Lipoprostaglandin E1 (lipo-PGE1) has been found accumulating in injured vascular regions. This study examined the localization of lipo-PGE1 in the anastomotic region. The study was divided into three parts. First, we performed anastomosis of the rat femoral artery and vein (n = 17). Lipo-PGE1 labeled with 1,1'-dioctadecyl-1,3,3',3'-tetramethyl-indocarbocyanine was infused intravenously. Hematoxylin-Eosin staining and fluorescence microscopic findings showed that lipo-PGE1 markedly accumulated at the anastomotic site when compared to the contralateral non anastomotic region. Then, we measured laser Doppler flow (LDF) of a lower leg before and after infusion of lipo-PGE1 (n = 7) and saline (n = 7). Increase of blood flow was maintained 1 hour after the infusion of lipo-PGE1 (144% ± 25.0%) when compared to saline infusion. Finally, we performed immunohistochemical and electron microscopic examinations and found that Lipo-PGE1 was incorporated in vascular smooth muscle cells of the anastomotic region. These findings suggest selective accumulation of the lipo-PGE1 in the vascular anastomosis site and affect on the blood flow of repaired vessels.