Objective: We analyzed, in vivo, whether the establishment of blood supply to implanted scaffolds can be accelerated by inosculation of an in situ-preformed microvascular network with the host microvasculature. Background: A rapid vascularization is crucial for the survival of scaffold-based transplanted tissue constructs. Methods: Poly-lactic-glycolic acid scaffolds were implanted into the flank of balb/c or green fluorescent protein (GFP)-transgenic mice for 20 days to create in situ a new microvascular network within the scaffolds. The prevascularized scaffolds were then transferred into the dorsal skinfold chamber of isogeneic recipient mice. Nonvascularized poly-lactic-glycolic acid scaffolds served as controls. Vascularization, blood perfusion, and cell survival of the implants were analyzed over 14 days using intravital fluorescence microscopy, histology, and immunohistochemistry. Results: Our results demonstrate that establishment of blood perfusion of prevascularized scaffolds is significantly accelerated and improved (136.7 Ϯ 23.2 pl/s) when compared with controls (6.9 Ϯ 1.9 pl/s), because the in situ-preformed microvessels were reperfused by forming interconnections to the host microvasculature. Apoptotic cell death within the implants was found only during the first 3 to 6 days after scaffold implantation during lack of blood perfusion, but not during the further 14-day observation period. Conclusions: Inosculation of in situ-preformed functional blood vessels represents a promising approach to improve the blood supply to implanted tissue constructs. (Ann Surg 2008;248: 939 -948)