Abnormal VWF distribution in Nbeal2^−/− megakaryocytes. (A) Immunofluorescence imaging of a representative permeabilized mature extending stage WT megakaryocyte shows elaboration of demarcation membranes stained with CD41 (green), dispersed VWF expression (red), and a peripheral tubulin cytoskeletal meshwork (violet). (B) A representative Nbeal2^−/− MK at the same stage shows a strong peripheral distribution of VWF, which is absent in Nbeal2^−/− platelets; this MK also contains the nucleus of an exogenous cell in its cytoplasm visible in the XZ view of the merged confocal panel. Comparisons of nonpermeabilized cells (C) confirm that VWF is abnormally concentrated near or on the surface of mature Nbeal2^−/− megakaryocytes (scale bars in ZY panels = 5 µm; scale bars in XY, XZ, and extended focus panels = 10 µm). (D) Transmission electron micrograph of immunogold labeled VWF present on the surface of a Nbeal2^−/− megakaryocyte. Magnification ×50 000; scale bar represents 500 nm.