Probing the effect of polymer molecular weight on penetration into the wood cell wall using polyethylenimine (PEI) as a model compound.

Decking is one of the largest applications for the treated wood market. The most challenging property to obtain for treated wood is dimensional stability, which can be achieved, in part, by cell wall bulking, cell wall polymer crosslinking and removal of hygroscopic components in the cell wall. A commonly accepted key requirement is for the actives to infuse through the cell wall, which has a microporosity of ∼5-13 nm. Equally as challenging is being able to measure and quantify the cell wall penetration. Branched polyethylenimine (PEI) was studied as a model polymer for penetration due to its water solubility, polarity, variable molecular weight ranges, and ability to form a chelation complex with preservative metals to treat lumbers. Two different molecular weight polyethylenimines (PEI), one with a weight average molecular weight (Mw) equal to 800 Da and the other 750 000 Da, were investigated for penetration by microscopy and spectroscopy techniques. Analytical methods were developed to both create smooth interfaces and for relative quantitation and visualisation of PEI penetration into the wood. The results showed both PEI with Mw of 800 Da and PEI with Mw of 750 000 Da coated the lumens in high density. However, only the PEI with Mw of 800 appeared to penetrate the cell walls in sufficient levels. Literature has shown the hydrodynamic radii of PEI 750 000 is near 29 nm, whereas a smaller PEI at 25 K showed 4.5 nm. Most importantly the results, based on methods developed, show how molecular weight and tertiary structure of the polymer can affect its penetration, with the microporosity of the wood being the main barrier.

Micro-patterning of mammalian cells on suspended MEMS resonant sensors for long-term growth measurements.

MEMS resonant mass sensors can measure the mass of individual cells, though long-term growth measurements are limited by the movement of cells off the sensor area. Micro-patterning techniques are a powerful approach to control the placement of individual cells in an arrayed format. In this work we present a method for micro-patterning cells on fully suspended resonant sensors through select functionalization and passivation of the chip surface. This method combines high-resolution photolithography with a blanket transfer technique for applying photoresist to avoid damaging the sensors. Cells are constrained to the patterned collagen area on the sensor by pluronic acting as a cell adhesion blocker. This micro-patterning method enables long-term growth measurements, which is demonstrated by a measurement of the change in mass of a human breast cancer cell over 18 h.

Effect of biointerfacing linker chemistries on the sensitivity of silicon nanowires for protein detection.

Point-of-care diagnostics show promise in removing reliance on centralized lab testing facilities and may help increase both the survival rate for infectious diseases as well as monitoring of chronic illnesses. CMOS compatible diagnostic platforms are currently being considered as possible solutions as they can be easily miniaturized and can be cost-effective. Top-down fabricated silicon nanowires are a CMOS-compatible technology which have demonstrated high sensitivities in detecting biological analytes, such as proteins, DNA, and RNA. However, the reported response of nanowires to these analytes has varied widely since several different functionalization protocols have been attempted with little characterization and comparison. Here we report protocols for fabrication and functionalization of silicon nanowires which yield highly stable nanowires in aqueous solutions and limits of detection to ∼1 pg/mL of the model protein used in the study. A thorough characterization was done into optimizing the release of the silicon nanowires using combined dry and wet etch techniques, which yielded nanowires that could be directly compared to increase output statistics. Moreover, a range of different linker chemistries were tried for reacting the primary antibody, and its response to target and nonspecific antigens, with polyethylene glycol based linker BS(PEG)5 providing the best response. Consequently, this chemistry was used to characterize different oxide thicknesses and their responses to the mouse IgG antigen, which with the smallest oxide thickness yielded 0.1-1 pg/mL limits of detection and a dynamic range over 3 orders of magnitude.

On-chip parallel detection of foodborne pathogens using loop-mediated isothermal amplification.

