Uniform Amplification of Phage with Different Growth Characteristics in Individual Compartments Consisting of Monodisperse Droplets

Jeder Klon zählt! Beim Phagen-Display gehen Klone, die das Phagenwachstum hemmende Liganden präsentieren, bei der Vervielfältigung verloren. In monodispersen Emulsionen, die mithilfe eines einfachen Mikrofluidiksystems erzeugt wurden, ist die Konkurrenz zwischen langsam (S) und schnell (R) wachsenden Phagen abgemildert, sodass das R/S-Verhältnis beibehalten bleibt. Die konkurrenzfreie Vervielfältigung von Phagen bewahrt Liganden, die im normalen Phagen-Display abhanden kommen.Chemistry and Chemical Biolog

Denaturation of Proteins by SDS and Tetraalkylammonium Dodecyl Sulfates

This article describes the use of capillary electrophoresis (CE) to examine the influence of different cations (C(+); C(+) = Na(+) and tetra-n-alkylammonium, NR(4)(+), where R = Me, Et, Pr, and Bu) on the rates of denaturation of bovine carbonic anhydrase II (BCA) in the presence of anionic surfactant dodecylsulfate (DS(-)). An analysis of the denaturation of BCA in solutions of Na(+)DS(-) and NR(4)(+)DS(-) (in Tris-Gly buffer) indicated that the rates of formation of complexes of denatured BCA with DS(-) (BCA(D)-DS(-)(n,sat)) are indistinguishable and independent of the cation below the critical micellar concentration (cmc) and independent of the total concentration of DS(-) above the cmc. At concentrations of C(+)DS(-) above the cmc, BCA denatured at rates that depended on the cation; the rates decreased by a factor >10(4) in the order of Na(+) ≈ NMe(4)(+) > NEt(4)(+) > NPr(4)(+) > NBu(4)(+), which is the same order as the values of the cmc (which decrease from 4.0 mM for Na(+)DS(-) to 0.9 mM for NBu(4)(+)DS(-) in Tris-Gly buffer). The relationship between the cmc values and the rates of formation of BCA(D)-DS(-)(n,sat()) suggested that the kinetics of denaturation of BCA involve the association of this protein with monomeric DS(-) rather than with micelles of (C(+)DS(-))(n). A less-detailed survey of seven other proteins (α-lactalbumin, β-lactoglobulin A, β-lactoglobulin B, carboxypeptidase B, creatine phosphokinase, myoglobin, and ubiquitin) showed that the difference between Na(+)DS(-) and NR(4)(+)DS(-) observed with BCA was not general. Instead, the influence of NR(4)(+) on the association of DS(-) with these proteins depended on the protein. The selection of the cation contributed to the properties (including the composition, electrophoretic mobility, and partitioning behavior in aqueous two-phase systems) of aggregates of denatured protein and DS(-). These results suggest that the variation in the behavior of NR(4)(+)DS(-) with changes in R may be exploited in methods used to analyze and separate mixtures of proteins.Chemistry and Chemical Biolog

A multi-color fast-switching microfluidic droplet dye laser

We describe a multi-color microfluidic dye laser operating in whispering gallery mode based on a train of alternating droplets containing solutions of different dyes; this laser is capable of switching the wavelength of its emission between 580 nm and 680 nm at frequencies up to 3.6 kHz -— the fastest among all dye lasers reported; it has potential applications in on-chip spectroscopy and flow cytometry

Continuously tunable microdroplet-laser in a microfluidic channel

This paper describes the generation and optical characterization of a series of dye-doped droplet-based optical microcavities with continuously decreasing radius in a microfluidic channel. A flow-focusing nozzle generated the droplets (~21 μm in radius) using benzyl alcohol as the disperse phase and water as the continuous phase. As these drops moved down the channel, they dissolved, and their size decreased. The emission characteristics from the drops could be matched to the whispering gallery modes from spherical micro-cavities. The wavelength of emission from the drops changed from 700 to 620 nm as the radius of the drops decreased from 21 μm to 7 μm. This range of tunability in wavelengths was larger than that reported in previous work on droplet-based cavities.Chemistry and Chemical Biolog

