A SOLID-PHASE ASSAY FOR THE DETERMINATION OF PROTEIN-TYROSINE KINASE-ACTIVITY OF C-SRC USING SCINTILLATING MICROTITRATION PLATES

A solid-phase assay for the determination of protein tyrosine kinase ( PTK) activity has been developed. The transfer of (PO4)-P-33 from ATP to the synthetic substrate poly(Glu, Tyr) 4:1 attached to the bioactiv e surface of scintillating microtiter plates served as the basis to ev aluate enzyme activity. The procedure eliminates detection with phosph otyrosine antibodies, tedious separation of phosphorylated peptides wi th phosphocellulose membranes, and extensive washing steps. For these reasons, the traditionally time-consuming procedure can be performed w ith a simple three-step protocol. The method is highly accurate, rapid , and robotics friendly. The advantages over existing assays make this procedure especially suited for high throughput applications. (C) 199 6 Academic Press, Inc.