Laccase, a blue multicopper oxidoreductase enzyme, is a robust enzyme that catalyzes the reduction of oxygen to water and has been shown previously to perform improved direct electron transfer in a biocathode when mixed with anthracene-modified multi-walled carbon nanotubes. Previous cathode construction used crude laccase enzyme isolated as a brown cell extract powder containing both active and inactive proteins. Purification of this enzyme, yielding a blue solution, resulted in greatly improved enzyme activity and removed insulating protein that competed for docking space in this cathodic system. Cyclic voltammetry of the purified biocathodes showed a background subtracted limiting current density of 1.84 (+/- 0.05) mA/cm(2) in a stationary air-saturated system. Galvanostatic and potentiostatic stability experiments show that the biocathode maintains up to 75% and 80% of the original voltage and current respectively over 24 hours of constant operation. Inclusion of the biocathode in a glucose/O-2 biofuel cell using a mediated glucose oxidase (GOx) anode produced maximum current and power densities of 1.28 (+/- 0.18) mA/cm(2) and 281 (+/- 50) mu W/cm(2) at 25 degrees C and 1.80 (+/- 0.06) mA/cm(2) and 381 (+/- 33) mu W/cm(2) at 37 degrees C, respectively. Enzymatic efficiency of this glucose/O-2 enzymatic fuel cell is among the highest reported for a glucose/O-2 enzymatic fuel cell. (C) 2012 The Electrochemical Society. [DOI: 10.1149/2.062212jes] All rights reserved.