Showing papers by "George M. Whitesides published in 2008"

Simple telemedicine for developing regions: camera phones and paper-based microfluidic devices for real-time, off-site diagnosis.

Abstract: This article describes a prototype system for quantifying bioassays and for exchanging the results of the assays digitally with physicians located off-site. The system uses paper-based microfluidic devices for running multiple assays simultaneously, camera phones or portable scanners for digitizing the intensity of color associated with each colorimetric assay, and established communications infrastructure for transferring the digital information from the assay site to an off-site laboratory for analysis by a trained medical professional; the diagnosis then can be returned directly to the healthcare provider in the field. The microfluidic devices were fabricated in paper using photolithography and were functionalized with reagents for colorimetric assays. The results of the assays were quantified by comparing the intensities of the color developed in each assay with those of calibration curves. An example of this system quantified clinically relevant concentrations of glucose and protein in artificial uri...

Three-dimensional microfluidic devices fabricated in layered paper and tape.

Abstract: This article describes a method for fabricating 3D microfluidic devices by stacking layers of patterned paper and double-sided adhesive tape. Paper-based 3D microfluidic devices have capabilities in microfluidics that are difficult to achieve using conventional open-channel microsystems made from glass or polymers. In particular, 3D paper-based devices wick fluids and distribute microliter volumes of samples from single inlet points into arrays of detection zones (with numbers up to thousands). This capability makes it possible to carry out a range of new analytical protocols simply and inexpensively (all on a piece of paper) without external pumps. We demonstrate a prototype 3D device that tests 4 different samples for up to 4 different analytes and displays the results of the assays in a side-by-side configuration for easy comparison. Three-dimensional paper-based microfluidic devices are especially appropriate for use in distributed healthcare in the developing world and in environmental monitoring and water analysis.

Eutectic Gallium-Indium (EGaIn) : A Liquid Metal Alloy for the Formation of Stable Structures in Microchannels at Room Temperature

Abstract: This paper describes the rheological behavior of the liquid metal eutectic gallium-indium (EGaIn) as it is injected into microfluidic channels to form stable microstructures of liquid metal. EGaIn is well-suited for this application because of its rheological properties at room temperature: it behaves like an elastic material until it experiences a critical surface stress, at which point it yields and flows readily. These properties allow EGaIn to fill microchannels rapidly when sufficient pressure is applied to the inlet of the channels, yet maintain structural stability within the channels once ambient pressure is restored. Experiments conducted in microfluidic channels, and in a parallel-plate rheometer, suggest that EGaIn’s behavior is dictated by the properties of its surface (predominantly gallium oxide, as determined by Auger measurements); these two experiments both yield approximately the same number for the critical surface stress required to induce EGaIn to flow (~0.5 N/m). This analysis–which shows that the pressure that must be exceeded for EGaIn to flow through a microchannel is inversely proportional to the critical (i.e., smallest) dimension of the channel–is useful to guide future fabrication of microfluidic channels to mold EGaIn into functional microstructures.

Electrostatic Charging Due to Separation of Ions at Interfaces: Contact Electrification of Ionic Electrets

Abstract: This Review discusses ionic electrets: their preparation, their mechanisms of formation, tools for their characterization, and their applications. An electret is a material that has a permanent, macroscopic electric field at its surface; this field can arise from a net orientation of polar groups in the material, or from a net, macroscopic electrostatic charge on the material. An ionic electret is a material that has a net electrostatic charge due to a difference in the number of cationic and anionic charges in the material. Any material that has ions at its surface, or accessible in its interior, has the potential to become an ionic electret. When such a material is brought into contact with some other material, ions can transfer between them. If the anions and cations have different propensities to transfer, the unequal transfer of these ions can result in a net transfer of charge between the two materials. This Review focuses on the experimental evidence and theoretical models for the formation of ionic electrets through this ion-transfer mechanism, and proposes--as a still-unproved hypothesis--that this ion-transfer mechanism may also explain the ubiquitous contact electrification ("static electricity") of materials, such as organic polymers, that do not explicitly have ions at their surface.

FLASH: A rapid method for prototyping paper-based microfluidic devices

Abstract: This article describes FLASH (Fast Lithographic Activation of Sheets), a rapid method for laboratory prototyping of microfluidic devices in paper. Paper-based microfluidic devices are emerging as a new technology for applications in diagnostics for the developing world, where low cost and simplicity are essential. FLASH is based on photolithography, but requires only a UV lamp and a hotplate; no clean-room or special facilities are required (FLASH patterning can even be performed in sunlight if a UV lamp and hotplate are unavailable). The method provides channels in paper with dimensions as small as 200 µm in width and 70 µm in height; the height is defined by the thickness of the paper. Photomasks for patterning paper-based microfluidic devices can be printed using an ink-jet printer or photocopier, or drawn by hand using a waterproof black pen. FLASH provides a straightforward method for prototyping paper-based microfluidic devices in regions where the technological support for conventional photolithography is not available.

