Chondrocytes in monolayer cultures lose their phenotype and capability to express type-II collagen, they dedifferentiate into a fibroblastic cell type. Using three-dimensional culture systems a redifferentiation of these cells may occur. In the present study we investigated the morphology and biosynthetic activity of human articular chondrocytes seeded on porous matrices of type I/III collagen (Chondrogide, Geistlich Biomaterials, Wolhusen, Switzerland). Microscopical examinations showed that chondrocytes adhere firmly to a collagen-I/III-membrane exhibiting their characteristic spherical cell shape. Cell numbers after enzymatic digestion of the membrane showed a 93% recovery of seeded cells. Immunohistological examination revealed positive staining for type-II collagen in some areas. The generated biocomposite withstands mechanical stress, keeps its size and design and does not shrink in culture. It is therefore easy to handle, can be sutured, glued or fixed with pins. This study shows, that in vitro production of autologous cartilage-like tissue could be established using a bilayer collagen type I/III fleece. This biocomposite carries active chondrocytes and is currently being evaluated in vivo in a sheep model as well as in a clinical trial for the repair of localized cartilage defects in the knee.