Transforming growth factor beta (TGF-beta) regulates cellular growth and differentiation and stimulates the synthesis and secretion of protein constituents of the extracellular matrix. Three isoforms of TGF-beta have been found in mammals. Although the biological activities of TGF-beta 1, TGF-beta 2, and TGF-beta 3 are similar at the level of cell culture, distinct in vivo functions for these molecules are emerging. To gain insight into the role of each isoform in wound repair, antibodies specific for each isoform of TGF-beta were used to examine excisional wound repair. Marked differences in the temporal and spatial relationships for immunoreactive TGF-beta 1, -beta 2, and -beta 3 were noted throughout the repair process. TGF-beta 2 and TGF-beta 3 were prevalent by 24 hours after excisional wounding, and strong immunoreactivity was observed in the migrating epidermis. Subtle changes in immunoreactivity occurred for TGF-beta 2 and TGF-beta 3 in cells of the epidermal appendages, mesenchymal derivatives, granulation tissue, and the underlying dermis throughout wound repair. In contrast, TGF-beta 1 was not associated with any undifferentiated cells and was not present in the dermis and most dermal structures in both nonwounded skin or wounds until day 5 after wounding, when re-epithelialization was complete. Following re-epithelialization, TGF-beta 2 and TGF-beta 3 were present in all four layers of stratum corneum of the differentiating epidermis. All three TGF-beta isoforms were present in mesenchymal cells and basal lamina, suggesting their role in the modulation of dermal-epidermal interaction during wound repair. Our observations support individual in vivo function for TGF-beta isoforms in cutaneous wound repair.