FIGURE 2.
HTS assay optimization for the Eya2 phosphatase assay using OMFP as a substrate. (A) Scattered plot of a DMSO plate test of the Eya2 phosphatase assay in a 1536-well plate. The wells in column 2 contain 1 mM EGTA that was used as a positive control. All other wells contain DMSO as the negative (no inhibition) control. (B) Zā and signal/background (S/B) values of a pilot screen using the LOPAC library. (C) The pilot screen revealed three compounds that inhibit Eya2 phosphatase activity. (D) Chemical structures of the three compounds in C. (E) Two pilot screen compounds inhibit Eya2 phosphate activity using the phospho-peptide substrate, pH2AX.