Biosensing using porous silicon double-layer interferometers: reflective interferometric Fourier transform spectroscopy

A simple, chip-based implementation of a double-beam interferometer that can separate biomolecules based on size and that can compensate for changes in matrix composition is introduced. The interferometric biosensor uses a double-layer of porous Si comprised of a top layer with large pores and a bottom layer with smaller pores. The structure is shown to provide an on-chip reference channel analogous to a double-beam spectrometer, but where the reference and sample compartments are stacked one on top of the other. The reflectivity spectrum of this structure displays a complicated interference pattern whose individual components can be resolved by fitting of the reflectivity data to a simple interference model or by fast Fourier transform (FFT). Shifts of the FFT peaks indicate biomolecule penetration into the different layers. The small molecule, sucrose, penetrates into both porous Si layers, whereas the large protein, bovine serum albumin (BSA), only enters the large pores. BSA can be detected even in a large (100-fold by mass) excess of sucrose from the FFT spectrum. Detection can be accomplished either by computing the weighted difference in the frequencies of two peaks or by computing the ratio of the intensities of two peaks in the FFT spectrum.     

Adsorption of beta-hairpin peptides on the surface of water: a neutron reflection study

Neutron reflectivity has been used to determine the thickness and surface coverage of monolayers of two 14-residue beta-hairpin peptides adsorbed at the air/water interface. The peptides differed only in that one was labeled with a fluorophore, while the other was not. The neutron reflection measurements were mainly made in null reflecting water, NRW, containing 8.1% D(2)O. Under this isotopic contrast the water is invisible to neutrons and the specular signal was then only from the peptide layer. At the highest concentration of ca. 4 microg/mL studied, the area per peptide molecule (A) was found to be 230 +/- 10 and 210 +/- 10 A(2) for the peptides with and without a BODIPY-based fluorophore, respectively. The thickness of the peptide layers was about 10 A for a Gaussian distribution. With decreasing bulk peptide concentration, both surface excess and layer thickness showed a steady trend of decrease. While the neutron results clearly indicate structural changes within the peptide monolayers with increasing bulk concentration, the outstanding structural feature is the formation of rather uniform peptide layers, consistent with the structural characteristics typical of beta-strand peptide conformations. These structural features are well supported by the parallel measurements of the adsorbed layers in D(2)O. With this isotopic contrast the neutron reflectivity provides an estimate about the extent of immersion of the peptide layers into water. The results strongly suggest that the 14-mer peptide monolayers were fully afloat on the surface of water, with only the carboxy groups on Glu residues hydrated.     

Characterization of the alignment of a nematic liquid crystal using half leaky guided modes

The excitation of half leaky guided optical modes to characterize fully the optical tensor profile in a thin liquid crystal layer has been used to evaluate the effect of rubbed polyimide aligning layers on the alignment of a nematic liquid crystal. A cell fabricated with rubbed polyimide alignment surfaces was studied at a wavelength of 632.8 nm. The liquid crystalline layer is sandwiched between a high refractive index top glass plate and a low refractive index glass substrate. Angular dependent reflectivities are recorded using a coupling prism and matching fluid with the same index as the top glass plate. Careful fitting of the predictions from multilayer optics theory to the observed angle dependent polarization conversion and reflectivity data yields the director profile within the liquid crystal layer in great detail.     

Effect of density fluctuating supercritical carbon dioxide on polymer interfaces

We investigated an effect of CO2 sorption on the compatibility of immiscible polystyrene (PS) and polybutadiene (PB) bilayers by using in situ neutron reflectivity. By labeling either polymer with deuterium, we found that the excess CO2 molecules were adsorbed to both top PS and bottom PB layers when the bilayers were exposed to CO2 at the narrow T and P regime near the critical point of pure CO2. Furthermore, we clarified that this excess sorption of CO2 molecules increased the interfacial width between the layers up to 100 angstroms even near room temperature, while the interfacial width without CO2 exposure has been reported to be at most 40 A even at the highest temperature (T congruent with 175 degrees C).     