Carbonic Anhydrase as a Model for Biophysical and Physical-Organic Studies of Proteins and Protein–Ligand Binding

Abstract: 1. Introduction: Overview of CA as a Model Carbonic anhydrase (CA, EC 4.2.1.1) is a protein that is especially well-suited to serve as a model in many types of studies in biophysics, bioanalysis, the physical-organic chemistry of inhibitor design, and medicinal chemistry. In vivo, this enzyme catalyzes the hydration of CO2 and the dehydration of bicarbonate (eq 1). CO2+H2O⇌HCO3−+H+ (1) The active site of α-CAs comprises a catalytic ZnII ion coordinated by three imidazole groups of histidines and by one hydroxide ion (or water molecule), all in a distorted tetrahedral geometry. This grouping is located at the base of a cone-shaped amphiphilic depression, one wall of which is dominated by hydrophobic residues and the other of which is dominated by hydrophilic residues.1 Unless otherwise stated, “CA” in this review refers to (i) various isozymes of α-CAs or (ii) the specific α-CAs human carbonic anhydrases I and II (HCA I and HCA II) and bovine carbonic anhydrase II (BCA II); “HCA” refers to HCA I and HCA II; and “CA II” refers to HCA II and BCA II. CA is particularly attractive for biophysical studies of protein–ligand binding for many reasons. (i) CA is a monomeric, single-chain protein of intermediate molecular weight (~30 kDa), and it has no pendant sugar or phosphate groups and no disulfide bonds. (ii) It is inexpensive and widely available. (iii) It is relatively easy to handle and purify, due in large part to its excellent stability under standard laboratory conditions. (iv) Amino acid sequences are available for most of its known isozymes. (v) The structure of CA, and of its active site, has been defined in detail by X-ray diffraction, and the mechanism of its catalytic activity is well-understood. (vi) As an enzyme, CA behaves not only as a hydratase/anhydrase with a high turnover number but also as an esterase (a reaction that is easy to follow experimentally). (vii) The mechanism of inhibition of CA by ligands that bind to the ZnII ion is fairly simple and well-characterized; it is, therefore, easy to screen inhibitors and to examine designed inhibitors that test theories of protein–ligand interactions. (viii) It is possible to prepare and study the metal-free apoenzyme and the numerous variants of CA in which the ZnII ion is replaced by other divalent ions. (ix) Charge ladders of CA II—sets of derivatives in which acylation of lysine amino groups (−NH3+ → −NHAc) changes the net charge of the protein—allow the influence of charge on properties to be examined by capillary electrophoresis. Some disadvantages of using CA include the following: (i) the presence of the ZnII cofactor, which can complicate biophysical and physical-organic analyses; (ii) a structure that is more stable than a representative globular protein and, thus, slightly suspect as a model system for certain studies of stability; (iii) a function—interconversion of carbon dioxide and carbonate—that does not involve the types of enzyme/substrate interactions that are most interesting in design of drugs; (iv) a catalytic reaction that is, in a sense, too simple (determining the mechanism of a reaction is, in practice, usually made easier if the reactants and products have an intermediate level of complexity); and (v) the absence of a solution structure of CA (by NMR spectroscopy). The ample X-ray data, however, paint an excellent picture of the changes (which are generally small) in the structure of CA that occur on binding ligands or introducing mutations. The most important class of inhibitors of CA, the aryl-sulfonamides, has several characteristics that also make it particularly suitable for physical-organic studies of inhibitor binding and in drug design: (i) arylsulfonamides are easily synthesized; (ii) they bind with high affinity to CA (1 μM to sub-nM); (iii) they share one common structural feature; and (iv) they share a common, narrowly defined geometry of binding that exposes a part of the ligand that can be easily modified synthetically. There are also many non-sulfonamide, organic inhibitors of CA, as well as anionic, inorganic inhibitors. We divide this review into five parts, all with the goal of using CA as a model system for biophysical studies: (I) an overview of the enzymatic activity and medical relevance of CA; (II) the structure and structure–function relationships of CA and its engineered mutants; (III) the thermodynamics and kinetics of the binding of ligands to CA; (IV) the effect of electrostatics on the binding of ligands to and the denaturation of CA; and (V) what makes CA a good model for studying protein–ligand binding and protein stability. 1.1. Value of Models CA serves as a good model system for the study of enzymes. That is, it is a protein having some characteristics representative of enzymes as a class, but with other characteristics that make it especially easy to study. It is a moderately important target in current medicinal chemistry: its inhibition is important in the treatment of glaucoma, altitude sickness, and obesity; its overexpression has recently been implicated in tumor growth; and its inhibition in pathogenic organisms might lead to further interesting drugs.2,3 More than its medical relevance, its tractability and simplicity are what make CA a particularly attractive model enzyme. The importance of models in science is often underestimated. Models represent more complex classes of related systems and contribute to the study of those classes by focusing research on particular, tractable problems. The development of useful, widely accepted models is a critical function of scientific research: many of the techniques (both experimental and analytical) and concepts of science are developed in terms of models; they are thoroughly engrained in our system of research and analysis. Examples of models abound in successful areas of science: in biology, E. coli, S. cerevisiae, Drosophila mela-nogaster, C. elegans, Brachydanio rerio (zebrafish), and the mouse; in chemistry, the hydrogen atom, octanol as a hydrophobic medium, benzene as an aromatic molecule, the 2-norbornyl carbocation as a nonclassical ion, substituted cyclohexanes for the study of steric effects, p-substituted benzoic acids for the study of electronic effects, cyclodextrins for ligand–receptor interactions; in physics, a vibrating string as an oscillator and a particle in a box as a model for electrons in orbitals. Science needs models for many reasons: Focus: Models allow a community of researchers to study a common subject. Solving any significant problem in science requires a substantial effort, with contributions from many individuals and techniques. Models are often the systems chosen to make this productive, cooperative focus possible. Research Overhead: Development of a system to the point where many details are scientifically tractable is the product of a range of contributions: for enzymes, these contributions are protocols for preparations, development of assays, determination of structures, preparation of mutants, definition of substrate specificity, study of rates, and development of mechanistic models. In a well-developed model system, the accumulation of this information makes it relatively easy to carry out research, since before new experiments begin, much of the background work–the fundamental research in a new system–has already been carried out. Recruiting and Interdisciplinarity: The availability of good model systems makes it relatively easy for a neophyte to enter an area of research and to test ideas efficiently. This ease of entry recruits new research groups, who use, augment, and improve the model system. It is especially important to have model systems to encourage participation by researchers in other disciplines, for whom even the elementary technical procedures in a new field may appear daunting. Comparability: A well-established model allows researchers in different laboratories to calibrate their experiments, by reproducing well-characterized experiments. Community: The most important end result of a good model system is often the generation of a scientific community–that is, a group of researchers examining a common problem from different perspectives and pooling information relevant to common objectives. One of the goals of this review is to summarize many experimental and theoretical studies of CA that have established it as a model protein. We hope that this summary will make it easier for others to use this protein to study fundamentals of two of the most important questions in current chemistry: (i) Why do a protein and ligand associate selectively? (ii) How can one design an inhibitor to bind to a protein selectively and tightly? We believe that the summary of studies of folding and stability of CA will be useful to biophysicists who study protein folding. In addition, we hope that the compilation of data relevant to CA in one review will ease the search for information for those who are beginning to work with this protein.

