Abstract The blood-brain barrier (BBB) protects the human brain from external aggression. Despite its great importance, very few in vitro models of the BBB reproducing its complex organization are available yet. Here we fabricated such a three-dimensional (3D) self-organized in vitro model of BBB microvasculature by means of a combination of collagen microfibers (CMF) and fibrin gel. The interconnected fibers supported human brain microvascular endothelial cell migration and the formation of a capillary-like network with a lumen diameter close to in vivo values. Fibrin, a protein involved in blood vessel repair, favored the further 3D conformation of the brain microvascular endothelial cells, astrocytes and pericytes, ensured gel cohesion and avoided shrinkage. The maturation of the BBB microvasculature network was stimulated by both the CMF and the fibrin in the hydrogel. The expression of essential tight-junction proteins, carriers and transporters was validated in regards to bidimensional simple coculture. The volume of gel drops was easily tunable to fit in 96-well plates. The cytotoxicity of D-Mannitol and its impacts on the microvascular network were evaluated, as an example of the pertinence of this 3D BBB capillary model for screening applications.