Planar lipid bilayer reconstitution with a micro-fluidic system

A planar lipid bilayer which is widely used for the electrophysiological study of membrane proteins in laboratories is reconstituted using a micro-fluidic system, in a manner that is suitable for automated processing. We fabricated micro-channels on both sides of the substrate, which are connected through a 100-200 microm aperture, and showed that the bilayer can be formed at the aperture by flowing the lipid solution and buffer, alternately. Parylene coating is found to be suitable for both bilayer formation and electric noise reduction. Future applications include a high-sensitivity ion sensor chip and a high-throughput drug screening device.     

Biophysical aspects of agar-gel supported bilayer lipid nembranes: a new method for forming and studying planar bilayer lipid membranes

Conventional bilayer lipid membranes (BLMs), formed by either the painting method or the Langmuir-Blodgett technique, lack the desired stability. This paper presents a simple method for forming long-lived BLMs on agar-gel supports. The supported BLM reported has a greatly improved mechanical stability and also has desirable dynamic properties. These self-assembled BLMs are of significant interest, in view of their similarity of biological membranes, their molecular dimension and their spontaneous formation.     

Highly selective acylation of ferrocene using microfluidic chip reactor

A rapid and highly selective acylation of ferrocene with various acid anhydrides using microfluidic chip as the reactor is described. The pressure driven glass microreactor was fabricated by standard photolithography and wet etching techniques. High conversions of ferrocene to the corresponding acylferrocenes were achieved at 25 °C and no diacylferrocene was observed in any case.     

Controlled delivery of proteins into bilayer lipid membranes on chip

The study and the exploitation of membrane proteins for drug screening applications requires a controllable and reliable method for their delivery into an artificial suspended membrane platform based on lab-on-a-chip technology. In this work, a polymeric device for forming lipid bilayers suitable for electrophysiology studies and biosensor applications is presented. The chip supports a single bilayer and is configured for controlled protein delivery through on-chip microfluidics. In order to demonstrate the principle of protein delivery, the potassium channel KcsA was reconstituted into proteoliposomes, which were then fused with the suspended bilayer on-chip. Fusion of single proteoliposomes with the membrane was identified electrically. Single channel conductance measurements of KcsA in the on-chip bilayer were recorded and these were compared to previously published data obtained with a conventional planar bilayer system.     

The sequence length in polymethyl methacrylate

The sequence length and tacticity of methyl methacrylate polymers prepared at 303–363°K in the presence of trichlorobromomethane as a transfer agent were measured from NMR spectra. The lengths did not change appreciably but there was an increase in syndiotacticity with increasing concentration of transfer agent. There is no marked change in tacticity with molecular weight.     

Hydrodynamic effects on spinodal decomposition kinetics in planar lipid bilayer membranes

The formation and dynamics of spatially extended compositional domains in multicomponent lipid membranes lie at the heart of many important biological and biophysical phenomena. While the thermodynamic basis for domain formation has been explored extensively in the past, domain growth in the presence of hydrodynamic interactions both within the (effectively) two-dimensional membrane and in the three-dimensional solvent in which the membrane is immersed has received little attention. In this work, we explore the role of hydrodynamic effects on spinodal decomposition kinetics via continuum simulations of a convective Cahn-Hilliard equation for membrane composition coupled to the Stokes equation. Our approach explicitly includes hydrodynamics both within the planar membrane and in the three-dimensional solvent in the viscously dominated flow regime. Numerical simulations reveal that dynamical scaling breaks down for critical lipid mixtures due to distinct coarsening mechanisms for elongated versus more isotropic compositional lipid domains. The breakdown in scaling should be readily observable in experiments on model membrane systems.     

Miniaturized biochemical sensing devices based on planar bilayer lipid membranes

Methods of in-vitro artificial formation of bilayer lipid membranes (BLM) and their analytical applications are reviewed, on the basis of 122 literature references. Different techniques for preparation of free-suspended planar BLMs, and gel-, filter-, and solid-supported systems are presented. The analytical applications developed are based on direct interaction of analytes with chemically unmodified BLMs, and with systems modified by use of redox mediators, ionophores, ion-channel forming species, enzymes, antibodies, or DNA.     

Diffusion of water vapour in polymethyl methacrylate

Certain aspects of the concentration dependence of the diffusion coefficient and on the mechanism of sorption of water vapour in PMMA are investigated. Early time integral sorption kinetics follow the Fickian pattern of initial linearity. Diffusion coefficients deduced from the slopes of these lines are independent of concentration and satisfactorily agree with those obtained from late-time analysis; however, desorption values are higher throughout the whole range of activities than the corresponding absorption ones. On the contrary, typical “non-Fickian” features are discerned in differential absorption kinetics on membranes of different thickness. These anomalies are attributed to rate-determining relaxation processes concurrent with diffusion, whereas other sorption models are incompatible with the general behaviour of this system. General trends of the equilibrium and kinetic data are compared with those of other polymer—water systems and the observed differences are interpreted in terms of the molecular relaxation model.     