Eutectic gallium-indium (EGaIn): a moldable liquid metal for electrical characterization of self-assembled monolayers.

Abstract: Herein we describe the formation of conformal electrodes from the fluid metal eutectic, Ga–In (which we abbreviate “EGaIn” and pronounce “e-gain”; 75% Ga, 25% In by weight, m.p.= 15.5 8C), and their use in studying charge transport across self-assembled monolayers (SAMs). Although EGaIn is a liquid at room temperature, it does not spontaneously reflow into the shape with the lowest interfacial free energy as do liquids such as Hg and H2O: as a result, it can be formed into metastable, nonspherical structures (e.g., cones, and filaments with diameters 1 mm). This behavior, along with its high electrical conductivity (3.4 4 10 Scm ) and its tendency to make low contact-resistance interfaces with a variety of materials, makes EGaIn useful for forming electrodes for thin-film devices. We discuss the convenience and precision of measurements of current density (J, Acm ) versus applied voltage (V, V) through SAMs of n-alkanethiolates on template-stripped, ultraflat Ag (Ag–SCn Ag–SCnH2n+1, n= 10, 12, 14, 16) using EGaIn. An ideal electrode for physical-organic studies of SAMs would 1) make conformal, but nondamaging, physical contacts, 2) readily form small-area (micrometer diameter) contacts, to minimize the contribution of defects in the SAM to J, 3) form without specialized equipment, and 4) be nontoxic. Point 3 is particularly important: elimination of procedures such as evaporating metals or lithographic patterning would allow a wide range of laboratories—including those without access to clean rooms—to survey relationships between structure and electrical conductivity. There are currently three general techniques for forming top contacts for large-area (i.e., more than a few molecules) electrical measurements on SAMs of organic molecules: 1) The direct deposition of metals such as Au or Ti by using electron-beam or thermal evaporation ensures atomic-level contact, but results in low yields of devices owing to damage to the organic monolayer by reaction with hot metal vapors, and in the formation of metal filaments that short the junctions. 2) The installation of an electrically conducting polymer between the SAM and a metallic top contact inhibits formation of metal filaments, but the instability of SAMs of alkanethiolates to the temperatures required to anneal most electroactive polymers limits the broad application of this approach. 3) The use of Hg allows formation of conformal contacts at room temperature, but Hg is toxic, amalgamates with metals, tends to form junctions that short, is difficult to form into small contacts, and measurements with Hg must be performed under a solvent bath. EGaIn does not flow until it experiences a critical surface stress (0.5 Nm ), at which point it yields (i.e., flows). EGaIn 1) makes conformal, nondamaging contacts at room temperature, 2) can be molded into nonspherical shapes with micrometer-scale (or larger) dimensions, 3) is commercially available, 4) can be deposited with a pipette or syringe without high temperatures or vacuum, 5) has a low vapor pressure, and 6) is nontoxic. The work function of EGaIn (4.1–4.2 eV) is close to that of Hg (4.5 eV), but EGaIn does not alloy with many metals. It is therefore an ideal replacement for Hg, especially in devices that incorporate SAMs (which are generally formed on Au or Ag). Auger spectroscopy on samples of EGaIn in air show that its surface is principally composed of oxides of Ga (see the Supporting Information); gallium oxide is an n-type semiconductor. There is undoubtedly an adsorbed film of water on this surface, as EGaIn has a high surface free energy (ca. 630 dynescm ), as do oxides formed from similar metals. During our measurements, there were no observable changes in the average magnitude or range of J when EGaIn was allowed to sit in air for extended periods before we deposited it on the SAM, or when we performed the measurements using the same drop of EGaIn to form between three and five junctions, or while we flowed dry N2 over the sample: therefore the contribution of the surface oxide to J was probably constant for the duration of the experiments. We formed EGaIn electrodes by suspending a drop of EGaIn from a metal 26s-gauge needle affixed to a 10-mL syringe, bringing the drop into contact with the bare surface of a sacrificial film of Ag using a micromanipulator, and retracting the needle slowly (ca. 50 mms ); the EGaIn adhered to both the needle and the Ag (Figure 1). The drop of EGaIn pinched into to an hour-glass shape until it bifurcated into two structures, one attached to the syringe (a cone approximately 0.05 mL in volume) and one (which was discarded) attached to the Ag. We produced conical tips of EGaIn with diameters ranging from less than 1 mm to 100 mm; the larger the bore of the needle, and the more rapidly we [*] Dr. R. C. Chiechi, Dr. E. A. Weiss, Dr. M. D. Dickey, Prof. G. M. Whitesides Department of Chemistry and Chemical Biology Harvard University 12 Oxford St., Cambridge, MA 02138 (USA) Fax: (+1)617-495-9857 E-mail: gwhitesides@gmwgroup.harvard.edu

Low-cost printing of poly(dimethylsiloxane) barriers to define microchannels in paper.

