| Assay Method Information | |
| | CDK1/Cyclin B1 ADP-Glo Kinase Assay |
| Description: | The luminescent signal generated is proportional to the ADP concentration produced and is correlated with the kinase activity. CDK1/Cyclin B1 was purchased from Carna (Cat 04-102). Typical reaction solutions (10 uL final reaction volume) contained 2% DMSO (f inhibitor), 10 mM MgCl2, 1 mM EGTA, 0.05% BSA, 2 mM DTT, 80 uM ATP (ATP Km=78.6 uM), 0.01% Brig-35, 0.75 uM substrate, and 4.917 nM CDK1/Cyclin Bl enzyme complex in 50 mM HEPES buffer at pH 7.5. The assay was initiated with the addition of ATP-containing substrate solution, following a 30-minute pre-incubation of enzyme and inhibitor at room temperature in the reaction mixture. The reaction was stopped after 90 minutes at room temperature by the addition of 10 uL of ADP-GLO Reagent. After a 90 minute incubation, 20 uL of Kinase Detection Reagent was added. Samples were incubated for 40 minutes, after which plate well luminescence was measured on a Envision microplate reader. |
| Affinity data for this assay | |
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