Figure 6.

Only PBK from mitotic HeLa cells becomes phosphorylated in an immunocomplex kinase assay. Preimmune serum or immune serum precipitates from asynchronous cell lysate (A) and from lysates of cells synchronized at S-phase (S) or M-phase (M) were assayed for kinase activity in kinase assay buffer supplemented with (³²P-γ)-ATP. The immunocomplexes were separated by SDS/PAGE and transferred to nitrocellulose. The blot was probed with anti-PBK antibody (Lower) to verify that PBK was immunoprecipitated in the presence of serum. PBK isolated from mitotic cells appears as a doublet. An autoradiograph of the same blot (Upper) shows that only PBK isolated from mitotic cells was phosphorylated.