| Assay Method Information | |
| | Kinase Assay |
| Description: | Approximately 5-20 μg of purified GST-RET proteins were incubated with 1 mM ATP in 20 μL kinase buffer (10 mM Tris-HCl, 5 mM MgCl2, pH 7.4), at 30° C. for 30 minutes. The kinase reactions were terminated by boiling the samples in SDS-PAGE sample buffer. Samples were resolved on 10% acrylamide gels by SDS-PAGE followed by Western blotting. Phosphorylation was detected using a pan-phosphotyrosine pY99 antibody (available from Santa Cruz Biotechnology) or RET phospho-specific antibody (pY905, available from Cell Signaling, Beverly, Mass., USA). |
| Affinity data for this assay | |
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