Louise M. Barrett

Rapid Microchip-Based Electrophoretic Immunoassays For The Detection Of Swine Influenza Virus.

Towards developing rapid and portable diagnostics for detecting zoonotic diseases, we have developed microchip-based electrophoretic immunoassays for sensitive and rapid detection of viruses. Two types of microchip-based electrophoretic immunoassays were developed. The initial assay used open channel electrophoresis and laser-induced fluorescence detection with a labeled antibody to detect influenza virus. However, this assay did not have adequate sensitivity to detect viruses at relevant concentrations for diagnostic applications. Hence, a novel assay was developed that allows simultaneous concentration and detection of viruses using a microfluidic chip with an integrated nanoporous membrane. The size-exclusion properties of the in situ polymerized polyacrylamide membrane are exploited to simultaneously concentrate viral particles and separate the virus/fluorescent antibody complex from the unbound antibody. The assay is performed in two simple steps--addition of fluorescently labeled antibodies to the sample, followed by concentration of antibody-virus complexes on a porous membrane. Excess antibodies are removed by electrophoresis through the membrane and the complex is then detected downstream of the membrane. This new assay detected inactivated swine influenza virus at a concentration four times lower than that of the open-channel electrophoresis assay. The total assay time, including device regeneration, is six minutes and requires <50 microl of sample. The filtration effect of the polymer membrane eliminates the need for washing, commonly required with surface-based immunoassays, increasing the speed of the assay. This assay is intended to form the core of a portable device for the diagnosis of high-consequence animal pathogens such as foot-and-mouth disease. The electrophoretic immunoassay format is rapid and simple while providing the necessary sensitivity for diagnosis of the illness state. This would allow the development of a portable, cost-effective, on-site diagnostic system for rapid screening of large populations of livestock, including sheep, pigs, cattle, and potentially birds.

Particle mixing and concentration through competing electrokinetic and hydrodynamic flows

We have developed a novel, low voltage particle concentration and separation paradigm that exploits the interplay between electrokinetic, dielectrophoretic, and pressure-driven flows. The devices presented utilize weak DC fields (5-25 V/cm) and patterned, insulating microfluidic channels. This approach has been applied to species varying in size by two orders of magnitude on the same chip (2 /spl mu/m-20 nm), can be applied to both biological and synthetic particles, and permits the channel geometry to be optimized to a specific size range.

Insulating dielectrophoresis for the continuous separation and concentration of Bacillus subtilis

This paper presents a novel microdevice for the dielectrophoretic manipulation of particles and cells for sample preparation and analysis. A two level isotropic etch of a glass substrate was used to create insulating ridges in micron sized channels. These ridges created a non-uniform field when a direct current field was applied across the channel and the dielectrophoretic force that resulted from the ridge was used to manipulate particles. We show the continuous concentration and separation of Bacillus subtilis from a two component sample mixture. When the applied voltage is at or above 30V/mm the flow of Bacillus subtilis was restricted to the central channel as a result of negative DEP away from the field concentration produced by the insulating ridges. Under the same applied electric fields the 200-nm polystyrene particles DEP away from the insulating ridges was negligible for the 200-nm particles, which flowed uninhibited down the three exit channels.

Continuous Particle Filtration and Concentration by Multigradient Dielectrophoresis

A novel methodology for designing selective particle concentrators in electrokinetic flows is presented. The technique is based on two-level etching of channels to produce ridges along which field gradients are patterned. The field gradients are then used to deflect particles using dielectrophoresis. Using uniform-field designs as a basis, fields in the vicinity of a single ridge are examined both experimentally and numerically. Although isotropic etching causes local deviations from piecewise continuous fields, ridges are found to serve as selective particle deflectors in experiments with both polystyrene beads and Bacillus subtilis. Sequences of parallel ridges are also tested, illustrating the efficacy of corrugated ridge structures for selective particle concentration.Copyright