According to estimates issued by the Center for Disease Control and Prevention, one out of six Americans will get sick during this year due to consumption of contaminated products and there will be 50,000 related hospitalizations. To control and treat the responsible foodborne diseases, rapid and accurate detection of pathogens is extremely important. A portable device capable of performing nucleic acid amplification will enable the effective detection of infectious agents in multiple settings, leading to better enforcement of food safety regulations. This work demonstrates the multiplexed detection of food pathogens through loop-mediated isothermal amplification on a silicon chip. Silane passivation is used to prevent the adsorption of the polymerase on silicon oxide, which can severely inhibit nucleic acid amplification. We demonstrate the multiplexed screening of virulence genes of Listeria monocytogenes, Escherichia coli, and Salmonella by dehydrating the corresponding primers in oxidized silicon wells. Droplets of 30 nL with reagents for nucleic acid amplification and lysate of suspected pathogens are arrayed on micro-machined wells with an automated microinjection system. We show that dehydrated primers re-suspend when other reagents are microinjected, and the resulting mix can be used to specifically amplify the targeted gene. Results of characterization experiments demonstrate sensitivity down to a few templates per reaction, specificity that enables multiplexed screening, and robustness that allows amplification without DNA extraction.

Ultralocalized thermal reactions in subnanoliter droplets-in-air.

Miniaturized laboratory-on-chip systems promise rapid, sensitive, and multiplexed detection of biological samples for medical diagnostics, drug discovery, and high-throughput screening. Within miniaturized laboratory-on-chips, static and dynamic droplets of fluids in different immiscible media have been used as individual vessels to perform biochemical reactions and confine the products. Approaches to perform localized heating of these individual subnanoliter droplets can allow for new applications that require parallel, time-, and space-multiplex reactions on a single integrated circuit. Our method positions droplets on an array of individual silicon microwave heaters on chip to precisely control the temperature of droplets-in-air, allowing us to perform biochemical reactions, including DNA melting and detection of single base mismatches. We also demonstrate that ssDNA probe molecules can be placed on heaters in solution, dried, and then rehydrated by ssDNA target molecules in droplets for hybridization and detection. This platform enables many applications in droplets including hybridization of low copy number DNA molecules, lysing of single cells, interrogation of ligand-receptor interactions, and rapid temperature cycling for amplification of DNA molecules.

Ultra-localized single cell electroporation using silicon nanowires.

Analysis of cell-to-cell variation can further the understanding of intracellular processes and the role of individual cell function within a larger cell population. The ability to precisely lyse single cells can be used to release cellular components to resolve cellular heterogeneity that might be obscured when whole populations are examined. We report a method to position and lyse individual cells on silicon nanowire and nanoribbon biological field effect transistors. In this study, HT-29 cancer cells were positioned on top of transistors by manipulating magnetic beads using external magnetic fields. Ultra-rapid cell lysis was subsequently performed by applying 600-900 mV(pp) at 10 MHz for as little as 2 ms across the transistor channel and the bulk substrate. We show that the fringing electric field at the device surface disrupts the cell membrane, leading to lysis from irreversible electroporation. This methodology allows rapid and simple single cell lysis and analysis with potential applications in medical diagnostics, proteome analysis and developmental biology studies.

Coupled heterogeneous nanowire-nanoplate planar transistor sensors for giant (>10 V/pH) Nernst response.

We offer a comprehensive theory of pH response of a coupled ISFET sensor to show that the maximum achievable response is given by ΔV/ΔpH = 59 mV/pH × α, where 59 mV/pH is the intrinsic Nernst response and α an amplification factor that depends on the geometrical and electrical properties of the sensor and transducer nodes. While the intrinsic Nernst response of an electrolyte/site-binding interface is fundamental and immutable, we show that by using channels of different materials, areas, and bias conditions, the extrinsic sensor response can be increased dramatically beyond the Nernst limit. We validate the theory by measuring the pH response of a Si nanowire-nanoplate transistor pair that achieves >10 V/pH response and show the potential of the scheme to achieve (asymptotically) the theoretical lower limit of signal-to-noise ratio for a given configuration. We suggest the possibility of an even larger pH response based on recent trends in heterogeneous integration on the Si platform.

Silicon nanowires with high-k hafnium oxide dielectrics for sensitive detection of small nucleic acid oligomers.