Complex layering observed in high internal phase emulsions at a silicon surface by neutron reflectometry

The neutron reflectivity profiles from the interface between silicon and aqueous phase-in-oil high internal phase emulsions of steadily increasing surfactant hydrophilicity, are reported for two isotopic contrasts for each surfactant. Layered models are required to fit the structured reflectivity profiles that demonstrate that the oxidised top layer of the silicon is always covered by a surfactant monolayer. Interposed between the surfactant monolayer and the bulk emulsion is a layer of oil--a geometric effect caused by reorganisation of the aqueous droplets. As the surfactant hydrophilicity increases, alternating aqueous and oil+surfactant layers are inserted between this topmost oil layer and the oxide attached surfactant monolayer. The resulting structures have compositions and layer spacings suggestive of sections from lamellar phases. This increase in layer ordering with increasing surfactant hydrophilicity is expected. The bulk emulsions are observed to exhibit lamellar or sponge phases increasingly as surfactant hydrophilicity increases.     

Structural rearrangements during the initial growth stages of organic thin films of F16CuPc on SiO2

We present an experimental study on the first stages of the thin film growth of the organic molecule F(16)CuPc (hexadecafluoro-copper-phthalocyanines) on SiO(2). By means of in situ X-ray reflectivity, in situ grazing incidence X-ray diffraction (GIXD), and ex situ atomic force microscopy (AFM), we provide a detailed picture of the film growth mode and its structural evolution at the nanometer scale. We discovered the formation of a low-density layer of molecular aggregates with heights between 5 and 10 A at the interface with the SiO(2) and show that, on top of this interfacial layer, the nucleation and two-dimensional growth of elongated islands of upright standing molecules take place. Structural changes are observed, pointing to significant relaxations of the lattice parameters within the first layers of standing molecules.     

Gas permeability in polymer- and surfactant-stabilized bubble films

The gas permeabilities of thin liquid films stabilized by poly(N-isopropylacrylamide) (PNIPAM) and PNIPAM-SDS (sodium dodecyl sulfate) mixtures are studied using the "diminishing bubble" method. The method consists of forming a microbubble on the surface of the polymer solution and measuring the shrinking rates of the bubble and the bubble film as the gas diffuses from the interior to the exterior of the bubble. PNIPAM-stabilized films exhibit variable thicknesses and homogeneities. Interestingly, despite these variable features, the gas permeability of the film is determined principally by the structure of the adsorbed polymer layer that provides an efficient gas barrier with a value of gas permeability coefficient that is comparable to that of an SDS Newton black film. In the presence of SDS, both the film homogeneity and the gas permeability coefficient increase. These changes are related to interactions of PNIPAM with SDS in the solution and at the interface, where coadsorption of the two species forms mixed layers that are stable but that are more porous to gas transfer. The mixed PNIPAM-SDS layers, studied previously for a single water-air interface by neutron reflectivity, are further characterized here in a vertical free-draining film using X-ray reflectivity.     

Annealing sequence dependent open-circuit voltage of inverted polymer solar cells attributable to interfacial chemical reaction between top electrodes and photoactive layers

The ability to laminate and delaminate top metal contacts during the processing and testing of inverted polymer solar cells has led us to uncover the peculiar dependence of their open-circuit voltage (V(oc)) on the annealing sequence. Specifically, thermally annealing inverted polymer solar cells having bulk-heterojunction photoactive layers after top electrode deposition above 100 °C leads to lower V(oc) compared to analogous devices with unannealed photoactive layers or photoactive layers that have been annealed prior to metal electrode deposition. This reduction in V(oc), however, can be reversed when the top electrodes are replaced. This observation is thus a strong indication that such changes in V(oc) with annealing sequence are manifestations of changes at the top electrode-photoactive layer interface, and not structural changes in the bulk of the photoactive layer. Electronic characterization conducted on the photoactive layers and metal contacts after dissection of the polymer solar cells via delamination suggests the reduction of V(oc) on thermal annealing in the presence of the metal top contacts to stem from an interfacial chemical reaction between the photoactive layers and the metal electrodes. This chemically generated interfacial layer is removed upon electrode delamination, effectively reverting the V(oc) to its original value prior to thermal annealing when the top electrodes are replaced.     

