A microbial fuel cell (MFC) was successfully developed as a toxicity biomonitoring system, giving a quick response to Cu(II) toxic events. The objective was to increase MFC sensitivity to Cu(II) toxicity by evaluating the impact of shear rate caused by mixing and intermittent nitrogen sparging on the biofilm structure. Low shear rate - achieved by continuously feeding the wastewater into the MFC at a low flow rate of 1.3 mL min(-1) during the enrichment period - resulted in low biomass density (124 g VSS L-1 of biofilm), high porosity and reduced levels of extracellular polymeric substances (EPS). Consequently, the sensitivity was improved. Scattered nitrogen sparging also increased the sensitivity by reducing the EPS level. It suggested that MFC enriched under low flow rate with intermittent nitrogen sparging could produce an anodic biofilm that was less dense, more porous, contained less EPS and ultimately displayed higher sensitivity to Cu(II) toxicity. (C) 2013 Elsevier Ltd. All rights reserved.