A dielectric-modulated field-effect transistor for biosensing

Interest in biosensors based on field-effect transistors (FETs), where an electrically operated gate controls the flow of charge through a semiconducting channel, is driven by the prospect of integrating biodetection capabilities into existing semiconductor technology. In a number of proposed FET biosensors, surface interactions with biomolecules in solution affect the operation of the gate or the channel. However, these devices often have limited sensitivity. We show here that a FET biosensor with a vertical gap is sensitive to the specific binding of streptavidin to biotin. The binding of the streptavidin changes the dielectric constant (and capacitance) of the gate, resulting in a large shift in the threshold voltage for operating the FET. The vertical gap is fabricated using simple thin-film deposition and wet-etching techniques. This may be an advantage over planar nanogap FETs, which require lithographic processing. We believe that the dielectric-modulated FET (DMFET) provides a useful approach towards biomolecular detection that could be extended to a number of other systems.     

A smart microfluidic affinity chromatography matrix composed of poly(N-isopropylacrylamide)-coated beads

The efficient upstream processing of complex biological or environmental samples for subsequent biochemical analysis remains a challenge in many analytical systems. New microfluidic platforms that provide multidiagnostic capabilities on single chips face a similar challenge in getting specific analytes purified or contaminants removed in different fluid streams. Here, stimuli-responsive polymers have been used to construct "smart" beads that can be reversibly immobilized on microfluidic channel walls to capture and release targets. The 100-nm latex beads were surface-modified with the temperature-sensitive polymer poly(N-isopropylacrylamide) (PNIPAAm). At room temperature, a suspension of these beads flows through a microfluidic channel constructed of poly(ethylene terephthalate). However, when the temperature in the channel is raised above the lower critical solution temperature (LCST) of PNIPAAm, the beads aggregate and adhere to the walls of the channel. The adhered beads are stable for long durations on the channel walls (demonstrated up to 70 min) in the presence of flow. The beads were further modified with the affinity moiety biotin, which tightly binds streptavidin. The dual-modified beads were adhered to the channel walls and functioned as a chromatographic affinity separation matrix, capable of binding streptavidin that was flowed through the microfluidic channel. Upon the reverse thermal stimulation to below the PNIPAAm LCST, the beads and captured streptavidin were observed to quickly dissolve and elute from the channel walls. This temperature-responsive affinity chromatography matrix can thus be flowed into a column and aggregated via temperature change, followed by the controlled release of affinity-captured targets back into the microfluidic flow stream.     

Nanofluidic concentration of selectively extracted biomolecule analytes by microtubules

We present a novel device for the selective extraction and high concentration of biomolecule analytes by integrating microtubules, one of cytoskeletal filaments, with nanofluidic technologies. Microtubules can be functionalized to provide numerous, nanoscopic binding sites for specific target biomolecules. The functionalized microtubules, with the target biomolecules bound to their surface, can be transported in the opposite direction to nontarget molecules using electrokinetic separation. Subsequently, the target molecule-bound microtubules are concentrated by a flat nanochannel structure, which filters the microtubules but allows ionic current and flow to pass through it. This device makes it possible to selectively extract target molecules such as streptavidin and bovine serum albumin and then highly concentrate them up to higher than 5 orders of magnitude from a complex mixture of analytes ranging from 1 nM to 10 fM. In addition, the device performs both extraction (separation) and concentration process simultaneously, which are typically performed in order in other devices, so that we significantly reduce analysis time and labor and even enable preconcentrated, identified target molecules to be available for postanalysis. Thus, we believe that the use of functionalized microtubules with nanofluidics will be a useful means to facilitate biochemical analysis systems.     

Continuous-flow particle separation by 3D Insulative dielectrophoresis using coherently shaped, dc-biased, ac electric fields

We present the development of a continuous-flow, "dielectrophoretic spectrometer" based on insulative DEP techniques and three-dimensional geometric design. Hot-embossed thermoplastic devices allow for high-throughput analysis and geometric control of electric fields via ridged microstructures patterned in a high width-to-depth aspect ratio (250:1) channel. We manipulate particles with dc-biased, ac electric fields and generate continuous-output streams of particles with a transverse outlet position specified by linear and nonlinear particle mobilities. We show, with simulation and experiment, that characteristic shape factors can be defined that capture the effects of constrictions in channel depth and that modulating the angle of these constrictions changes the resulting local DEP force. Microdevices are fabricated with an insulative constriction in channel depth, whose angle of incidence with the direction of flow varies continuously across the channel width. The resulting electric field gradients enable demonstration of a dielectrophoretic spectrometer that separates particles and controls their transverse channel position.     

