a high-throughput format. This EnzChek kit contains a starch derivative the DQ starch substrate that is labeled with BODIPY FL dye (Figure 1) to such a degree that the fluorescence is quenched. This substrate is efficiently degraded by amylase; digestion relieves the quenching and yields highly fluorescent fragments. The accompanying increase in fluorescence is proportional to amylase activity (Figures 2 and 3) and can be monitored with a fluorescence microplate reader or fluorometer, using standard fluorescein filters.

In tests using a-amylase from Bacillus sp. (Sigma A-6380) and 200 g/mL of the DQ starch substrate (30 minute incubation at room temperature), the EnzChek Ultra Amylase Assay Kit could be used to detect a-amylase activity down to a final concentration of 2 10-3 U/mL, where one unit is defined as the amount of enzyme required to liberate 1 mg of maltose from starch in 3 minutes at 20C, pH 6.9.