Local anesthetics facilitate ion transport across lipid planar bilayer membranes under an electric field: dependence on type of lipid bilayer

In order to elucidate the role of structural change of lipid membrane bilayer in the mode of action of local anesthetic, we studied the effects of local anesthetics, charged tetracaine and uncharged benzocaine, on ion permeability across various lipid planar bilayers (PC, mixed PC/PS (4/1, mol/mol); mixed PC/PE (1/1, mol/mol); mixed PC/SM (4/1, mol/mol)) under a constant applied voltage. The membrane conductances increased in the order of PC  PC/PS ≤ PC/SM  PC/PE. When the constant voltage of −100 or −70 mV was applied through the lipid bilayer membranes in the presence of positively charged tetracaine, the fluctuating current pulses with the large amplitude generated, but not appeared in the absence of tetracaine. The addition of uncharged benzocaine generated the fluctuating currents with the small amplitude. Both charged tetracaine and uncharged benzocaine facilitated electrophoretically the transport of small ions such as KCl in the buffer solution through the fluctuating pores in the lipid bilayer membranes formed by interaction with the local anesthetic under the negative applied membrane potential. The current pulses also contained actual transport of charged tetracaine together with the transport of the small ions. The amplitude and the duration time of the electrical current generated by adding the local anesthetics were dependent on the type of the lipid, the applied voltage and its voltage polarity.     

Photochemical reaction of nitroanthracene derivatives in polymethyl methacrylate

Irradiation of 9-cyano-10-nitroanthracene or 9-benzoyl-10-nitroanthracene in polymethyl methacrylate (PMMA), at room temperature, gives the 9-cyano- or 9-benzoyl-10-anthryloxy radical which disappears slowly after irradiation, following the second order dark reaction, and finally gives the substituted anthranol. In the presence of TEA, the corresponding anthryloxy anions are obtained and their absorption spectra are greatly red-shifted compared with those in fluid basic polar solutions. The existence of 9-benzoyl-10-anthryloxy anion in basic PMMA is very interesting, because 9-benzoyl-10-anthranol is obtained in methyl methacrylate monomer (MMA) even in the presence of TEA. This is explained in terms of the increase of the microscopic medium polarity of MMA by the polymerisation.     

Deposition of polymethyl methacrylate latex on fibers

Summary In order to study the deposition of polymethyl methacrylate latex on fibers,viz., polyamide (Nylon 6), polyacrylonitrile (Vonnel), polyester (Tetoron), and cotton, the amount of latex deposited was measured and the interaction energy between fiber and latex particles was determined from -potentials by the application of heterocoagulation theory. The amount of latex deposited decreased and the total interaction energy increased with increasing pH for all fibers. However, it was found that the deposition of latex on Vonnel and cotton could not be interpreted satisfactorily by only the interaction energy, because latex deposited on those fibers in spite of the large total interaction energy.From the observation of the state of deposition of latex particles in terms of scanning electron microscope, it was found that latex particles deposited on Nylon and Tetoron in a single particle layer, while those on Vonnel in a multiple or blocky layer. It was suggested that the deposition of latex on fiber was influenced by the surface roughness of fiber and the uniformity of the filament diameter.

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Zusammenfassung Um die Ablagerung von Polymethylmethacrylat auf Fasern Polyamid (Nylon 6), Polyacrylnitril (Vonnel), Polyester (Tetoron) and Baumwolle zu studieren, wurde die Menge des abgelagerten Latex gemessen, and die Wechselwirkungsenergie zwischen Faser and Latex-Teilchen wurde aus den -Potentialen durch die Anwendung der Heterokoagulationstheorie bestimmt. Die Menge des abgelagerten Latex nahm ab und die totale Wechselwirkungsenergie stieg mit steigendem pH für alle Fasern an. Jedoch zeigte sich, daß die Ablagerung von Latex an Vonnel und Baumwolle nicht zufriedenstellend allein durch die Wechselwirkungsenergie erklärt werden konnte, weil Latex sich an diese Fasern trotz der großen totalen Wechselwirkungsenergie ablagerte.Aus der Beobachtung des Ablagerungszustandes von Latex-Teilchen im Rasterelektronenmikroskop fand man heraus, daß Latex-Teilchen sich an Nylon und Tetoron in einer einfachen, an Vonnel aber in einer mehrfachen oder dickeren Schicht ablagerten. Es wurde darauf hingewiesen, daß die Ablagerung von Latex an Fasern durch die Oberflächenrauheit der Fasern und die Einheitlichkeit des Faser-Durchmessers beeinflußt wurde.

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With 7 figures     

Influence of inhalation anesthetics on ion transport across a planar bilayer lipid membrane

Ion transport from one aqueous phase (W1) to another (W2) across a planar bilayer lipid membrane (BLM) in the presence of inhalation anesthetics was electrochemically investigated. In the absence of inhalation anesthetics in the BLM system, no ion transport current flowed between W1 and W2 across the BLM. When inhalation anesthetics such as halothane, chloroform, diethyl ether and trichloroethylene were added to the two aqueous phases or the BLM, the ion transport current quite clearly appeared. When the ratio of the concentration of KCl or NaCl in W1 to that in W2 was varied, the zero current potential across the BLM was shifted. By considering the magnitude of the potential shift, we concluded that the ion transport current can be predominantly ascribed to the transport of Cl(-) across the BLM. Since the dielectric constants of these anesthetics are larger than that of the inner hydrophobic domain of the BLM, the concentration of hydrophilic electrolyte ions in the BLM increases with the increase in the dielectric constant of the inner hydrophobic domain caused by addition of these anesthetics. These situations lead to an increase in the ion permeability coefficient.     

