Localized necrotic cell death in Rb-null fetal liver. (A-B) TUNEL immunohistochemistry on sagittal sections of wild-type and Rb-null E13.5 fetal liver. (C-D) Representative flow cytometric analysis of annexin-V and TER119 staining on disaggregated whole E13.5 fetal liver from wild-type (C, 7.1% ± 0.6%, P < .002) and Rb-null (D, 2.8% ± 1.0%, P < .002) embryos to detect phosphotidyl serine externalization. (E-G) TUNEL staining for DNA fragmentation of wild-type (E) and Rb-null (F-G) E13.5 fetal livers (1000× magnification). The arrow in panel E identifies a phagocytic macrophage and s indicates a liver sinusoid. The dorsal region of the Rb-null fetal liver (F) shows markedly less TUNEL positivity than the ventral end of the liver that is distant from the hepatic vasculature. (H-J) Protein blue staining of sections of E13.5 wild-type fetal liver (H), more dorsal, healthy regions of the Rb-null fetal liver (I), and dying regions in the ventral end of the Rb^−/− fetal liver (J) with black arrows indicating the presence of phagocytic macrophages and red arrows indicating distended, remnant nuclei. See ″Fluorescence microscopy and island counts″ for image acquisition information.