Abstract: This paper describes the use of a modified x,y-plotter to generate hydrophilic channels by printing a solution of hydrophobic polymer (pol(dimethylsiloxane; PDMS) dissolved in hexanes onto filter paper. The PDMS penetrates the depth of the paper and forms a hydrophobic wall that aqueous solutions cannot cross. The minimum size of printed features is ∼1 mm; this resolution is adequate for the rapid prototyping of hand-held, visually read, diagnostic assays (and other microfluidic systems) based on paper. After curing the printed PDMS, the paper-based devices can be bent or folded to generate three-dimensional systems of channels. Capillary action pulls aqueous samples into the paper channels. Colorimetric assays for the presence of glucose and protein are demonstrated in the printed devices; spots of Bromothymol Blue distinguished samples with slightly basic pH (8.0) from samples with slightly acidic pH (6.5). The work also describes using printed devices that can be loaded using multipipets and printed fl...

Emulsification in a microfluidic flow-focusing device: effect of the viscosities of the liquids

Abstract: We report the results of a comparative study of microfluidic emulsification of liquids with different viscosities. Depending on the properties of the fluids and their rates of flow, emulsification occurred in the dripping and jetting regimes. We studied the characteristic features and typical dependence of the size and of the size distribution of droplets in each regime. For each liquid, we identified a range of hydrodynamic conditions promoting generation of highly monodisperse droplets. Viscosity played an important role in emulsification: highly viscous liquids were emulsified into larger droplets with lower polydispersity. Although it was not possible to provide a unified scaling for the volumes of the droplets, our results suggest that the break-up dynamics of the lower viscosity fluids resembles the rate-of-flow-controlled break-up, as reported earlier for the formation of bubbles in flow-focusing geometries [Garstecki P, Stone HA, Whitesides GM (2005) Phys Rev Lett 94:164501]. The results of this study can be helpful for a rationalized selection of liquids for the controlled formation of droplets with a predetermined size and with a narrow distribution of sizes.

Methods of etching articles via microcontact printing

Abstract: Improved methods of forming a patterned self-assembled monolayer on a surface and derivative articles are provided. According to one method, an elastomeric stamp is deformed during and/or prior to using the stamp to print a self-assembled molecular monolayer on a surface. According to another method, during monolayer printing the surface is contacted with a liquid that is immiscible with the molecular monolayer-forming species to effect controlled reactive spreading of the monolayer on the surface. Methods of printing self-assembled molecular monolayers on nonplanar surfaces and derivative articles are provided, as are methods of etching surfaces patterned with self-assembled monolayers, including methods of etching silicon. Optical elements including flexible diffraction gratings, mirrors, and lenses are provided, as are methods for forming optical devices and other articles using lithographic molding. A method for controlling the shape of a liquid on the surface of an article is provided, involving applying the liquid to a self-assembled monolayer on the surface, and controlling the electrical potential of the surface.

Dynamically reconfigurable liquid-core liquid-cladding lens in a microfluidic channel

Abstract: This paper describes the design and operation of a liquid-core liquid-cladding (L2) lens formed by the laminar flow of three streams of liquids in a microchannel whose width expands laterally in the region where the lens forms. Two streams of liquid with a lower refractive index (the cladding) sandwich a stream of liquid with a higher refractive index (the core). As the core stream enters the expansion chamber, it widens and becomes biconvex in shape, for some rates of flow. This biconvex fluidic element focuses light. Manipulating the relative rates of flow of the streams reconfigures the shape, and therefore the focal distance, of the L2 lens. This paper also describes a technique for beam tracing, and for characterization of a lens in an enclosed micro-scale optical system. The use of a cladding liquid with refractive index matched to that of the material used in the fabrication of the microfluidic system (here, poly(dimethylsiloxane)) improves the quality of the focused beam.

Directional control of cell motility through focal adhesion positioning and spatial control of Rac activation

Abstract: Local physical interactions between cells and extracellular matrix (ECM) influence directional cell motility that is critical for tissue development, wound repair, and cancer metastasis. Here we test the possibility that the precise spatial positioning of focal adhesions governs the direction in which cells spread and move. NIH 3T3 cells were cultured on circular or linear ECM islands, which were created using a microcontact printing technique and were 1 μm wide and of various lengths (1 to 8 μm) and separated by 1 to 4.5 μm wide nonadhesive barrier regions. Cells could be driven proactively to spread and move in particular directions by altering either the interisland spacing or the shape of similar-sized ECM islands. Immunofluorescence microscopy confirmed that focal adhesions assembled preferentially above the ECM islands, with the greatest staining intensity being observed at adhesion sites along the cell periphery. Rac-FRET analysis of living cells revealed that Rac became activated within 2 min afte...

