| Assay Method Information | |
| | PIM1/2/3 Kinase Activity Assay |
| Description: | PIM1, PIM2, and PIM3 (all three kinases are from Carna) were each prepared with 1× Kinase buffer into a kinase solution at a concentration that was 2.5 times the final concentration. 10 μL of kinase solution at a concentration that was 2.5 times the final concentration was added to each of the compound wells and the positive control well, and 10 μL of 1× Kinase buffer was added to the negative control well. The plate was centrifuged at 1000 rpm for 30 s, mixed well by shaking, and then incubated at room temperature for 10 min. A mixed solution of ATP and Kinase substrate 20 (from GL) at a concentration that was 25/15 times the final concentration was prepared with 1× Kinase buffer. The reaction was started by adding 15 μL of the mixed solution of ATP and Kinase substrate 20 at a concentration that was 25/15 times the final concentration. The 384-well plate was centrifuged at 1000 rpm for 30 s, mixed well by shaking, and then incubated at room temperature for 60 min. 30 μL of stop assay buffer was added to stop the kinase reaction. The plate was centrifuged at 1000 rpm for 30 s and mixed well by shaking. The conversion rates were read using Caliper EZ Reader. |
| Affinity data for this assay | |
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