A microfluidics approach to the problem of creating separate solution environments accessible from macroscopic volumes

We report on a microfluidic device that generates separate solution environments in macroscopic volumes. Spatially distinct patterns are created by emitting fluids from 16 different sources (closely spaced microchannels) into a solution-filled macroscopic chamber. The fluid in neighboring microchannels couples viscously in the macroscopic container, generating one single interdigitated stream. Scanning nanoelectrode amperometry was used for characterizing the concentration landscape and the diffusion zones between solutions running in parallel at different coordinates in the stream. These experiments were complemented by finite element simulations of the Navier-Stokes and mass transport equations to describe the velocity distributions and the diffusion behavior. For in channel flow velocities of 50 mm.s(-1), patterns could persist on the order of millimeters to centimeters in the open volume. The most narrow diffusion zones with widths less than 10 microm (5-95% concentration change) were found some tens of micrometers out in the macroscopic container. We demonstrate that a 14-microm-diameter nearly spherical object (biological cell) attached to a micropipet can be moved from one solution environment to another by a lateral displacement of only 8 microm. The device is suitable for applications where the solution environment around a microscopic or nanoscopic sensor needs to be changed multiple times, i.e., in order to build layered structures, for obtaining binding isotherms, and kinetic information, for example, on ion channels, enzymes, and receptors as well as in applications where different loci on an object need to be exposed to different environments or where complex solution environments need to be created for studies of interfacial chemistry between two streaming layers.     

Nanoscale adhesion,friction and wear studies of biomolecules on silane polymer-coated silica and alumina-based surfaces

Proteins on biomicroelectromechanical systems (BioMEMS) confer specific molecular functionalities. In planar FET sensors (field-effect transistors, a class of devices whose protein-sensing capabilities we demonstrated in physiological buffers), interfacial proteins are analyte receptors, determining sensor molecular recognition specificity. Receptors are bound to the FET through a polymeric interface, and gross disruption of interfaces that removes a large percentage of receptors or inactivates large fractions of them diminishes sensor sensitivity. Sensitivity is also determined by the distance between the bound analyte and the semiconductor. Consequently, differential properties of surface polymers are design parameters for FET sensors. We compare thickness, surface roughness, adhesion, friction and wear properties of silane polymer layers bound to oxides (SiO₂ and Al₂O₃, as on AlGaN HFETs). We compare those properties of the film–substrate pairs after an additional deposition of biotin and streptavidin. Adhesion between protein and device and interfacial friction properties affect FET reliability because these parameters affect wear resistance of interfaces to abrasive insult in vivo. Adhesion/friction determines the extent of stickage between the interface and tissue and interfacial resistance to mechanical damage. We document systematic, consistent differences in thickness and wear resistance of silane films that can be correlated with film chemistry and deposition procedures, providing guidance for rational interfacial design for planar AlGaN HFET sensors.     

Two-color mixing for classifying agricultural products for safety and quality

We show that the chromaticness of the visual signal that results from the two-color mixing achieved through an optically enhanced binocular device is directly related to the band ratio of light intensity at the two selected wavebands. A technique that implements the band-ratio criterion in a visual device by using two-color mixing is presented here. The device will allow inspectors to identify targets visually in accordance with a two-wavelength band ratio. It is a method of inspection by human vision assisted by an optical device, which offers greater flexibility and better cost savings than a multispectral machine vision system that implements the band-ratio criterion. With proper selection of the two narrow wavebands, discrimination by chromaticness that is directly related to the band ratio can work well. An example application of this technique for the inspection of carcasses chickens of afficted with various diseases is given. An optimal pair of wavelengths of 454 and 578 nm was selected to optimize differences in saturation and hue in CIE LUV color space among different types of target. Another example application, for the detection of chilling injury in cucumbers, is given, here the selected wavelength pair was 504 and 652 nm. The novel two-color mixing technique for visual inspection can be included in visual devices for various applications, ranging from target detection to food safety inspection.     

Accumulation and separation of membrane-bound proteins using hydrodynamic forces

The separation of molecules residing in the cell membrane remains a largely unsolved problem in the fields of bioscience and biotechnology. We demonstrate how hydrodynamic forces can be used to both accumulate and separate membrane-bound proteins in their native state. A supported lipid bilayer (SLB) was formed inside a microfluidic channel with the two proteins streptavidin (SA) and cholera toxin (CT) coupled to receptors in the lipid bilayer. The anchored proteins were first driven toward the edge of the lipid bilayer by hydrodynamic forces from a flowing liquid above the SLB, resulting in the accumulation of protein molecules at the edge of the bilayer. After the concentration process, the bulk flow of liquid in the channel was reversed and the accumulated proteins were driven away from the edge of the bilayer. Each type of protein was found to move at a characteristic drift velocity, determined by the frictional coupling between the protein and the lipid bilayer, as well as the size and shape of the protein molecule. Despite having a similar molecular weight, SA and CT could be separated into monomolecular populations using this approach. The method also revealed heterogeneity among the CT molecules, resulting in three subpopulations with different drift velocities. This was tentatively attributed to multivalent interactions between the protein and the monosialoganglioside G(M1) receptors in the lipid bilayer.     

