Immunohistochemical staining.

Transverse-sectioned E9.5- or E10.5-derived embryos and sagittal-sectioned adult hearts were stained with anti-ALCAM antibody. (A) The E9.5-derived dorsal aorta or vitelline artery was weakly positive for ALCAM (Ai). The vitelline artery in the square area is shown in high magnification (Aii). Black arrowheads indicate positively stained endotheliallike cells. In yolk sac, both mesenchymal cells (M) (white arrowheads) and endothelial cells (Ec) (black arrowheads) were ALCAM⁺ (Aiii). (B) In the E10.5 embryo dorsal aorta, endothelial cells were strongly positive for ALCAM (square area in panel Bi; high magnification of the square area: black arrowheads in panel Bii). Similarly, M (white arrowheads) and Ec (black arrowheads) were ALCAM⁺ in E10.5 yolk sac (Biii). (C) Adult mouse, heart, and aorta were examined for the expression of ALCAM. Antiheart muscle antibody (Ci), antismooth muscle antibody (Cii), and anti-CD31 antibody (Ciii) (black arrowheads indicate endocardium; black arrows, wall of aorta) were used for control, respectively. Note that ALCAM was negative for endocardium and endothelium of aorta (Civ). Scale bar, 200 μm. Da indicates dorsal aorta; N, notocord; Va, vitelline artery; Fp, floor plate; My, myocardium; Tr, trachea; Eo, esophagus.