Chorioallantoic membrane (CAM) and chorioallantoic fluid (CAF) of the chick embryo were studied for the presence of immunoreactive and biologically active basic fibroblast growth factor (bFGF) from Day 6 to Day 18 of incubation. An immunoreactive M(r) 16,000 bFGF-like molecule was detected both in CAM and in CAF. This molecule was identified as bFGF on the basis of its molecular weight, its affinity for heparin, and its capacity to induce plasminogen activator production in cultured endothelial GM 7373 cells. The levels of biologically active and immunoreactive bFGF vary in CAM and CAF during embryonic development, maximal concentrations being observed between Days 10 and 14 of incubation. At all time points investigated, absolute concentrations of bFGF were significantly higher in CAM (ranging from 25 to 183 ng/g of wet tissue) than in CAF (ranging from 0.2 to 4 ng/ml). In a parallel series of experiments performed at Day 8 and evaluated at Day 12 of chick embryo development, human recombinant bFGF and neutralizing anti-bFGF antibody were investigated for their capacity to affect the vasoproliferative processes of the CAM. The two molecules either were applied onto the surface of the CAM or were injected into the allantoic sac. When bFGF or anti-bFGF antibodies were absorbed on methylcellulose discs and applied on the top of the CAM, they exerted a strong angiogenic or anti-angiogenic effect, respectively. On the contrary, when bFGF or the corresponding neutralizing antibody was injected into the allantoic sac, no modifications of the vasoproliferative processes of the CAM were observed at either the macroscopic or the microscopic level. These results provide evidence indicating that endogenous bFGF has a rate-limiting role in the vascularization of the CAM during chick embryogenesis. bFGF located within the CAM, rather than that present in the CAF, appears to be involved in this developmental process.