Interaction of charged lipid vesicles with planar bilayer lipid membranes: detection by antibiotic membrane probes.

A technique has been developed for monitoring the interaction of charged phospholipid vesicles with planar bilayer lipid membranes (BLM) by use of the antibiotics Valinomycin, Nonactin, and Monazomycin as surface-charge probes. Anionic phosphatidylserine vesicles, when added to one aqueous compartment of a BLM, are shown to impart negative surface charge to zwitterionic phosphatidylcholine and phosphatidylethanolamine bilayers. The surface charge is distributed asymmetrically, mainly on the vesicular side of the BLM, and is not removed by exchange of the vesicular aqueous solution. Possible mechanisms for the vesicle-BLM interactions are discussed.     

Current development in microfluidic immunosensing chip

This review accounts for the current development in microfluidic immunosensing chips. The basic knowledge of immunoassay in relation to its microfluidic material substrate, fluid handling and detection mode are briefly discussed. Here, we mainly focused on the surface modification, antibody immobilization, detection, signal enhancement and multiple analyte sensing. Some of the clinically important currently implemented on the microfluidic immunoassay chips are C-reactive protein (CRP), prostate specific antigen (PSA), ferritin, vascular endothelial growth factor (VEGF), myoglobin (Myo), cardiac troponin T (cTnT), cardiac troponin I (cTnI), and creatine kinase-cardiac muscle isoform (CK-MB). The emerging microfludic immunosensor technology may be a promising prospect that can propel the improvement of clinical and medical diagnosis.     

Arachidonic acid-induced channel- and carrier-type ion transport across planar bilayer lipid membranes.

Transmembrane ion transport by arachidonic acid (AA) through bilayer lipid membranes (BLMs) was investigated by means of electrochemical measurements to provide a basis for designing a sensor membrane. We found that AA induces a channel-type current, in addition to a carrier-type current, across planar BLMs. A linear relation between the logarithmic value of the AA concentration and the current responses (given as integrated currents) was observed for a carrier-type current, while a sigmoid relation was found for a channel-type current. Although AA transports Na+, Ca2+ and Mg2+ and exhibits ion selectivity between Na+ and Mg2+ for the carrier-type current, ion transport for the channel-type current was non-selective. It was found that ion transport via the channel mechanism occurs frequently for AA, while channel-type currents were only occasionally observed for y-linolenic acid and prostaglandin D2. No channel-type currents were induced by other fatty acids (oleic, linoleic, stearic, myristic, eicosapentanoic and docosahexanoic acids) and metabolites of AA (12-HETE and 5-HETE). The carrier-type ion transport occurs selectively to these compounds if the concentration is below 1.0 microM. These results suggest that AA selectively facilitates an ion flux through the BLMs, generating channel-type and/or carrier-type currents, which can be used as a measure of the AA concentration.     

Application of polymethyl methacrylate in cathode materials of lithium-ion batteries

A study of the Li₂FeSiO₄/C cathode material doped with Mn demonstrated that introduction of polymethyl methacrylate results in a substantial decrease in the particle size and increase in the specific surface area of the cathode material. Polymethyl methacrylate strongly improves the cyclic stability of the cathode material. The discharge capacity after the first cycle was 218 mA h g^–1, and that upon stabilization of the structure of the cathode material, 170 mA h g^–1.     

On the kinetics of radical oxidation in polymethyl methacrylate films

Radical oxidation controlled by oxygen diffusion in a solid medium consisting of regions of size »» 60 with different diffusion coefficients, has been studied by computer simulation. It is shown that the rate constant distribution coincides with the diffusion coefficient distribution within a wide range of oxygen concentrations.     

Flexible planar microfluidic chip employing a light emitting diode and a PIN-photodiode for portable flow cytometers

Detection of fluorescence particles is a key method of flow cytometry. We evaluate the performance of a design for a microfluidic fluorescence particle detection device. Due to the planar design with low layer thicknesses, we avoid optical components such as lenses or dichroic mirrors and substitute them with a shadow mask and colored film filters. A commercially available LED is used as the light source and a PIN-photodiode as detector. This design approach reduces component cost and power consumption and enables supplying the device with power from a standard USB port. From evaluation of this design, we obtain a maximum particle detection frequency of up to 600 particles per second at a sensitivity of better than 4.7 × 10(5) MESF (molecules of equivalent soluble fluorochrome) measured with particles for FITC sensitivity calibration. Lowering the flow rate increases the instrument sensitivity by an order of magnitude enabling the detection of particles with 4.5 × 10(4) MESF.