It has recently been shown that some growth factors (GFs) have strong interactions with nonproteoglycan extracellular matrix proteins. Relevant here, the 12th-14th type three repeats of fibronectin (FN III12-14) have been shown to bind insulin-like growth factor binding-protein-3, fibroblast growth factor (FGF)-2, and vascular endothelial growth factor (VEGF)-A with high affinity. Since FN III12-14 is known to bind GFs from different families, we hypothesized that this domain could be highly promiscuous in its GF-binding capacity. We used biochemical approaches and surface plasmon resonance to investigate such interactions with recombinant FN III12-14. We found that FN III12-14 binds most of the GFs from the platelet-derived growth factor (PDGF)/VEGF and FGF families and some GFs from the transforming growth factor-β and neurotrophin families, with K(D) values in the nanomolar range, without inhibiting GF activity. Overall, 25 new binding interactions were identified. In a clinically relevant fibrin matrix, a fibrin-binding variant of FN III12-14 was highly effective as a GF delivery system. For instance, in matrices functionalized with FN III12-14, PDGF-BB-induced sprouting of human smooth muscle cell spheroids was greatly enhanced. We show that FN III12-14 is a highly promiscuous ligand for GFs and also holds great potential in clinical healing applications.