Please cite this article in press as: M. Varshney et al., A label-free, microfluidics and interdigitated array microelectrode-based impedance biosensor in combination with nanoparticles immunoseparation for detection of Escherichia coli O157:H7 in food samples, Sens. Actuators B: Chem. (2007), doi:10.1016/j.snb.2007.03.045 ARTICLE IN PRESS +Model SNB-10055; No. of Pages 9 Sensors and Actuators B xxx (2007) xxx–xxx A label-free, microfluidics and interdigitated array microelectrode-based impedance biosensor in combination with nanoparticles immunoseparation for detection of Escherichia coli O157:H7 in food samples Madhukar Varshney a , Yanbin Li a,∗ , Balaji Srinivasan b , Steve Tung b a Department of Biological and Agricultural Engineering, University of Arkansas, Fayetteville, AR 72701, United States b Department of Mechanical Engineering, University of Arkansas, Fayetteville, AR 72701, United States Received 14 November 2006; received in revised form 12 March 2007; accepted 26 March 2007 Abstract A microfluidic flow cell with embedded gold interdigitated array microelectrode (IDAM) was developed and integrated with magnetic nanoparticle-antibody conjugates (MNAC) into an impedance biosensor to rapidly detect pathogenic bacteria in ground beef samples. The flow cell consisting of a detection microchamber and inlet and outlet microchannels was fabricated by bonding an IDAM chip to a poly(dimethylsiloxane) (PDMS) microchannel. The detection microchamber with a dimension of 6 mm × 0.5 mm × 0.02 mm and a volume of 60 nL was used to collect bacterial cells in the active layer above the microelectrode for sensitive impedance change. MNAC were prepared by conjugating streptavidin- coated magnetic nanoparticles with biotin-labeled polyclonal goat anti-E. coli antibodies and were used in the separation and concentration of target bacteria. The cells of E. coli O157:H7 inoculated in a food sample were first captured by the MNAC, separated, and concentrated by applying a magnetic field, washed, and then suspended in mannitol solution and finally injected through the microfluidic flow cell for impedance measurement. This impedance biosensor was able to detect as low as 1.6 × 10 2 and 1.2 × 10 3 cells of E. coli O157:H7 cells present in pure culture and ground beef sample, respectively. The total detection time from sampling to measurement was 35min. Equivalent circuit analysis indicated that the bulk medium resistance, double layer capacitance, and dielectric capacitance were responsible for the impedance change due to the presence of E. coli O157:H7 cells on the surface of IDAM. Sample pre-enrichment, secondary antibodies on the microelectrode surface, and redox probes were not required in this impedance biosensor. © 2007 Elsevier B.V. All rights reserved. Keywords: Impedance biosensor; E. coli O157:H7; Interdigitated array microelectrodes; Microfluidic flow cell; Label-free detection; Magnetic nanoparticles 1. Introduction The Centers for Disease Control and Prevention [1] esti- mates that foodborne diseases cause approximately 76 million illnesses, 325,000 hospitalizations, and 5000 deaths in the United States each year. USDA Economic Research Services [2] reported that medical costs, productivity losses, and costs of pre- mature deaths for diseases caused by major foodborne pathogens total to $6.9 billion per year. Among all pathogens, Escherichia coli O157:H7 is a leading cause of foodborne illness. Based on a 1999 estimate, 73,000 cases of infection and 61 deaths occur in the United States each year due to E. coli O157:H7 only [1]. ∗ Corresponding author. Tel.: +1 479 575 2424; fax: +1 479 575 7139. E-mail address: yanbinli@uark.edu (Y. Li). As the loss caused by E. coli O157:H7 is enormous in terms of medical cost and product recall, it is extremely important to rapidly and specifically detect E. coli O157:H7 in food products. Until recently, most biosensors studied for pathogenic bac- teria detection are label-dependent immunosensors that use labeled secondary antibodies to convert the antibody/antigen interaction into detectable optical or electrochemical signals. Label-free biosensors such as quartz crystal microbalance and surface plasmon resonance have attractive advantages with respect to speed, cost, and simplicity of operation. Impedance technique is yet another rapid and inexpensive alternative for label-free biosensors. Traditionally, macro-sized metal rods or wires were used as electrodes immersed in a medium to mea- sure impedance [3–5]. Several electrode geometries have been developed during the last decades to add functionalities, improve sensitivities, and lower the detection limits of impedance 0925-4005/$ – see front matter © 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.snb.2007.03.045