The fabrication of nanostructured electrodes for enzymatic glucose-oxygen biofuel cells is reported. The electrodes are based on enzyme encapsulation in sol-gel silica matrices and incorporate carbon nanotubes within the matrix to provide enhanced electronic conduction. The silica matrix is designed to be sufficiently porous that both glucose and oxygen have access to the enzymes and yet provides a protective cage for immobilizing the biomolecules without affecting biological function. Voltammetry experiments indicate that the effect of the silica matrix on mediator diffusion is minimal, although for one mediator, 2,2'-azino-bis ( 3-ethylbenzothiazoline-6-sulfonic acid ) diammonium salt, chemical modification of the solvent phase with polyethylene glycol is necessary. The polyethylene glycol addition also results in a more uniform dispersion of the carbon nanotubes. The enzymes maintain their biocatalytic activity in the sol-gel matrix. A glucose- oxygen biofuel cell based on the nanostructured silica sol-gel/carbon nanotube composite electrodes generates similar to 120 mu W/cm(2) at 0.24 V when operated at room temperature. (c) 2006 The Electrochemical Society.