In cardiovascular tissue engineering, synthetic or biologic scaffolds serve as templates for tissue development. Currently used scaffolds showing toxic degradation and immunogenic reactions are still far from ideal. We present a new alternative method to develop completely autologous human tissue without using any scaffold materials. Human vascular cells of arterial and venous origin were cultured to form cell sheets over a 4 week period under standard conditions. Thereafter, cell sheets of each origin were folded and cultured in a newly developed frame device for an additional 4 weeks. Controls remained under standard culture conditions. Tissue development was evaluated by morphology and biochemical assays. The formation of multilayered cell sheets and production of extracellular matrix were observed in all groups. Folded and framed neo-tissue showed a solid structure, with increased matrix formation and tissue organization when compared with the control groups. DNA content indicated significantly lower cell proliferation, and hydroxyproline assay indicated significantly higher collagen content in the framed cell sheets. We present a new approach to the engineering of cardiovascular tissue without the use of biodegradable scaffold material. Three-dimensional, completely autologous human tissue may be developed on the basis of this structure, thus avoiding scaffold induced toxic degradation or inflammatory reaction.