Fig. 5. Regulation of S phase entry in newt cardiomyocytes. (A) Dependence of entry on serum concentration. Cells were exposed to various concentrations of FBS, as described in Materials and Methods, and pulsed with BrdU 4 days later for 8 hours. Each point is from a separate dish of the same culture, and the peak at approximately 10% FBS was observed in three other comparable experiments. (B,C) Detection of Rb in cardiomyocytes by indirect immunofluorescence with an antibody recognising phosphoserine 608. The cells were stained with anti-MyHC and Hoechst 33258 (DNA; B), anti-phosphoserine 608 (in Rb; C). In control experiments the intensity of nuclear staining was significantly diminished by digestion of fixed cells with lambda phosphatase prior to reaction with the antibody (not shown). Note that most of the nuclei in B are positive for the Rb phosphorylated epitope in C. Scale bar: 50 µm.

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