The nanocomposite enzymatic cathode and anode with carbon nanotube were fabricated and optimized for the application to biofuel cells. The nanocomposite electrodes consisted of polyion complex matrix where enzyme, mediator, and carbon nanotube as an electron transport enhancer were immobilized on the glassy carbon electrode. For the fabrication of anode and cathode, glucose oxidase and tetrathiafulvalene and bilirubin oxidase and 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium, respectively, were immobilized together in the polymer matrix. Electrochemical observation confirmed the bioelectrocatalytic ability of glucose oxidation and oxygen reduction at the anode and cathode, respectively, due to the enzymatic activity. The biological-fuel cell fabricated by combining the nanocomposite enzyme electrodes demonstrated open circuit voltage of 0.65 V, and its maximum power was 150 µW cm⁻².