A simple assay method for determination of the specific radioactivity of the γ-phosphate group of 32P-labelled ATP.

Abstract

A simple and rapid assay method is described for determining the specific radioactivity of the gamma-phosphate group of 32P-labelled ATP. Labelled ATP is incubated with cyclic AMP, cyclic AMP-dependent protein kinase and histone H2A under conditions leading to maximum phosphorylation of the histone. The specific radioactivity of the gamma-phosphate group of the 32P-labelled ATP is calculated from the total amount of [32P]Pi incorporated into a standard amount of histone. This assay method uses inexpensive commercially available materials, and it yields an accurate specific radioactivity with as little as 0.25 nmol of 32P-labelled ATP.