A mathematical model describing voltammograms and transport numbers under intensive electrodialysis modes

A three-layer mathematical model of overlimiting state is developed. A reactive layer with a thickness depending on the current density is introduced into the model. A decrease in the thickness of diffusion layer, which donates the counterions, with increasing current density as a result of electroconvection is also taken into account. A boundary-value problem is formulated within the Nernst-Planck and Poisson’s model in the three-layer region with the boundary conditions of constant concentrations in the bulk solution. It is shown that an increase in the reactive layer thickness with increasing current density determines the behavior of effective transport numbers in the overlimiting state of ion-exchange electromembrane system. In the current range under consideration (from 1 to 20 limiting currents), the reactive layer thickness is several tens nanometers and reaches 70 nm at a 100-fold excess over the limiting current. To calculate the voltammograms, the dependence of effective thickness δ_N of diffusion layer on the current density is required. This dependence can be obtained by solving an inverse problem, from the laser interferometry experiments, or calculated by the Navier-Stokes hydrodynamic model. The model enables one to calculate the distribution of electric field strength, potential, concentrations in the diffusion layers and membrane.     

Surface enrichment of proteins at quartz/water interfaces: a neutron reflectivity study

Neutron reflectivity (NR) was used to study the adsorption of human serum albumin and human fibrinogen on quartz. The proteins were individually and sequentially adsorbed from heavy water and heavy water/methanol mixtures at pH 4 and 7.0. The technique allows for the subnanometer resolution of the adsorbed layer thickness and gross morphology. Under the conditions of our measurements we found that fibrinogen formed a distinct layer that we interpret as a mat of the protein three layers thick whereas albumin formed only diffuse layers. The adsorption pattern of the two proteins changed radically when one protein was adsorbed on top of the other (previously adsorbed). In general our measurements indicate that the adsorbed protein layers on quartz are rather loosely bound and that these layers, incorporating as much as 80% water, extend further into the bulk fluid than might have been expected.     

Adsorption and Exchange Dynamics in Aging Hydroxyethylcellulose Layers on Silica

The adsorption kinetics of hydroxyethylcellulose (HEC) on silica and relaxations in adsorbed HEC layers were probed using total internal reflectance fluorescence and near-Brewster reflectivity. Like many random-coil polymers, HEC was found to adsorb at the transport-limited rate. Relaxations occurred at nearly constant interfacial mass when HEC layers were exposed to aqueous solvent, causing the subsequent exchange of chains between the layer and the free solution to become increasingly hindered. Eventually, on the time scale of a day, layers became immobilized and unable to accommodate chains from free solution. A continued fluorescence decay, beyond time scales that could be probed with self exchange, suggested further relaxations of the adsorbed HEC. The polydisperse HEC system (with an average molecular weight near 450,000) behaved qualitatively similar to molecular weight standard polyethylene oxide (PEO) layers on silica. For instance, relaxations in PEO layers occurred on a time scale of 10-20 h, like the HEC layers. Young layers of the latter, however, exhibited self-exchange kinetics that were an order of magnitude slower than PEO layers of similar age. This difference in adsorbed layer dynamics was attributed to HEC's stiffer backbone, compared with flexible PEO. Copyright 2000 Academic Press.     

Adsorption configuration and local ordering of silicotungstate anions on Ag(100) electrode surfaces

X-ray reflectivity, cyclic voltammetry, and scanning tunneling microscopy (STM) are used to examine the structure of alpha-SiW12O4(4-) or silicotungstic acid (STA) adsorbed on Ag(100) in acid solution. The voltammetry shows that STA passivates the Ag surface relative to electron transfer to a solution redox species. STM images reveal the formation of a series of lattice structures, one of which can be associated with a commensurate ( radical13x radical13)R33.69 degrees structural model. X-ray reflectivity measurements show uniquely that STA orients with its four-fold axis perpendicular to the Ag(100) surface and that the center of the STA molecule is 4.90 A above the top layer of the Ag substrate. Analysis of bond lengths leads to a footprint of STA on Ag(100), in which the four terminal O atoms are located near the hollow sites and have a Ag-O bond length of 2.06 A. This bond length is consistent with a strong covalent interaction between STA and the Ag surface.     

The growth of compact layers at the electrode interface: Part IV. Fractal character of uracil films at the mercury-aqueous electrolyte interface

Compact layers of uracil films grown on the mercury electrode-aqueous solution interface were studied by means of time-resolved FFT impedance spectroscopy. The films are characterized by the fractal dimension which evolves with time. The results are discussed in terms of the cluster-cluster aggregation model and the hole formation mechanism proceeding in the compact layer.     