Nanoskiving: A New Method To Produce Arrays of Nanostructures

Abstract: This Account reviews nanoskiving--a new technique that combines thin-film deposition of metal on a topographically contoured substrate with sectioning using an ultramicrotome--as a method of fabricating nanostructures that could replace conventional top-down techniques in selected applications. Photolithography and scanning beam lithography, conventional top-down techniques to generate nanoscale structures and nanostructured materials, are useful, versatile, and highly developed, but they also have limitations: high capital and operating costs, limited availability of the facilities required to use them, an inability to fabricate structures on nonplanar surfaces, and restrictions on certain classes of materials. Nanoscience and nanotechnology would benefit from new, low-cost techniques to fabricate electrically and optically functional structures with dimensions of tens of nanometers, even if (or perhaps especially if) these techniques have a different range of application than does photolithography or scanning beam lithography. Nanoskiving provides a simple and convenient procedure to produce arrays of structures with cross-sectional dimensions in the 30-nm regime. The dimensions of the structures are determined by (i) the thickness of the deposited thin film (tens of nanometers), (ii) the topography (submicrometer, using soft lithography) of the surface onto which the thin film is deposited, and (iii) the thickness of the section cut by the microtome (> or =30 nm by ultramicrotomy). The ability to control the dimensions of nanostructures, combined with the ability to manipulate and position them, enables the fabrication of nanostructures with geometries that are difficult to prepare by other methods. The nanostructures produced by nanoskiving are embedded in a thin epoxy matrix. These epoxy slabs, although fragile, have sufficient mechanical strength to be manipulated and positioned; this mechanical integrity allows the nanostructures to be stacked in layers, draped over curved surfaces, and suspended across gaps, while retaining the in-plane geometry of the nanostructures embedded in the epoxy. After removal of the polymer matrix by plasma oxidation, these structures generate suspended and draped nanostructures and nanostructures on curved surfaces. Two classes of applications, in optics and in electronics, demonstrate the utility of nanostructures fabricated by nanoskiving. This technique will be of primary interest to researchers who wish to generate simple nanostructures, singly or in arrays, more simply and quickly than can be accomplished in the clean-room. It is easily accessible to those not trained in top-down procedures for fabrication and those with limited or no access to the equipment and facilities needed for photolithography or scanning-beam fabrication. This Account discusses a new fabrication method (nanoskiving) that produces arrays of metal nanostructures. The defining process in nanoskiving is cutting slabs from a polymeric matrix containing embedded, more extended metal structures.

Formation of Bubbles and Droplets in Parallel, Coupled Flow‐Focusing Geometries

Abstract: This paper describes the mechanism of formation of bubbles of nitrogen in water containing Tween 20 as a surfactant, and of droplets of water in hexadecane containing Span 80 as a surfactant. The study of these microfluidic systems compares two or four flow-focusing generators coupled through shared inlets, supplying the continuous phase, and through a common outlet channel. The processes that form bubbles in neighboring generators interact for a wide range of flow parameters; the formation of bubbles alternates in time and space, and the bubbles assemble into complex patterns in the outlet channel. The dynamics of formation of bubbles in these systems are stable for long time (at least 10 min). For a certain range of flow parameters, the coupled flow-focusing generators exhibit two stable modes of operation for a single set of flow parameters. The dynamics of formation of droplets of water in hexadecane by the coupled flow-focusing generators are simpler--the adjacent generators produce only monodisperse droplets over the entire range of flow parameters that are explored. These observations suggest that the mechanism of interaction between coupled flow-focusing generators relies on the compressibility of the dispersed phase (e.g., the gas or liquid), and on variations in pressure at the flow-focusing orifices induced by the breakup of bubbles or droplets.

Egg beater as centrifuge: isolating human blood plasma from whole blood in resource-poor settings

Abstract: This paper demonstrates that a hand-powered egg beater can be modified to serve as a centrifuge for separating plasma from human whole blood. Immunoassays used to diagnose infectious diseases often require plasma from whole blood, and obtaining plasma typically requires electrically-powered centrifuges, which are not widely available in resource-limited settings. Human whole blood was loaded into polyethylene (PE) tubing, and the tubing was attached to the paddle of an egg beater. Spinning the paddle pelleted the blood cells to the distal end of the PE tubing; the plasma remained as the supernatant. A cholesterol assay (run on patterned paper) demonstrated the suitability of this plasma for use in diagnostic assays. The physics of the system was also analyzed as a guide for the selection of other rotating systems for use in centrifugation. Egg beaters, polyethylene tubing, and paper are readily available devices and supplies that can facilitate the use of point-of-care diagnostics at sites far from centralized laboratory facilities.

Fabrication of a modular tissue construct in a microfluidic chip

Abstract: By combining microfluidics and soft-lithographic molding of gels containing mammalian cells, a device for three-dimensional (3D) culture of mammalian cells in microchannels was developed. Native components of the extracellular matrix, including collagen or Matrigel™, made up the matrix of each molded piece (module) of cell-containing gel. Each module had at least one dimension below ∼300 μm; in modules of these sizes, the flux of oxygen, nutrients, and metabolic products into and out of the modules was sufficient to allow cells in the modules to proliferate to densities comparable to those of native tissue (108–109cells cm−3). Packing modules loosely into microfluidic channels and chambers yielded structures permeated with a network of pores through which cell culture medium could flow to feed the encapsulated cells. The order in the packed assemblies increased as the width of the microchannels approached the width of the modules. Multiple cell types could be spatially organized in the small microfluidic channels. Recovery and analysis of modules after 24 h under constant flow of medium (200 μL h−1) showed that over 99% of encapsulated cells survived this interval in the microfluidic chamber.

