Megakaryopoiesis during murine neonatal development. MK concentration (A) and MK diameter (B) were measured in the liver (orange triangles and solid line), BM (green diamonds and interrupted line), and spleen (blue squares and solid line) of newborn mice on days P1, P3, P5, P7, P10, and P14, and in adult mice (>60 days) after immunohistochemical staining with anti-VWF. Data presented are means ± SD from at least 4 mice at each time point. (A) MK concentration, expressed as number of MKs present in an area of tissue measuring 250 × 250 µm at ×400. (B) MK diameter (in μm) measured under the microscope at ×400. (C) MK maturation was analyzed by immunofluorescent staining for VWF (green) and PF4 (red). Shown are representative neonatal MKs (spleen, upper row) and adult MKs (BM, lower row). Note a blood vessel in the upper panel, with visible endothelial cells expressing only VWF (green). All photomicrographs were obtained at ×400, Bar: 10 μm. (D) MKs isolated from the spleen of newborn mice (P5-P10, top) were compared with MKs from the spleen of adult mice (bottom). The ploidy of CD41/CD61 double-positive cells was determined by flow cytometry after staining with Hoechst 33342. (E) Ultrastructure was analyzed by electron microscopy in flow-sorted 2-4N MKs isolated from the spleen of neonatal mice (P5-P10). Despite their small size, ∼30% of 2-4N neonatal MKs were mature, as indicated by the presence of an open demarcation membrane system. (Bar: 2 μm)