Release of a dye from hydrogen-bonded and electrostatically assembled polymer films triggered by adsorption of a polyelectrolyte

The absorption of dyes within hydrogen-bonded and electrostatically assembled multilayers and subsequent release of the dyes from the films were studied in situ using FTIR-ATR. Multilayers were composed of poly(methacrylic acid), PMAA, and poly(ethylene oxide), PEO (hydrogen-bonded multilayers), or of PMAA and 22% quarternized copolymer of N-ethyl-4-vinylpyridium bromide and 4-vinylpyridine, Q22 (electrostatically stabilized multilayers). After multilayer deposition, the solution pH was changed to produce excess charge within the films. Dyes with charge opposite to the excess charge of the film (Rhodamine 6G for hydrogen-bonded films or Bromophenol Blue for electrostatically assembled multilayers) were then allowed to absorb within multilayers. In both systems, the dyes were uniformly included within the films. The top layers largely affected the loading capacity of the multilayers, suggesting weaker binding of the dyes with the top layers. Dye release into a 0.01 M phosphate buffer was significantly smaller as compared to release in the presence of 0.05-0.5 mg/mL solutions of adsorbing polymers whose charge was the same as the excess charge within the films. We found that with the PMAA/PEO films, dye release did not depend on the concentration of polymer in solution, but was largely controlled by the amount of charge accumulated within the adsorbing polymer layer on the top of the film. For electrostatically stabilized PMAA/Q22 systems, dye release increased with increasing concentration of Q22 in solution, suggesting a significant contribution of the competition of solution species in the release mechanism. Our findings contribute to the understanding of interactions of small molecules with polymer multilayers and might have ramifications for novel applications of multilayer films as new materials for the controlled delivery of chemicals.     

Temperature-triggered micellization of block copolymers on an ionic liquid surface

In situ neutron reflectivity was used to study thermally induced structural changes of the lamellae-forming polystyrene-block-poly(2-vinylpyridine) (PS-b-P2VP) block copolymer thin films floating on the surface of an ionic liquid (IL). The IL, 1-butyl-3-methylimidazolium trifluoromethanesulfonate, is a nonsolvent for PS and a temperature-tunable solvent for P2VP, and, as such, micellization can be induced at the air-IL interface by changing the temperature. Transmission electron microscopy and scanning force microscopy were used to investigate the resultant morphologies of the micellar films. It was found that highly ordered nanostructures consisting of spherical micelles with a PS core surrounded by a P2VP corona were produced. In addition, bilayer films of PS homopolymer on top of a PS-b-P2VP layer also underwent micellization with increasing temperature but the micellization was strongly dependent on the thickness of the PS and PS-b-P2VP layers.     

Surface-induced unfolding of human lactoferrin

We have determined the structural conformations of human lactoferrin adsorbed at the air/water interface by neutron reflectivity (NR) and its solution structure by small angle neutron scattering (SANS). The neutron reflectivity measurements revealed a strong structural unfolding of the molecule when adsorbed at the interface from a pH 7 phosphate buffer solution (PBS with a total ionic strength at 4.5 mM) over a wide concentration range. Two distinct regions, a top dense layer of 15-20 angstroms on the air side and a bottom diffuse layer of some 50 angstroms into the aqueous subphase, characterized the unfolded interfacial layer. At a concentration around 1 g dm(-3), close to the physiological concentration of lactoferrin in biological fluids, the adsorbed amount was 5.5 x 10(-8) mol m(-2) in the absence of NaCl, but the addition of 0.3 M NaCl reduced protein adsorption to 3.5 x 10(-8) mol m(-2). Although the polypeptide distributions at the interface remained similar, quantitative analysis showed that the addition of NaCl reduced the layer thickness. Parallel measurements of lactoferrin adsorption in D2O instead of null reflecting water confirmed the unfolded structure at the interface. Furthermore, the D2O data indicated that the polypeptide in the top layer was predominantly protruded out of water, consistent with it being hydrophobic. In contrast, the scattering intensity profiles from SANS were well described by a cylindrical model with a diameter of 47 angstroms and a length of 105 angstroms in the presence of 0.3 M NaCl, indicating a retention of the globular framework in the bulk solution. In the absence of NaCl but with the same amount of phosphate buffer, the length of the cylinder increased to some 190 angstroms and the diameter remained constant. The length increase is indicative of changes in distance and orientation between the bilobal monomers due to the change in charge interactions. The results thus demonstrate that the surface structural unfolding was caused by the exposure of the protein molecule to the unsymmetrical energetic balance following surface adsorption.     

