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Fig. 5. Apoptosis and proliferation analysis of Mp+/+ and Mp–/– EBs. (A) TUNEL analysis of the Mp+/+ and Mp–/– EBs. (a) TUNEL staining. Fluorescent green cells are TUNEL-positive apoptotic cells. Left panels are TUNEL staining; right panels are the overlapped images of DAPI and TUNEL staining. Note the pattern of apoptosis in Mp+/+ (below) and Mp–/– (above) EBs. Arrows indicate the apoptotic cells. (b) Analysis of TUNEL-positive apoptotic cells in Mp+/+ and Mp–/– EBs. N, the number of cells counted. No significant difference is observed between the two classes of EBs. (B) Mp–/– EBs have a reduced rate of cell proliferation. (a) The mitotic index of EBs was determined by immunostaining with anti-phosphorylated histone 3 (H3P). Arrows indicate the proliferating cells. (b) Statistical analysis of the percentage of H3P-positive cells in Mp+/+ and Mp–/– EBs. N, the number of cells counted. Note that there is a significant difference between the two classes of EBs.