Diffusion-limited patterning of molecules in nanofluidic channels

Diffusion-limited patterning (DLP) is a new technique that enables patterning of labile molecular species in solution phase onto surfaces that are not easily accessible. This technique is self-aligning and is simple to implement for patterning multiple species. We demonstrated DLP by patterning alternating bands of fluorescently labeled and unlabeled streptavidin in biotin-functionalized nanofluidic channels with spatial resolution better than 1 microm. The methodology of DLP also enables experimental measurement of a unique parameter that relates molecular surface grafting density, concentration, diffusivity, and channel geometry.     

Three-color mixing for classifying agricultural products for safety and quality

A three-color mixing application for food safety inspection is presented. It is shown that the chromaticness of the visual signal resulting from the three-color mixing achieved through our device is directly related to the three-band ratio of light intensity at three selected wavebands. An optical visual device using three-color mixing to implement the three-band ratio criterion is presented. Inspection through human vision assisted by an optical device that implements the three-band ratio criterion would offer flexibility and significant cost savings as compared to inspection with a multispectral machine vision system that implements the same criterion. Example applications of this optical three-color mixing technique are given for the inspection of chicken carcasses with various diseases and for apples with fecal contamination. With proper selection of the three narrow wavebands, discrimination by chromaticness that has a direct relation with the three-band ratio can work very well. In particular, compared with the previously presented two-color mixing application, the conditions of chicken carcasses were more easily identified using the three-color mixing application. The novel three-color mixing technique for visual inspection can be implemented on visual devices for a variety of applications, ranging from target detection to food safety inspection.     

Patterned solvent delivery and etching for the fabrication of plastic microfluidic devices

A very simple method for micropatterning flat plastic substrates that can be used to build microfluidic devices is demonstrated. Patterned poly(dimethylsiloxane) elastomer is used as a template to control the flow path of an etching solvent through a channel design to be reproduced on the plastic substrate. The etching solvent was a acetone/ethanol mixture for poly(methyl methacrylate) substrates or a dimethylformamide/acetone mixture for polystyrene. The method is extremely fast in that duplicate plastic substrates can be patterned in just a few minutes each. We identified conditions that lead to smooth channel surfaces and characterized the rate of etching under these conditions. We determined that, for sufficiently short etching times (shallow channel depths), the etch rate is independent of the linear flow rate. This is very important since it means that the etch depth is approximately constant even in complex channel geometries where there will be a wide range of etchant flow rates at different positions in the pattern to be reproduced. We also demonstrate that the method can be used to produce channels with different depths on the same substrate as well as channels that intersect to form a continuous fluid junction. The method provides a nice alternative to existing methods to rapidly fabricate microfluidic devices in rigid plastics without the need for specialized equipment.     

Silicon nanoribbons for electrical detection of biomolecules

Direct electrical detection of biomolecules at high sensitivity has recently been demonstrated using semiconductor nanowires. Here we demonstrate that semiconductor nanoribbons, in this case, a thin sheet of silicon on an oxidized silicon substrate, can approach the same sensitivity extending below the picomolar concentration regime in the biotin/streptavidin case. This corresponds to less than approximately 20 analyte molecules bound to receptors on the nanoribbon surface. The micrometer-size lateral dimensions of the nanoribbon enable optical lithography to be used, resulting in a simple and high-yield fabrication process. Electrical characterization of the nanoribbons is complemented by computer simulations showing enhanced sensitivity for thin ribbons. Finally, we demonstrate that the device can be operated both in inversion as well as in accumulation mode and the measured differences in detection sensitivity are explained in terms of the distance between the channel and the receptor coated surface with respect to the Debye screening length. The nanoribbon approach opens up for large scale CMOS fabrication of highly sensitive biomolecule sensor chips for potential use in medicine and biotechnology.     

Apta-biosensors for nonlabeled real time detection of human IgE based on carbon nanotube field effect transistors

In this study, we demonstrated the aptamer-based biosensor (apta-biosensor) using CNT-FET devices for label free detection of allergy diagnosis by IgE detection. In order to detect the IgE, two kinds of receptor (monoclonal IgE antibody and anti-IgE aptamer)-modified CNT-FET devices were fabricated. The binding event of the target IgE onto receptors was detected by monitoring the gating effect caused by the charges of the target proteins. Since the CNT-FET biosensors were used in buffer solution, it was crucial to use small-size receptors like aptamers than whole antibodies so that the charged target IgE could approach the CNT surface within the Debye length distance to give a large gating effect. The results show that CNT-FET biosensors using monoclonal IgE antibody had very low sensitivity (minimum detectable level 1000 ng/mL), while those based on anti-IgE aptamer could detect 50 ng/mL. Moreover, the aptamer-modified CNT-FET herein could successfully block non-target proteins and could selectively detect the target protein in an environment similar to human serum electrolyte. Therefore, aptamer-based CNT-FET devices enable the production of label-free ultrasensitive electronic biosensors to detect clinically important biomarkers for disease diagnosis.     