Using ratchets and sorters to fractionate motile cells of Escherichia coli by length

Abstract: This paper describes the fabrication of a composite agar/PDMS device for enriching short cells in a population of motile Escherichia coli. The device incorporated ratcheting microchannels, which directed the motion of swimming cells of E. coli through the device, and three sorting junctions, which isolated successively shorter populations of bacteria. The ratcheting microchannels guided cells through the device with an average rate of displacement of (32 ± 9) μm s−1. Within the device, the average length of the cells decreased from 3.8 μm (Coefficient of Variation, CV: 21%) at the entrance, to 3.4 μm (CV: 16%) after the first sorting junction, to 3.2 μm (CV: 19%) after the second sorting junction, to 3.0 μm (CV: 19%) after the third sorting junction.

Cell Encapsulation in Sub-mm Sized Gel Modules Using Replica Molding

Abstract: For many types of cells, behavior in two-dimensional (2D) culture differs from that in three-dimensional (3D) culture. Among biologists, 2D culture on treated plastic surfaces is currently the most popular method for cell culture. In 3D, no analogous standard method—one that is similarly convenient, flexible, and reproducible—exists. This paper describes a softlithographic method to encapsulate cells in 3D gel objects (modules) in a variety of simple shapes (cylinders, crosses, rectangular prisms) with lateral dimensions between 40 and 1000 mm, cell densities of 10 5 –1 0 8 cells/cm 3 , and total volumes between 1610 27 and 8610 24 cm 3 . By varying (i) the initial density of cells at seeding, and (ii) the dimensions of the modules, the number of cells per module ranged from 1 to 2500 cells. Modules were formed from a range of standard biopolymers, including collagen, Matrigel TM , and agarose, without the complex equipment often used in encapsulation. The small dimensions of the modules allowed rapid transport of nutrients by diffusion to cells at any location in the module, and therefore allowed generation of modules with cell densities near to those of dense tissues (10 8 –1 0 9 cells/cm 3 ). This modular method is based on soft lithography and requires little special equipment; the method is therefore accessible, flexible, and well suited to (i) understanding the behavior of cells in 3D environments at high densities of cells, as in dense tissues, and (ii) developing applications in tissue engineering.

Fabrication of Arrays of Metal and Metal Oxide Nanotubes by Shadow Evaporation

Abstract: This paper describes a simple technique for fabricating uniform arrays of metal and metal oxide nanotubes with controlled heights and diameters. The technique involves depositing material onto an a...

Fabrication of Surface Plasmon Resonators by Nanoskiving Single-Crystalline Gold Microplates

Abstract: This paper demonstrates the sectioning of chemically synthesized, single-crystalline microplates of gold with an ultramicrotome (nanoskiving) to produce single-crystalline nanowires; these nanowires act as low-loss surface plasmon resonators. This method produces collinearly aligned nanostructures with small, regular changes in dimension with each consecutive cross-section: a single microplate thus can produce a number of “quasi-copies” (delicately modulated variations) of a nanowire. The diamond knife cuts cleanly through microplates 35 μm in diameter and 100 nm thick without bending the resulting nanowire and cuts through the sharp edges of a crystal without deformation to generate nanoscale tips. This paper compares the influence of sharp tips and blunt tips on the resonator modes in these nanowires.

Using magnetic levitation to distinguish atomic-level differences in chemical composition of polymers, and to monitor chemical reactions on solid supports.

Abstract: This communication describes a density-based method that uses magnetic levitation for monitoring solid-supported reactions and for distinguishing differences in chemical composition of polymers. The method is simple, rapid, and inexpensive and is similar to thin-layer chromatography (TLC; for solution-phase chemistry) in its potential for monitoring reactions in solid-phase chemistry. The technique involves levitating a sample of beads (taken from a reaction mixture) in a cuvette containing a paramagnetic solution (e.g., GdCl3 dissolved in H2O) positioned between two NdFeB magnets. The vertical position at which the beads levitate corresponds to the density of the beads and correlates with the progress of a chemical reaction on a solid support. The method is particularly useful for monitoring the kinetics of reactions occurring on polymer beads.

Fabrication of Arrays of Metal and Metal Oxide Nanotubes by Shadow

Abstract: This paper describes a simple technique for fabricating uniform arrays of metal and metal oxide nanotubes with controlled heights and diameters. The technique involves depositing material onto an anodized aluminumoxide(AAO)membranetemplateusingacollimatedelectronbeamevaporationsource.Theevaporating material enters the porous openings of the AAO membrane and deposits onto the walls of the pores. The membrane is tilted with respect to the column of evaporating material, so the shadows cast by the openings of the pores onto the inside walls of the pores define the geometry of the tubes. Rotation of the membrane during evaporation ensures uniform deposition inside the pores. After evaporation, dissolution of the AAO in base easily removesthetemplatetoyieldanarrayofnanotubesconnectedbyathinbackingofthesamemetalormetaloxide. The diameter of the pores dictates the diameter of the tubes, and the incident angle of evaporation determines theheightofthetubes.Tubesupto1.5minheightand20-200nmindiameterwerefabricated.Thismethod is adaptable to any material that can be vapor-deposited, including indiumtin oxide (ITO), a conductive, transparent material that is useful for many opto-electronic applications. An array of gold nanotubes produced bythistechniqueservedasasubstrateforsurface-enhancedRamanspectroscopy:theRamansignal(permolecule) from a monolayer of benzenethiolate was a factor of510 5 greater than that obtained using bulk liquid benzenethiol.