Nanobiosensors based on silicon nanowire field effect transistors offer advantages of low cost, label-free detection, and potential for massive parallelization. As a result, these sensors have often been suggested as an attractive option for applications in point-of-care (POC) medical diagnostics. Unfortunately, a number of performance issues, such as gate leakage and current instability due to fluid contact, have prevented widespread adoption of the technology for routine use. High-k dielectrics, such as hafnium oxide (HfO(2)), have the known ability to address these challenges by passivating the exposed surfaces against destabilizing concerns of ion transport. With these fundamental stability issues addressed, a promising target for POC diagnostics and SiNWFETs has been small oligonucleotides, more specifically, microRNA (miRNA). MicroRNAs are small RNA oligonucleotides which bind to mRNAs, causing translational repression of proteins, gene silencing, and expressions are typically altered in several forms of cancer. In this paper, we describe a process for fabricating stable HfO(2) dielectric-based silicon nanowires for biosensing applications. Here we demonstrate sensing of single-stranded DNA analogues to their microRNA cousins using miR-10b and miR-21 as templates, both known to be upregulated in breast cancer. We characterize the effect of surface functionalization on device performance using the miR-10b DNA analogue as the target sequence and different molecular weight poly-l-lysine as the functionalization layer. By optimizing the surface functionalization and fabrication protocol, we were able to achieve <100 fM detection levels of the miR-10b DNA analogue, with a theoretical limit of detection of 1 fM. Moreover, the noncomplementary DNA target strand, based on miR-21, showed very little response, indicating a highly sensitive and highly selective biosensing platform.

Label-free electrical detection of pyrophosphate generated from DNA polymerase reactions on field-effect devices.

We introduce a label-free approach for sensing polymerase reactions on deoxyribonucleic acid (DNA) using a chelator-modified silicon-on-insulator field-effect transistor (SOI-FET) that exhibits selective and reversible electrical response to pyrophosphate anions. The chemical modification of the sensor surface was designed to include rolling-circle amplification (RCA) DNA colonies for locally enhanced pyrophosphate (PPi) signal generation and sensors with immobilized chelators for capture and surface-sensitive detection of diffusible reaction by-products. While detecting arrays of enzymatic base incorporation reactions is typically accomplished using optical fluorescence or chemiluminescence techniques, our results suggest that it is possible to develop scalable and portable PPi-specific sensors and platforms for broad biomedical applications such as DNA sequencing and microbe detection using surface-sensitive electrical readout techniques.

High-k dielectric Al2O3 nanowire and nanoplate field effect sensors for improved pH sensing.

Over the last decade, field-effect transistors (FETs) with nanoscale dimensions have emerged as possible label-free biological and chemical sensors capable of highly sensitive detection of various entities and processes. While significant progress has been made towards improving their sensitivity, much is yet to be explored in the study of various critical parameters, such as the choice of a sensing dielectric, the choice of applied front and back gate biases, the design of the device dimensions, and many others. In this work, we present a process to fabricate nanowire and nanoplate FETs with Al(2)O(3) gate dielectrics and we compare these devices with FETs with SiO(2) gate dielectrics. The use of a high-k dielectric such as Al(2)O(3) allows for the physical thickness of the gate dielectric to be thicker without losing sensitivity to charge, which then reduces leakage currents and results in devices that are highly robust in fluid. This optimized process results in devices stable for up to 8 h in fluidic environments. Using pH sensing as a benchmark, we show the importance of optimizing the device bias, particularly the back gate bias which modulates the effective channel thickness. We also demonstrate that devices with Al(2)O(3) gate dielectrics exhibit superior sensitivity to pH when compared to devices with SiO(2) gate dielectrics. Finally, we show that when the effective electrical silicon channel thickness is on the order of the Debye length, device response to pH is virtually independent of device width. These silicon FET sensors could become integral components of future silicon based Lab on Chip systems.

Silicon field effect transistors as dual-use sensor-heater hybrids.

We demonstrate the temperature mediated applications of a previously proposed novel localized dielectric heating method on the surface of dual purpose silicon field effect transistor (FET) sensor-heaters and perform modeling and characterization of the underlying mechanisms. The FETs are first shown to operate as electrical sensors via sensitivity to changes in pH in ionic fluids. The same devices are then demonstrated as highly localized heaters via investigation of experimental heating profiles and comparison to simulation results. These results offer further insight into the heating mechanism and help determine the spatial resolution of the technique. Two important biosensor platform applications spanning different temperature ranges are then demonstrated: a localized heat-mediated DNA exchange reaction and a method for dense selective functionalization of probe molecules via the heat catalyzed complete desorption and reattachment of chemical functionalization to the transistor surfaces. Our results show that the use of silicon transistors can be extended beyond electrical switching and field-effect sensing to performing localized temperature controlled chemical reactions on the transistor itself.