Surface tension driven flows in shallow layers due to heat or mass transfer to the free surface

In a number of physical systems a surface tension driven flow is established in a shallow layer of liquid as a result of heat or mass transfer to the free surface. Such transfer processes often produce a thin temperature or concentration boundary layer near the free surface. We have considered the relatively simple situation when this occurs in shallow two-dimensional channel flow under steady conditions. It is shown that the properties of the boundary layer can be obtained by solving a sequence of parabolic partial differential equations and that the shape of the free surface results from the solution of an integral equation. The simple case of uniform surface transfer has been considered, but the analysis developed can be extended to more complex situations.     

ATTRIBUTES FOR EXPEDIENT COMPUTER VISION INSPECTION

The visual inspection of parts as they progress through die manufacturing process is an important task in all industries. Visual inspection, when performed by humans is a tedious task and is prone to error. This is precisely what makes it a good candidate for automation. Although computer vision systems have been around for over 30 years, die industrial applications of vision systems have become practical only in die last decade. Image processing and pattern recognition algorithms used in industrial vision systems are built upon a broad body of knowledge in vision research. But the use of computer vision systems in quality control has been limited to replicating die visual inspection tasks as they would be performed by a human operator. It is die contention of this study that when computerized inspection is employed, quality control inspection plans suitable for computerized inspection should also be employed to assure cost-effectiveness.

We study a simple gauging inspection task and propose a quality control plan that exploits die characteristics of computer vision systems in order to improve cost-effectiveness.     

Control of nanogap separation by surface-catalyzed chemical deposition

The nanogap devices, which comprise multiple electrodes separated by a few to a few tens of nanometers, have opened up new possibilities in biomolecular sensing as well as various frontier electronics. One of the key aspects of the nanogap device research is how to control the gap distance following each specific needs of the gap structure. Here, we report the extensive study on the fine control of the gap distance between electrodes within the range of 1-80 nm via surface-catalyzed chemical deposition. The initial gap electrodes were prepared via conventional e-beam lithography, and the gap distance was narrowed to a designed value through the surface-catalyzed reduction of gold ion on the predefined electrode surfaces, by simple dipping of the electrodes into the aqueous solution of gold chloride and hydroxylamine. The final gap distance was controlled by adjusting the repetition number, reductant concentration, reaction time, and reaction temperature. The dependence of the gap-narrowing reaction on these parameters was systematically examined based on the results of field emission scanning electron microscopy and atomic-force microscopy.     

An experimental investigation of the cavitational characteristics of converging nozzles

We consider problems associated with the appearance and development of cavitation in the flow of a liquid through channels with local constriction. For the case of separation flow we have derived an equation which associates the parameters of three sections of the channel: at the inlet, at the cross section of the stream, and at the outlet; this equation has been confirmed through experimentation on cold and hot water, as well as on kerosene.     

Stabilization of high-resistance seals in patch-clamp recordings by laminar flow

The formation of a high-resistance electrical seal between a cell membrane and a glass micropipet tip is essential in patch-clamp experiments. We have studied the electrical properties and the mechanical stability of the seal using a microfluidic chip generating laminar flow in open volumes. We show that, by using fluid flow (1-10 mm/s) acting along the symmetry axis of the cell-pipet, seals of a higher mechanical stability with increased resistances can be achieved, allowing up to 100% longer recording times and over 40% decreased noise levels (Irms). These improved properties are beneficial for high-sensitivity patch-clamp recordings, in particular, in longtime studies of ion channel receptor systems that are relevant in biosensor applications of the technique. Furthermore, these observations support the combination of patch-clamp with microfluidic devices, for example, for rapid solution exchange around a single cell sensor for high-throughput electrophysiology and for highly resolved kinetic studies.     

Flow rate optimization for the quadrupole magnetic cell sorter.

The quadrupole magnetic cell sorter is a form of split-flow thin-channel (SPLITT) separation device. It employs a quadrupole magnetic field and annular channel geometry. Immunomagnetic labels are used to bind to specific receptors on the surface of the cells of interest. It is the interaction of these labels with the magnetic field that brings about the selective isolation of these cells. The SPLITT separation devices have generally been based on parallel-plate geometry, usually with effectively constant field strength applied across the channel thickness. The nonconstant field strength and annular channel geometry of the magnetic cell sorter require that a new strategy be developed for optimization of inlet and outlet flow rates. We present such a strategy here based on a consideration of certain specific cell trajectories within the system.