Diazenedicarboxylic acid bis (N,N-dimethylamide), commonly called diamide, inhibits purified protein kinase isolated from swine kidney. The extent of inhibition is increased in the presence of cysteine, reduced glutathione and dithiothreitol. The inhibition of protein kinase is reversible and greater than 80% of the activity of an inhibited preparation can be recovered by dialysis. Diamide appears to be specific for the inhibition of protein kinase. The activities of various other kinases acting on low molecular weight substrates were unaffected by diamide. Diamide is the first known low molecular weight inhibitor of protein kinase and may prove to be an extremely useful agent in elucidating the physiological role of this enzyme. © 1975.