Externally Applied Electric Fields up to 1.6 × 10 5 V/m Do Not Affect the Homogeneous Nucleation of Ice in Supercooled Water

The freezing of water can initiate at electrically conducting electrodes kept at a high electric potential or at charged electrically insulating surfaces. The microscopic mechanisms of these phenomena are unknown, but they must involve interactions between water molecules and electric fields. This paper investigates the effect of uniform electric fields on the homogeneous nucleation of ice in supercooled water. Electric fields were applied across drops of water immersed in a perfluorinated liquid using a parallel-plate capacitor; the drops traveled in a microchannel and were supercooled until they froze due to the homogeneous nucleation of ice. The distribution of freezing temperatures of drops depended on the rate of nucleation of ice, and the sensitivity of measurements allowed detection of changes by a factor of 1.5 in the rate of nucleation. Sinusoidal alternation of the electric field at frequencies from 3 to 100 kHz prevented free ions present in water from screening the electric field in the bulk of drops. Uniform electric fields in water with amplitudes up to (1.6 ± 0.4) × 105 V/m neither enhanced nor suppressed the homogeneous nucleation of ice. Estimations based on thermodynamic models suggest that fields in the range of 107−108 V/m might cause an observable increase in the rate of nucleation.Chemistry and Chemical Biolog

Optofluidic ultrahigh-throughput detection of fluorescent drops

This paper describes an optofluidic droplet interrogation device capable of counting fluorescent drops at a throughput of 254000 drops per second. To our knowledge, this rate is the highest interrogation rate published thus far. Our device consists of 16 parallel microfluidic channels bonded directly to a filter-coated two-dimensional Complementary Metal-Oxide-Semiconductor (CMOS) sensor array. Fluorescence signals emitted from the drops are collected by the sensor that forms the bottom of the channel. The proximity of the drops to the sensor facilitates efficient collection of fluorescence emission from the drops, and overcomes the trade-off between light collection efficiency and field of view in conventional microscopy. The interrogation rate of our device is currently limited by the acquisition speed of CMOS sensor, and is expected to increase further as high-speed sensors become increasingly available

Filter-Based Assay for Escherichia coli in Aqueous Samples Using Bacteriophage-Based Amplification

This paper describes a method to detect the presence of bacteria in aqueous samples, based on the capture of bacteria on a syringe filter, and the infection of targeted bacterial species with a bacteriophage (phage). The use of phage as a reagent provides two opportunities for signal amplification: i) the replication of phage inside a live bacterial host (1000-fold amplification for M13 phage in E. coli K12), and ii) the rapid conversion of a colorless substrate to a colored or fluorescent product by an enzyme that is co-expressed with the phage (in this demonstration β- galactosidase, which has a turnover rate of ~ 600 molecules/second). This method can detect a single colony-forming unit (CFU) of E. coli in one liter of water with an overnight culture-based assay, or 50 CFUs of E. coli in 1 liter of water (or 10 mL of orange juice, or 10 mL of skim milk) in less than four hours with a solution-based assay with visual readout. The solution-based assay does not require specialized equipment or access to a laboratory, and is more rapid than existing tests that are suitable for use at the point of access. This method could be applied to the detection of many different bacteria, in parallel, with bacteriophages that express enzymes not natively expressed in the target bacteria.Chemistry and Chemical Biolog