Localized heating on silicon field effect transistors: device fabrication and temperature measurements in fluid.

We demonstrate electrically addressable localized heating in fluid at the dielectric surface of silicon-on-insulator field-effect transistors via radio-frequency Joule heating of mobile ions in the Debye layer. Measurement of fluid temperatures in close vicinity to surfaces poses a challenge due to the localized nature of the temperature profile. To address this, we developed a localized thermometry technique based on the fluorescence decay rate of covalently attached fluorophores to extract the temperature within 2 nm of any oxide surface. We demonstrate precise spatial control of voltage dependent temperature profiles on the transistor surfaces. Our results introduce a new dimension to present sensing systems by enabling dual purpose silicon transistor-heaters that serve both as field effect sensors as well as temperature controllers that could perform localized bio-chemical reactions in Lab on Chip applications.

A detection instrument for enhanced-fluorescence and label-free imaging on photonic crystal surfaces.

We report on the design and demonstration of an optical imaging system capable of exciting surface-bound fluorophores within the resonant evanescent electric field of a photonic crystal surface and gathering fluorescence emission that is directed toward the imaging objective by the photonic crystal. The system also has the ability to quantify shifts in the local resonance angle induced by the adsorption of biomolecules on the photonic crystal surface for label-free biomolecular imaging. With these two capabilities combined within a single detection system, we demonstrate label-free images self-registered to enhanced fluorescence images with 328x more sensitive fluorescence detection relative to a glass surface. This technique is applied to a DNA microarray where label-free quantification of immobilized capture DNA enables improved quality control and subsequent enhanced fluorescence detection of dye-tagged hybridized DNA yields 3x more genes to be detected versus commercially available microarray substrates.

Formation of tethered bilayer lipid membranes on gold surfaces: QCM-Z and AFM study.

Recently, tethered bilayer lipid membranes (tBLMs) have shown high potential as biomimetic systems due to their high stability and electrical properties, and have been used in applications ranging from membrane protein incorporation to biosensors. However, the kinetics of their formation remains largely uninvestigated. By using quartz crystal microbalance with impedance analysis (QCM-Z), we were able to monitor both the kinetics and viscoelastic properties of tether adsorption and vesicle fusion. Formation of the tether monolayer was shown to follow pseudo-first-order Langmuir kinetics with association and dissociation rate constants of 21.7 M-1 s(-1) and 7.43 x 10-6 s(-1), respectively. Moreover, the QCM-Z results indicate a rigid layer at the height of deposition, which then undergoes swelling as indicated by AFM. The deposition of vesicles to the tether layer also followed pseudo-first-order Langmuir kinetics with observed rate constants of 5.58 x 10(-2) and 2.41 x 10-2 s(-1) in water and buffer, respectively. Differential analysis of the QCM-Z data indicated deposition to be the fast kinetic step, with the rate-limiting steps being water release and fusion. Atomic force microscopy pictures taken complement the QCM-Z data, showing the major stages of tether adsorption and vesicle fusion, while providing a road map to successful tBLM formation.

AFM study of micelle chaining in surface films of polystyrene-block-poly(ethylene oxide) stars at the air/water interface.

A series of three-arm star block copolymers were examined using atomic force microscopy (AFM). These stars consisted of a polystyrene core composed of ca. 111 styrene units/branch with poly(ethylene oxide) (PEO) chains at the star periphery. Each star contained different amounts of PEO, varying from 107 to 415 ethylene oxide units/branch. The stars were spread as thin films at the air/water interface on a Langmuir trough and transferred onto mica at various surface pressures. Circular domains representing 2D micelle-like aggregated molecules were observed at low pressures. Upon further compression, these domains underwent additional aggregation in a systematic manner, including micellar chaining. At this point, domain area and the number of molecules/domain increased with increasing pressure. In addition, it was found that longer PEO chains led to greater intermolecular separation and less aggregation. These AFM results correspond to attributes seen in the surface pressure-area isotherms of the stars. In addition, they demonstrate the viability of AFM as a quantitative characterization technique.