Bone morphogenetic protein-2 is a very promising candidate for the treatment of bone diseases and defects, but more effective therapeutic methods are required due to its instability in vivo. A controlled and localized delivery system of Bone morphogenetic protein-2 would be appropriate for effective bone regeneration. Here, we report a novel delivery system of bone morphogenetic protein-2 using vitrigel (a novel stable collagen gel membrane prepared from vitrified type 1 collagen) for in vivo bone regeneration. Scanning electron microscopy revealed that the collagen vitrigel formed a tightly woven network with average pore sizes of about 1-2 mu m. The vitrigel scaffold delivery system exhibited sustained release of bone morphogenetic protein-2 and >80% of the total bone morphogenetic protein-2 was still retained in the vitrigel after 15 days in phosphate-buffered saline in vitro. Bone morphogenetic protein-2-containing vitrigel was transplanted into mouse calvarial defects. The enhanced mechanical strength of the vitrigel made it easier to implant into defects without damage. Obvious bone regeneration was observed in the defects of mice treated with as little as 0.19 mu g of bone morphogenetic protein-2 at 4 weeks after the transplantation. The local and sustained delivery system for bone morphogenetic protein-2 developed in the present study may represent a powerful modality for bone regeneration. (C) 2008, The Society for Biotechnology, Japan. All rights reserved.