Quencher-labeled substrate (represented by the forked lines) is phosphorylated by PI 3-K and binds to the Sensor (yellow circle) via the phosphate-metal ion interaction. The fluorescence is quenched (gray circle) as a result of this interaction (i.e., turn-off mode). The assay can also be read in turn-on mode by measuring the emission of the energy transferred from Sensor to fluorescent dye.
The activity of PI 3-Kb was measured using the various wavelength modes as outlined in the Detailed Protocol.
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