Contractile C2C12 myotube line patterns embedded in a fibrin gel have been developed to afford a physiologically relevant and stable bioassay system. The C2C12 myotube/fibrin gel system was prepared by transferring a myotube monolayer from a glass substrate to a fibrin gel while retaining the original line patterns of myotubes. To endow the myotubes with contractile activity, a series of electrical pulses was applied through a pair of carbon electrodes placed at either side of a fibrin gel separately. The frequency and magnitude of myotube contraction were functions of the pulse frequency and duration, respectively. We found that the myotubes supported by an elastic fibrin gel maintained their line patterns and contractile activities for a longer period of time (1 week) than myotubes adhered on a conventional culture dish. Biotechnol. Bioeng. 2010; 105: 1161-1167. (C) 2009 Wiley Periodicals, Inc.