Specific antiserum was used to investigate the distribution of the extracellular glycoprotein, fibronectin, in rat skin and tongue tissue by light and electron microscopy with immunofluorescence and immunoperoxidase techniques. We conclude that fibronectin is absent from stable, differentiated parts of tissues, such as the sebaceous glands or the matrix, medulla, cortex, and cuticles of the hair and the inner and outer root sheaths, or even in tissues in which there is some cell movement, such as the epidermis. It is, however, characteristic of sites at which cell division is occurring in contact with an extracellular scaffolding, such as basement membrane or loose connective tissue. Conspicuous examples were in the glassy membrane and connective tissue sheath associated with the follicular epithelium, the basement membrane underlying vascular endothelial cells, the connective tissues surrounding and investing nerve and muscle fibre bundles, and the dermal connective tissue where fibronectin was often associated closely with collagen fibres. At the basement membrane of the dermal/epidermal junction, fibronectin occurred at the plasma membrane of the basal cells and in the lamina lucida area. There was no correlation with specific areas of cell-substrate adhesion, such as the hemidesmosomes. The endoplasmic reticulum of fibroblasts stained strongly suggesting that these cells represent a major site of synthesis.