In this study, we developed an automated strategy to manufacture an enzyme BFC powered by glucose/O2. The bioanode consists of GOx enzyme and PQQ redox mediator adsorbed over night on MWCNTs then deposited by means of AC-electrophoresis at 30 Hz and 160 Vp–p and, finally stabilized by electropolymerized polypyrrole. The biocathode is constructed from LAc enzyme and ABTS redox mediator adsorbed over night on MWCNTs, then electrophoretically deposited under AC-electric field at 30 Hz and 160 Vp–p and, finally stabilized by electrodeposited polypyrrole. The BFC was studied under air in phosphate buffer solution pH 7.4 containing 10 mM glucose and in human serum with 5 mM glucose addition at the physiological temperature of 37°C. Under these conditions, the maximum power density reaches 1.1 µW · mm−2 at a cell voltage of 0.167 V in buffer solution and 0.69 µW · mm−2 at cell voltage of 0.151 V in human serum. Such automated BFCs have a great potential to be optimized, miniaturized to micro and nanoscale devices suitable for in vivo studies. Biotechnol. Bioeng. 2012; 109:1601–1609. © 2012 Wiley Periodicals, Inc.