ELECTROKINETICS-ASSISTED ELECTRICAL SENSORS FOR RAPID DETECTION OF BACTERIA

Abstract

An array of microfabricated interdigitated electrodes (IDEs) is the most commonly used form of electrode geometry for dielectrophoretic manipulation of biological particles in microfluidic biochips owing to simplicity of fabrication and ease of analysis. However, the dielectrophoretic force dramatically reduces as the distance from the electrode surface increases; therefore, the effective region is usually close to the electrode surface for a given electric potential difference. Here, I present a novel two-dimensional computational method for generating planar electrode patterns with enhanced volumetric electric fields, which I call the “microelectrode discretization (MED)” method. It involves discretization and reconstruction of planar electrodes followed by selection of the electrode pattern that maximizes a newly defined objective function, factor S, which is determined by the electric potentials on the electrode surface alone. In this study, IDEs were used as test planar electrodes. Two arrays of IDEs and respective MED-optimized electrodes were implemented in microfluidic devices for the selective capture of Escherichia coli against 1-μm-diameter polystyrene beads, and I experimentally observed that 1.4 to 35.8 times more bacteria were captured using the MED-optimized electrodes than the IDEs (p < 0.0016), with a bacterial purity against the beads of more than 99.8%. This simple design method offered simplicity of fabrication, highly enhanced electric field, and uniformity of particle capture, and can be used for many dielectrophoresis-based sensors and microfluidic systems. Dielectrophoresis (DEP) is usually effective close to the electrode surface. Several techniques have been developed to overcome its drawbacks and to enhance dielectrophoretic particle capture. Here a simple technique was presented of superimposing alternating current DEP (high-frequency signals) and electroosmosis (EO; low-frequency signals) between two coplanar electrodes (gap: 25 μm) using a lab-made voltage adder for rapid and selective concentration of bacteria, viruses, and proteins, where the voltages and frequencies of DEP and EO were controlled separately. This signal superimposition technique enhanced bacterial capture (Escherichia coli K-12 against 1-μm-diameter polystyrene beads) more selectively (>99 %) and rapidly (~30 s) at lower DEP (5 Vpp) and EO (1.2 Vpp) potentials than those used in the conventional DEP capture studies. Nanometer-sized MS2 viruses and troponin I antibody proteins were also concentrated using the superimposed signals, and significantly more MS2 and cTnI-Ab were captured using the superimposed signals than the DEP (10 Vpp) or EO (2 Vpp) signals alone (p < 0.035) between the two coplanar electrodes and at a short exposure time (1 min). This technique has several advantages, such as simplicity and low cost of electrode fabrication, rapid and large collection without electrolysis. Electrokinetic technologies such as AC electro-osmosis (EO) and dielectrophoresis (DEP) have been used for effective manipulation of bacteria to enhance the sensitivity of an assay, and many previously reported electrokinetics-enhanced biosensors are based on stagnant fluids. An effective region for positive DEP for particle capture is usually too close to the electrode for the flowing particles to move toward the detection zone of a biosensor against the flow direction; this poses a technical challenge for electrokinetics-assisted biosensors implemented within pressure-driven flows, especially if the particles flow with high speed and if the detection zone is small. Here, a microfluidic single-walled carbon nanotubes (SWCNTs)-based field-effect transistor immunosensor was presented with electrohydrodynamic (EHD) focusing and DEP concentration for continuous and label-free detection of flowing Staphylococcus aureus in a 0.01× phosphate buffered saline (PBS) solution. The EHD focusing involved AC EO and negative DEP to align the flowing particles along lines close to the bottom surface of a microfluidic channel for facilitating particle capture downstream in the detection zone. For feasibility, 380-nm-diameter fluorescence beads suspended in 0.001× PBS were tested, and 14.6 times more beads were observed to be concentrated on the detection area with EHD focusing. Moreover, label-free, continuous, and selective measurement of S. aureus in 0.01× PBS was demonstrated, showing good linearity between the relative changes in electrical conductance of the SWCNTs and logarithmic S. aureus concentrations, a capture/detection time of 35 min, and limit of detection of 150 CFU/mL, as well as high specificity through electrical manipulation and biological interaction.