Bioinspired self-repairing slippery surfaces with pressure-stable omniphobicity

Creating a robust synthetic surface that repels various liquids would have broad technological implications for areas ranging from biomedical devices and fuel transport to architecture but has proved extremely challenging. Inspirations from natural nonwetting structures particularly the leaves of the lotus, have led to the development of liquid-repellent microtextured surfaces that rely on the formation of a stable air–liquid interface. Despite over a decade of intense research, these surfaces are, however, still plagued with problems that restrict their practical applications: limited oleophobicity with high contact angle hysteresis, failure under pressure and upon physical damage inability to self-heal and high production cost. To address these challenges, here we report a strategy to create self-healing, slippery liquid-infused porous surface(s) (SLIPS) with exceptional liquid- and ice-repellency, pressure stability and enhanced optical transparency. Our approach—inspired by Nepenthes pitcher plants—is conceptually different from the lotus effect, because we use nano/microstructured substrates to lock in place the infused lubricating fluid. We define the requirements for which the lubricant forms a stable, defect-free and inert ‘slippery’ interface. This surface outperforms its natural counterparts and state-of-the-art synthetic liquid-repellent surfaces in its capability to repel various simple and complex liquids (water, hydrocarbons, crude oil and blood), maintain low contact angle hysteresis (<2.5°), quickly restore liquid-repellency after physical damage (within 0.1–1 s), resist ice adhesion, and function at high pressures (up to about 680 atm). We show that these properties are insensitive to the precise geometry of the underlying substrate, making our approach applicable to various inexpensive, low-surface-energy structured materials (such as porous Teflon membrane). We envision that these slippery surfaces will be useful in fluid handling and transportation, optical sensing, medicine, and as self-cleaning and anti-fouling materials operating in extreme environments.Engineering and Applied Science

Cofabrication: A Strategy for Building Multicomponent Microsystems

This Account describes a strategy for fabricating multicomponent microsystems in which the structures of essentially all of the components are formed in a single step of micromolding. This strategy, which we call “cofabrication”, is an alternative to multilayer microfabrication, in which multiple layers of components are sequentially aligned (“registered”) and deposited on a substrate by photolithography. Cofabrication has several characteristics that make it an especially useful approach for building multicomponent microsystems. It rapidly and inexpensively generates correctly aligned components (for example, wires, heaters, magnetic field generators, optical waveguides, and microfluidic channels) over very large surface areas. By avoiding registration, the technique does not impose on substrates the size limitations of common registrations tools, such as steppers and contact aligners. We have demonstrated multicomponent microsystems with surface areas exceeding 100 $cm^2$, but in principle, device size is only limited by the requirements of generating the original master. In addition, cofabrication can serve as a low-cost strategy for building microsystems. The technique is amenable to a variety of laboratory settings and uses fabrication tools that are less expensive than those used for multistep microfabrication. Moreover, the process requires only small amounts of solvent and photoresist, a costly chemical required for photolithography; in cofabrication, photoresist is applied and developed only once to produce a master, which is then used to produce multiple copies of molds containing the microfluidic channels. From a broad perspective, cofabrication represents a new processing paradigm in which the exterior (or shell) of the desired structures are produced before the interior (or core). This approach, generating the insulation or packaging structure first and injecting materials that provide function in channels in liquid phase, makes it possible to design and build microsystems with component materials that cannot be easily manipulated conventionally (such as solid materials with low melting points, liquid metals, liquid crystals, fused salts, foams, emulsions, gases, polymers, biomaterials, and fragile organics). Moreover, materials can be altered, removed, or replaced after the manufacturing stage. For example, cofabrication allows one to build devices in which a liquid flows through the device during use, or is replaced after use. Metal wires can be melted and reset by heating (in principle, repairing a break). This method leads to certain kinds of structures, such as integrated metallic wires with large cross-sectional areas or optical waveguides aligned in the same plane as microfluidic channels, that would be difficult or impossible to make with techniques such as sputter deposition or evaporation. This Account outlines the strategy of cofabrication and describes several applications. Specifically, we highlight cofabricated systems that combine microfluidics with (i) electrical wires for microheaters, electromagnets, and organic electrodes, (ii) fluidic optical components, such as optical waveguides, lenses, and light sources, (iii) gels for biological cell cultures, and (iv) droplets for compartmentalized chemical reactions, such as protein crystallization.Chemistry and Chemical Biolog