Lysine acetylation can generate highly charged enzymes with increased resistance toward irreversible inactivation

Abstract: This paper reports that the acetylation of lysine e-NH3 + groups of α-amylase—one of the most important hydrolytic enzymes used in industry—produces highly negatively charged variants that are enzymatically active, thermostable, and more resistant than the wild-type enzyme to irreversible inactivation on exposure to denaturing conditions (e.g., 1 h at 90°C in solutions containing 100-mM sodium dodecyl sulfate). Acetylation also protected the enzyme against irreversible inactivation by the neutral surfactant TRITON X-100 (polyethylene glycol p-(1,1,3,3-tetramethylbutyl)phenyl ether), but not by the cationic surfactant, dodecyltrimethylammonium bromide (DTAB). The increased resistance of acetylated α-amylase toward inactivation is attributed to the increased net negative charge of α-amylase that resulted from the acetylation of lysine ammonium groups (lysine e-NH3 + → e-NHCOCH3). Increases in the net negative charge of proteins can decrease the rate of unfolding by anionic surfactants, and can also decrease the rate of protein aggregation. The acetylation of lysine represents a simple, inexpensive method for stabilizing bacterial α-amylase against irreversible inactivation in the presence of the anionic and neutral surfactants that are commonly used in industrial applications.

Size-dependent charge collection in junctions containing single-size and multi-size arrays of colloidal CdSe quantum dots.

Abstract: This paper describes the electrical characteristics of junctions composed of three-dimensional arrays of colloidal CdSe quantum dots (QDs) with tin-doped indium oxide (ITO)/poly(3,4-ethylenedioxy-thiophene):poly(styrenesulfonate) (PEDOT:PSS) and eutectic Ga−In (EGaIn) electrodes. It focuses on a comparison of junctions containing QDs of one size to those of arrays containing QDs of multiple sizes. This comparison makes it possible to estimate the relative contributions of transport across various interfaces (e.g., between the QDs and between the QDs and the electrodes) to the observed electrical characteristics of the junction and to evaluate the dependence of these contributions on the locations of various sizes of QDs within the junction. The junctions were diodes, and their turn-on voltage depended on the size of the QDs next to the PEDOT:PSS. We describe this dependence using a Marcus model to estimate the barrier for charge transfer induced by the difference in energies between the orbitals of the QD...

Fluid Flow Induced Calcium Response in Bone Cell Network.

Abstract: In our previous work, bone cell networks with controlled spacing and functional intercellular gap junctions had been successfully established by using microcontact printing and self assembled monolayers technologies [Guo, X. E., E. Takai, X. Jiang, Q. Xu, G. M. Whitesides, J. T. Yardley, C. T. Hung, E. M. Chow, T. Hantschel, and K. D. Costa. Mol. Cell. Biomech. 3:95–107, 2006]. The present study investigated the calcium response and the underlying signaling pathways in patterned bone cell networks exposed to a steady fluid flow. The glass slides with cell networks were separated into eight groups for treatment with specific pharmacological agents that inhibit pathways significant in bone cell calcium signaling. The calcium transients of the network were recorded and quantitatively evaluated with a set of network parameters. The results showed that 18α-GA (gap junction blocker), suramin (ATP inhibitor), and thapsigargin (depleting intracellular calcium stores) significantly reduced the occurrence of multiple calcium peaks, which were visually obvious in the untreated group. The number of responsive peaks also decreased slightly yet significantly when either the COX-2/PGE2 or the NOS/nitric oxide pathway was disrupted. Different from all other groups, cells treated with 18α-GA maintained a high concentration of intracellular calcium following the first peak. In the absence of calcium in the culture medium, the intracellular calcium concentration decreased slowly with fluid flow without any calcium transients observed. These findings have identified important factors in the flow mediated calcium signaling of bone cells within a patterned network.

Interfacial instabilities in a microfluidic Hele-Shaw cell

Abstract: This paper describes surfactant-sensitive, dynamic instabilities that occur to aqueous droplets translating in a continuous flow of hexadecane in a microfluidic Hele-Shaw cell (HSC). A very low interfacial tension (on the order of 0.01 mN m−1) between water and hexadecane allowed for deformation of the droplets along the fields of flow and tip-streaming from moving droplets. In the system of water and hexadecane that we investigated, the use of surfactants in both fluids was necessary to achieve interfacial tension sufficiently low for the instabilities to occur. The droplets entering the HSC stretched orthogonally to the main direction of flow into elongated shapes, with aspect ratios greater than ten to one (width to length). These droplets exhibited two types of instabilities. The first included elongation of droplets, and Rayleigh–Plateau instabilities in the stretched droplets. Arrays of these stretched droplets formed three characteristic patterns that depended on the rates of flow of water and hexadecane. The second was driven by the shear stress exerted on the interface between the two fluids by the top and bottom boundaries of the HSC; this instability is named a “shear-driven instability” (SDI). Our observations supported that the SDI—an effect similar to tip-streaming—resulted from a redistribution of surfactants at the interface between the two fluids.