Radiometric trace analysis of cobalt with diethyldithiocarbamate-35S,or 203Hg

Two radiometric methods for the determination of submugram amounts of cobalt are described. (A) Cobalt is extracted from an ammoniacal solution with a zinc-diethyldithiocarbamate-³⁵S solution in chloroform. Excess reagent and interfering metals are removed with mercury(II) and cyanide. The ³⁵S in the final organic layer is a measure of the cobalt in this layer. (B) Cobalt is extracted from an ammoniacal solution with a fixed amount of zinc-DDC in chloroform. Excess reagent and complexes of foreign metals are removed by exchange with ²⁰³Hg⁺² and the ²⁰³Hg in the chloroform (compared with a blank) acts as a measure of the cobalt. Method A is applicable to 0.1 μg of cobalt and method B to 0.8 μg. As the efficiency of both processes is variable, isotope dilution with ⁶⁰Co is carried out, A 10-fold excess of foreign metals is permitted in method A and a 4-fold excess in method B ; larger amounts are previously removed, e.g. by extraction with inactive zinc-DDC from sodium hydroxide media.     

Surface and solution behavior of the mixed dialkyl chain cationic and nonionic surfactants

The surface and solution behavior of the mixed dialkyl chain cationic and nonionic surfactant mixture of dihexadecyldimethylammonium bromide, DHDAB, and hexaethylene monododecyl ether, C12E6, has been investigated, using primarily the scattering techniques of small-angle neutron scattering and neutron reflectivity. Within the time scale of the measurements, the adsorption of the pure component C12E6 at the air-solution interface shows no time dependence. In contrast, the adsorption of the DHDAB/C12E6 mixture and pure DHDAB has a pronounced time dependence. The characteristic time for adsorption varies with surfactant concentration, composition, and temperature. It is approximately 2-3 h for the DHDAB/C12E6 mixture, dependent upon concentration and composition, and approximately 50 min for DHDAB. At the air-solution interface, the equilibrium composition of the adsorbed layer shows a marked departure from ideal mixing, which is dependent upon both the solution concentration and the concentration of added electrolyte. In contrast, the composition of the aggregates in the bulk solution that are in equilibrium with the surface is close to ideal mixing, as expected for solution concentrations well in excess of the critical micellar concentration. The structure of the mixed adsorbed layer has been measured and compared with the structure of the equivalent pure surfactant monolayer, and no substantial changes in structure or conformation are observed. The extreme departure from ideal mixing in the adsorption behavior of the DHDAB/C12E6 mixture is discussed in the context of the structure of the adsorbed layer, changes in the underlying solution structures, and the failure of regular solution theory to predict such behavior.     

Structure and dynamics of crystalline protein layers bound to supported lipid bilayers

We study proteins at the surface of bilayer membranes using streptavidin and avidin bound to biotinylated lipids in a supported lipid bilayer (SLB) at the solid-liquid interface. Using X-ray reflectivity and simultaneous fluorescence microscopy, we characterize the structure and fluidity of protein layers with varied relative surface coverages of crystalline and noncrystalline protein. With continuous bleaching, we measure a 10-15% decrease in the fluidity of the SLB after the full protein layer is formed. We propose that this reduction in lipid mobility is due to a small fraction (0.04) of immobilized lipids bound to the protein layer that create obstacles to membrane diffusion. Our X-ray reflectivity data show a 40 A thick layer of protein, and we resolve an 8 A layer separating the protein layer from the bilayer. We suggest that the separation provided by this water layer allows the underlying lipid bilayer to retain its fluidity and stability.     

Generation of complex, static solution gradients in microfluidic channels

A microfluidic device is used to generate a complex gradient of diffusible molecules in a static solution. The gradient is precise and steady both in space and in time. This device, made from poly(dimethylsiloxane), consists of three layers. The molecules in reservoirs on the top layer diffuse through the flat middle layer of hydrogel and reach an equilibrium distribution. Microfluidic channels on the bottom layer that are in close contact with the hydrogel contain free solution that has concentration gradients based on the gradient in the gel. The gradient profile in the channel can be designed to have an arbitrary form (within the range of the existing gradient in the hydrogel) by controlling the local direction of the channel at each point.