Tibiae of young rats were examined by scanning electron microscopy (SEM) and transmission electron microscopy (TEM) in order to study the origin and structure of matrix vesicles in periosteal bone. SEM studies have shown that periosteal osteoblasts have elongated processes with globular structures of 0.1 micron in diameter attached to the cell surface and processes. Similar structures were found to cover the inner surface of osteoblastic lacunae. The SEM studies have further shown that in the periosteal surface (forming bone) the above-described globules once mineralized, aggregated to form larger, nonhomogeneous mineralized spherules in which, by proper treatment with NaOCl, hydroxyapatite crystals could be exposed. Endosteal osteoblasts had fewer processes, devoid of the globular structures. Similarly, osteocytic and osteoclastic processes, although elongated and numberous, were not covered by the globular structures. In the matrix, collagen fibers of forming bone were randomly orientated, while in the deeper areas of bone they formed bundles with a longitudinal orientation. TEM studies have shown that the structures found on the osteoblastic surface and in the matrix are membrane-bound matrix vesicles which seem to be formed by budding from cell processes. Preformed membrane-bound vesicles were also observed by TEM inside sections of osteoblastic processes. These vesicles resembled the extracellular matrix vesicles in size and shape, thus giving the impression that at least some of the matrix vesicles are preformed cellular structures. While comparing SEM with TEM, it can be conducted that in bone, as in cartilage, matrix vesicles which probably serve as the initial locus of calcification, are formed directly by osteoblasts.