Tmod3^−/−fetal liver erythroblasts display increased apoptosis throughout terminal differentiation. (A) Percentage of apoptotic cells based on Annexin V-positive cells as a proportion of total 7-AAD–negative fetal liver cells at E14.5. (B) Percentage of apoptotic cells within each R population. The 7-AAD–negative cells were first gated on CD71/Ter119 to identify R populations as in Figure 4C, and percentages for each R was calculated as the number of Annexin V-positive cells as a fraction of the total cell number in that R population. All values are means ± SD with at least 3 replicates with samples prepared from different individual embryos. *P < .05. (C) Cytospins of fetal liver cells stained with lamin B (red) for nuclear envelope, Ter119-Alexa488 (green) for a GPA-associated antigen to identify late erythroblasts, and Hoechst (blue) for nuclei. (D) Cytospins of fetal liver cells stained with active-caspase3 (red) for apoptosis, Ter119-Alexa488 (green), and Hoechst (blue). (E) Cytospins of fetal liver cells stained with Annexin V (green) for apoptosis, Ter119-PE (red), and Hoechst (blue). Panels C-E were acquired with a Zeiss LSM 780 laser scanning confocal fluorescence microscope using a Zeiss 100× oil-immersion objective (N.A. 1.4) at a zoom of 2. Scale bar, 4 µm.