| Assay Method Information | |
| | Biochemical Assay |
| Description: | Biochemical IC50s were measured at Invitrogen using Z-lyte technology. Briefly, for STK4, the 2×STK4 (MST1)/Ser/Thr 07 mixture was prepared in 50 mM HEPES pH 7.5, 0.01% BRIJ35, 10 mM MgCl2, 1 mM EGTA. The final 10 μL Kinase Reaction consisted of 6.25-160 ng STK4 (MST1) and 2 μM Ser/Thr 07 in 50 mM HEPES pH 7.5, 0.01% BRIJ-35, 10 mM MgCl2, 1 mM EGTA. After the 1 hour Kinase Reaction incubation, 5 μL of a 1:100000 dilution of Development Reagent A was added. Compounds were screened in a 10 point titration with 3-fold serial dilutions starting at a concentration of 10 μM. The biochemical IC50s against other STK family members were measured similarly. The ATP concentration used matched the Km for each kinase. |
| Affinity data for this assay | |
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