Localization of Tmtsp protein on endothelial cells. (A) Tissue distribution of Tmtsp mRNA by Northern blotting analysis. (B) Immunohistochemistry of adult mouse brain with anti-Tmtsp mAb. Tmtsp-specific signals were detected on the blood vessel–like structure in the forebrain. (C) Immunohistochemistry of the fetal liver (E15.5) with anti-Tmtsp mAb. Tmtsp protein (red) was detected in VE-cadherin–positive cells (green) with spindle nuclei located along luminal duct filled with enucleated erythrocytes and DAPI-positive blood cells. Nuclei were visualized by DAPI staining (blue). (D) Flow cytometric analysis of endothelial cells. Tmtsp protein was detected on CD45^–CD31⁺ endothelial cells, but not on CD45^–CD31^– nonendothelial cells prepared from adult mouse liver and bEND.3 cells, a mouse brain–derived endothelioma cell line. (E) Double staining of cultured endothelial cells differentiated from ES cells on OP-9 stromal cells. Tmtsp protein (green) was localized on the surface of VE-cadherin–positive endothelial cells (red). Nuclei were visualized with DAPI staining (blue).