Adaptation of the protein kinase filter paper assay to a 96-well microtiter format.
The most widely used method for assaying protein kinase activities involves incorporation of radioactive phosphate into a protein or peptide substrate with subsequent binding or precipitaion of the radiolabeled substrate onto filter paper squares. We have adapted this assay for use with readily available 96-well microtiter plate technologies. The sensitivity and reproducibility of the modified 96-well protein kinase assay are comparable to standard filter paper assay methods, but assay throughput is much greater and reagent costs are significantly reduced. This technique has been used to assay several different protein kinase activities using both P81 phosphocellulose and 3MM filter papers. The general methods described here could be readily adapted for use with other kinds of filter materials or used with automated assay systems.
Protein Kinase Activities; Filter Papers; Radioactive Phosphate
Amino Acid Sequence; Indicators and Reagents; Microchemistry; Phosphorus Radioisotopes; Protein Kinases; Scintillation Counting; Substrate Specificity