| Assay Method Information | |
| | DYRK1A Inhibition Assay |
| Description: | 1. Add 5 μL enzyme & substrate mixture to each well in Column 2-23 and A24-H24 wells of the 384-well assay plate;2. Add 5 μL 0% Phosphorylation control to A1-H1 and I24-P24 wells of the assay plate;3. Add 5 μL 100% Phosphorylation control to I1-P1 wells of the assay plate;4. Spin the assay plate (1000 rpm, 1 minute @23° C.);5. Incubate enzyme with compounds for 15 minutes at 23° C.;6. Add 5 μL ATP solution to each well of the assay plate;7. Spin the plate (1000 rpm, 1 minute @23° C.);8. Incubate the assay plate for 90 minutes at 23° C.;9. Add 10 μL Development reagent A to each well of the assay plate;10. Centrifuge the plate at 1000 rpm about 15 seconds and seal a film over assay plate. Incubate the assay plate for 30 minutes at 23° C.11. Read assay plates on Envision (see Tables A, B, and C).Final Compound ConcentrationsAssay buffer: 50 mM Hepes pH7.5, 10 mM MgCl2, 1 mM EDTA, 0.01% Brij-35DYRK1A: 1 nMATP: 20 μMSer/Thr 18 peptide: 2 μMReaction time: 90 minutes. |
| Affinity data for this assay | |
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