Antibodies which completely inhibited the enzymatic activity of the protein kinase associated with virions of frog virus were obtained by immunization of rabbits with the purified enzyme. This inhibition provided a specific probe for the frog virus protein kinase, since this antiserum had no inhibitory effect on a variety of other protein kinases, including the activity of uninfected cells, or the protein kinase associated with vesicular stomatitis virus or vaccinia virus cultivated in the same cell line as frog virus. The frog virus protein kinase was characterized as a virus-specified protein on the basis of the following observations: (a) the virion protein kinase was antigenically distinct from essentially all of the protein kinase expressed in uninfected cells; (b) following infection by frog virus more than a 15-fold increase was detected in the specific activity of intracellular protein kinase and most of this activity was antigenically related to the virion enzyme; (c) when frog virus was grown in cells derived from widely different species, the antigenic and biochemical specificities of the virion protein kinase remained identical; and (d) screening of cells infected with different temperature-sensitive mutants of frog virus indicated that certain viral mutants failed to synthesize this protein kinase when cultivated at the nonpermissive temperature.