Quorum sensing (QS), the process of autoinducer-mediated cell-cell signaling among bacteria, facilitates biofilm formation, virulence, and many other multicellular phenotypes. QS inhibitors are being investigated as antimicrobials because of their potential to reduce symptoms of infectious disease while slowing the emergence of resistant strains. Autoinducer-2 (AI-2) analogs have been shown to inhibit genotypic QS responses among many bacteria. We demonstrate for the first time, the ability of C1-alkyl AI-2 analog, isobutyl-DPD, to significantly inhibit the maturation of Escherichia coli biofilms grown in vitro. Using a novel microfluidic device that incorporates dynamic, real-time measurements of biofilm density, we also show that a combinatorial approach wherein isobutyl-DPD ((S)-4,5-dihydroxy-2,3-pentanedione) is used with the antibiotic gentamicin is quite effective in rendering near complete clearance of pre-existing E. coli biofilms. Similarly, another AI-2 analog, phenyl-DPD, also used in combination with near MIC levels of gentamicin, resulted in clearance of preformed Pseudomonas aeruginosa biofilms. Clearance of pre-existing biofilms has remained a significant health care challenge; these results warrant consideration of a new approach based on the combination of "quenching" QS signal transduction processes with traditional antibiotic treatment.