Patterns of Electrostatic Charge and Discharge in Contact Electrification

Abstract: Here we describe a study of the charging and discharging of solids in a system comprising a metal sphere that rolls across an electrically insulating plate. There are two kinetically distinct processes: 1) charging at a constant rate; 2) abrupt discharging, when the potential difference between sphere and surface reaches a critical value determined by the dielectric strength of air. This work has two objectives: 1) to develop a procedure for examining the rate of charging and discharging as a function of a range of relevant variables; 2) to use this information to test the hypotheses that charge separation involved ions and that discharge of the potential produced involved a breakdown of air. In published work, we have described this system; this study demonstrates the wealth of quantitative information it can provide as a tool for studying the atomic/molecular mechanisms of contact electrification. These mechanisms are relevant to processes ranging from lightning to xerography, and are a subject of active controversy. Three mechanisms appear to contribute to contact electrification: 1) ion transfer between surfaces having mobile ions, 2) partitioning of ions from adsorbed water onto the surfaces of non-ionic insulators, and 3) electron transfer between conductors and semiconductors (materials with mobile electrons and well-defined Fermi surfaces). We have concluded—in agreement with a hypothesis by Diaz —that the transfer of ions between the contacting surfaces is the most common mechanism for charge separation when organic materials are involved. The data we present here are consistent with contact charging by the slow transfer of ions, interrupted by episodic, rapid discharge events involving ionized plasmas when the difference in electrical potential between the surfaces exceeds the breakdown limit of air. These experiments used the rolling sphere tool (RST, Figure 1) developed by Grzybowski et al. We investigated contact electrification between stainless steel spheres (d= 3.2 mm) and three different surfaces (relative humidity, RH = 20–25 %, T 22 8C, w = 80 rpm). The Supporting Information contains the experimental procedures we followed for preparing the insulating surfaces: 1) glass (a 1.0 mm thick, 76 mm diameter wafer of low-alkali glass); 2) glass silanized with N-trimethoxysilylpropyl-N,N,N-trimethylammonium chloride; 3) glass silanized with 3-(trihydroxysilyl)-1-propane-sulfonic acid. When the sphere was far (more than ca. 2.5 cm) from the electrode (width 5 mm, 0.2 radians), the electrometer reported only the charge on the portion of the insulator (the glass plate) to which the electrode coupled (Qw). When the sphere passed over the electrode, the electrometer registered a peak in the charge, the height of which was the sum (Qs+w) of the charges that the electrode sensed on the sphere (Qs) and Qw. Figure 2a shows the charge the electrometer recorded for one revolution of the sphere. The fullwidth at half-maximum of the peak was about 0.63 radians. Figure 2b shows a representative plot of the complex pattern of charge (Q, in picocoulomb, pC: 1 pC = 6.2 ; 10 elementary charges) the electrometer reported as the sphere rolled on a glass wafer. When the sphere was directly over the electrode, the electrometer measured a fraction of the charge on the sphere (80–90 %; see the Supporting Information). Qw (grey dot-dash guidelines) and Qs+w (black dashed guidelines) increased linearly with time. Sharp discontinuities— discharge events through air—interrupted the charging. Subtracting Qw from Qs+w gave Qs—the charge that the electrometer sensed on the rolling sphere alone—as a function of time (Figure 2c). We have observed qualitatively similar behavior on a variety of materials, including organic polymers; we will detail these experiments in a full paper. The polarity of charge separation was invariant when a steel sphere (positive) rolled on a clean glass wafer (negative) (Figure 3a). When the sphere rolled on a surface with bound Figure 1. Illustration of the “rolling sphere tool” to measure the kinetics of contact electrification between rolling stainless steel spheres and insulating surfaces. The Supporting Information contains additional graphical representations.

Using ion channel-forming peptides to quantify protein-ligand interactions.

Abstract: This paper proposes a method for sensing affinity interactions by triggering disruption of self-assembly of ion channel-forming peptides in planar lipid bilayers. It shows that the binding of a derivative of alamethicin carrying a covalently attached sulfonamide ligand to carbonic anhydrase II (CA II) resulted in the inhibition of ion channel conductance through the bilayer. We propose that the binding of the bulky CA II protein (MW approximately 30 kD) to the ion channel-forming peptides (MW approximately 2.5 kD) either reduced the tendency of these peptides to self-assemble into a pore or extracted them from the bilayer altogether. In both outcomes, the interactions between the protein and the ligand lead to a disruption of self-assembled pores. Addition of a competitive inhibitor, 4-carboxybenzenesulfonamide, to the solution released CA II from the alamethicin-sulfonamide conjugate and restored the current flow across the bilayer by allowing reassembly of the ion channels in the bilayer. Time-averaged recordings of the current over discrete time intervals made it possible to quantify this monovalent ligand binding interaction. This method gave a dissociation constant of approximately 2 microM for the binding of CA II to alamethicin-sulfonamide in the bilayer recording chamber: this value is consistent with a value obtained independently with CA II and a related sulfonamide derivative by isothermal titration calorimetry.

Fabrication of Conjugated Polymer Nanowires by Edge Lithography

Abstract: This paper describes the fabrication of conjugated polymer nanowires by a three stage process: (i) spin-coating a composite film comprising alternating layers of a conjugated polymer and a sacrificial material, (ii) embedding the film in an epoxy matrix and sectioning it with an ultramicrotome (nanoskiving), and (iii) etching the sacrificial material to reveal nanowires of the conjugated polymer. A free-standing, 100-layer film of two conjugated polymers was spin-coated from orthogonal solvents: poly(2-methoxy-5-(2′-ethylhexyloxy)-1,4-phenylenevinylene) (MEH-PPV) from chloroform and poly(benzimidazobenzophenanthroline ladder) (BBL) from methanesulfonic acid. After sectioning the multilayer film, dissolution of the BBL with methanesulfonic acid yielded uniaxially aligned MEH-PPV nanowires with rectangular cross sections, and etching MEH-PPV with an oxygen plasma yielded BBL nanowires. The conductivity of MEH-PPV nanowires changed rapidly and reversibly by >103 upon exposure to I2 vapor. The result suggests...