An experimental study was done to evaluate factors influencing guided regeneration of bone in standardized calvarial bony defect. An 8 mm circular transosseous calvarial bony defect was made. Various material such as demineralized freeze-dried bone (DFDB), BioMesh , Millipore filter and its combination was placed in the bony defect. A sequential histopathologic, histochemical, immunohistochemical, and histomorphometric studies were done on the guided bone regeneration in the calvarial bony defect. Bone formation was sigificantly enhanced when the DFDB was retained within the bony defect with a protective bioabsorbable membrane. Inframembranous DFDB-filling was required to prevent collapse of the membrane and preserve spaces for bone regeneration. The bioabsorbable membrane should presumably remain intact for longer than at least 5 weeks to facilitate bone regeneration. The new bone formation was dependent on the barrier-effect (preserving secluded spaces) and inflammation-inducing property of membrane, and guiding bone regeneration of the grafts. Macrophages recruited by grafts were partly involved in decrease of bone regeneration via the sequential events of release of fibronectin, chemotactic effect of the fibronectin to fibroblasts, and collagen lay-down.