Quantitation of D-type cyclins and their CDK subunits in 32D cells. Lysates from equal numbers of parental 32D cells (lanes 1 and 2) or from clone 2C cells grown in the absence (lane 3) or presence (lane 4) of zinc were precipitated with nonimmune rabbit serum (lane 1, NRS) or with antisera to various cyclins or CDKs (lanes 2 to 4) as indicated at the right of each panel. Immune precipitates were electrophoretically separated on denaturing gels together with the indicated amounts (ng) of corresponding recombinant proteins and were then blotted with the same antiserum as used for immunoprecipitation. The quantities (in ng) of cyclin D2 (A), D3 (B), CDK4 (C), and CDK6 (D) present in the cell lysates were estimated by comparative densitometry and are indicated by the numbers below lanes 2 to 4 at the left of each panel. Note that with the antisera used, D2 was detected at greater sensitivity than D3, and CDK6 was somewhat more readily detected than CDK4; nonetheless, D3 and CDK4 were the predominant proteins in the lysates (see text).