| Solution Information | help | |
| Enzyme: | Glycogen synthase kinase-3 beta | |
| inhibitor: | BDBM68345 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | 1) Dispense 1 uL/well of CABPE, 0.5 uL of ATP, and 1 uL of positive control GW8510 or AB in respective wells according to plate design to 1536-well assay ready plates (Aurora 29847) generated by acoustic transfer using Labcyte Echo that contain 7.5 nL/well of 10 mM compound using BioRAPTR (Beckman) to start the reaction. Incubate at room temperature for 60 minutes. 2) Add 2.5 uL/well of ADP-glo (Promega, V9103) with BioRAPTR, incubate at room temperature for 40 minutes 3) Add 5 uL/well of ADP-glo (Promega, V9103) with Combi nL (Thermo), incubate at room temperature for 30 minutes 4) Read on Viewlux (PerkinElmer) for luminescence Solutions: AB: 25 mM Tris 7.5 10 mM MgCl2 GW8510 (in AB, Sigma G7791) 50 uM GW8510 CABPE (in AB): 12.5 mM DTT (Sigma 43816) 0.25 mg/mL BSA (Sigma A4503) 0.5 U/mL Heparin (Baxter NDC 0641-2440-41) 8 uM Peptide (American Peptide) 9 nM GSK3beta (BPS Biosciences) ATP (in AB, Promega V9103 component): 125 uM ATP | |
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