| Assay Method Information | |
| | Kinase Assay |
| Description: | JAK2 and JAK2 [V617F] Kinase Assay: In 5× Kinase Buffer A, JAK2 or JAK2 [V617F] kinase was mixed with pre-diluted compounds at different concentrations in duplicate for 10 minutes. The corresponding substrate and ATP were added and reacted at room temperature for 20 minutes (in which negative and positive controls were set: the negative control was a blank control and the positive control was erlotinib). After the reaction was completed, a detection reagent (the reagent in the HTRF Kinase TK kit) was added. After incubation at room temperature for 30 minutes, the enzyme activities in the presence of the compounds disclosed herein at each concentration were measured by an Evnvision microplate reader, and inhibitory activities of compounds at different concentrations on the enzyme activity were calculated. The inhibitory activities of compounds at different concentrations on enzyme activity were then fitted according to the four-parameter equation using Graphpad 5.0 software, and the IC50 values were calculated. |
| Affinity data for this assay | |
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