| Assay Method Information | |
| | HPK1 Inhibition Assay |
| Description: | A stock solution of 10 mM test compound was prepared in DMSO. The compound was prepared by 5-fold and 10-point serial dilutions with HPK1 kinase assay buffer. The HPK1 kinase assay buffer contained 40 mM Tris, pH 7.5; 20 mM MgCl2, 0.1 mg/ml BSA, and 50 μM DTT. 7.5 ng/μl of Active HPK1 (Promega, V4098) and 0.1 μg/ml MBP substrate protein were added to the reaction system. The final concentration of ATP in the HPK1 kinase reaction system was 10 μM. Serial diluted compounds were transferred to the reaction system and incubated at room temperature for 1 hr. 5 μL of ADP-Glo™ Reagent (Promega, V9101) was added and incubated for 40 min to stop the kinase reaction and deplete the unconsumed ATP. After that, 10 ul of kinase detection reagent was added and incubated at room temperature for 30-60 min. The luminescence was measured with a plate-reading luminometer. |
| Affinity data for this assay | |
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