| Assay Method Information | |
| | Eya Phosphatase Assay |
| Description: | Eya activity was measured in 50-μl reactions using black, 96-well, half-volume microtiter plates (Greiner Bio-one) with the substrate OMFP (3-O-methylfluoresceinphosphate, Sigma-Aldrich), which is converted to a fluorescent product OMF upon dephosphorylation. The assay conditions used were 50 mM MES, pH 6.5, 50 mM NaCl, 1.25 mM MgCl2, 0.05% BSA, and 1mM DTT, and the reaction contained 50 nM Eya2 ED and 100 μM OMFP. Reactions were started by the addition of OMFP, preceded for 1 h at room temperature followed by terminating the reaction with the addition of 75mMEDTA. Fluorescence intensity was measured at 485/515 nm excitation/emission on a Fluoromax-3 plate reader (Horiba Jobin Yvon). |
| Affinity data for this assay | |
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