Null Ellipsometry Research Articles

Biochip Surfaces Containing Recombinant Cell-Binding Domains of Fibronectin

Surface immobilization and characterization of the functional activity of fibronectin (Fn) type-III domains are reported. The domains FnIII9-10 or FnIII10 containing the RGD loop and PHSRN synergy site were recombinantly produced and covalently bound to chemically activated PEG methacrylate (MA) hydrogel coatings by microcontact printing. Such fabricated biochip surfaces were 6 mm in diameter and consisted of 190 µm wide protein stripes separated by 200 µm spacing. They were analyzed by imaging null ellipsometry, atomic force microscopy and fluorescence microscopy. Also, the coatings were tested in human foreskin fibroblast and HeLa cultures for at least 96 h, thus evaluating their suitability for controlled cell adhesion and proliferation. However, while HeLa cultures were equally well responsive to the FnIII9-10, FnIII10 and Fn surfaces, the fibroblasts displayed lower cell and lower focal adhesion areas, as well as lower proliferation rates on the Fn fragment surfaces as compared to Fn. Nevertheless, full functional activity of the fibroblasts was confirmed by immunostaining of Fn produced by the cells adherent on the biochip surfaces. The observed interaction differences that were either cell type or surface composition-dependent demonstrate the potential use of specifically engineered Fn and other ECM protein-derived domains in biochip architectures.

A comparison between the structures of reconstituted salivary pellicles and oral mucin (MUC5B) films

HypothesisSalivary pellicles i.e., thin films formed upon selective adsorption of saliva, protect oral surfaces against chemical and mechanical insults. Pellicles are also excellent aqueous lubricants. It is generally accepted that reconstituted pellicles have a two-layer structure, where the outer layer is mainly composed of MUC5B mucins. We hypothesized that by comparing the effect of ionic strength on reconstituted pellicles and MUC5B films we could gain further insight into the pellicle structure. ExperimentsSalivary pellicles and MUC5B films reconstituted on solid surfaces were investigated at different ionic strengths by Force Spectroscopy, Quartz Crystal Microbalance with Dissipation, Null Ellipsometry and Neutron Reflectometry. FindingsOur results support the two-layer structure for reconstituted salivary pellicles. The outer layer swelled when ionic strength decreased, indicating a weak polyelectrolyte behavior. While initially the MUC5B films exhibited a similar tendency, this was followed by a drastic collapse indicating an interaction between exposed hydrophobic domains. This suggests that mucins in the pellicle outer layer form complexes with other salivary components that prevent this interaction. Lowering ionic strength below physiological values also led to a partial removal of the pellicle inner layer. Overall, our results highlight the importance that the interactions of mucins with other pellicle components play on their structure.

Photografting and Patterning of Poly(ethylene glycol) Methacrylate Hydrogel on Glass for Biochip Applications.

An efficient procedure for chemical initiator-free, in situ synthesis of a functional polyethylene glycol methacrylate (PEG MA) hydrogel on regular glass substrates is reported. It is demonstrated that self-initiated photografting and photopolymerization driven by UV irradiation can yield tens of nanometer-thick coatings of carboxy-functionalized PEG MA on the aldehyde-terminated borosilicate glass surface. The most efficient formulation for hydrogel synthesis contained methyl methacrylic acid (MAA), 2-hydroxyethyl methacrylate (HEMA), and PEG methacrylate (PEG10MA) monomers (1:1:1). The resulting HEMA/PEG10MA/MAA (HPMAA) coatings had a defined thickness in the range from 11 to 50 nm. The physicochemical properties of the synthesized HPMAA coatings were analyzed by combining water contact angle measurements, stylus profilometry, imaging null ellipsometry, and atomic force microscopy (AFM). The latter technique was employed in the quantitative imaging mode not only for direct probing of the surface topography but also for swelling behavior characterization in the pH range from 4.5 to 8.0. The estimated high swelling ratios of the HPMAA hydrogel (up to 3.2) together with its good stability and resistance to nonspecific protein binding were advantageous in extracellular matrix mimetics via patterning of fibronectin (FN) at a resolution close to 200 nm. It was shown that the fabricated FN micropatterns on HPMAA were equally suitable for single-cell arraying, as well as controlled cell culture lasting at least for 96 h.

Influence of Cu impurities and surface temperature to the formation of thin a-C:H(Me) film

The aim of this experimental work is identify most important physical and chemical processes on the surface during amorphous carbon films formation on the silicon surface with copper nanoclusters (Cu). Duration of films deposition on n type silicon (100) surface by using different substrate temperatures (25 °C, 100 oC and 250 °C) was 45s. Velocity of film growth from C2H2 gas plasma depended on surface temperature and was variable (0.2-0.5 nm/s). Data of null ellipsometry, Raman spectroscopy and element analysis showed that formation of amorphous carbon film is in the early stage and mixture of Si-C, Si-COH and GLC fragments is dominant on the surface. The experimental RS curves were fitted by few Gaussian-shape lines in the spectral range from 500 cm-1 to 700 cm-1, from 700 cm-1 to 900 cm-1, from 900 cm-1 to 1100 cm-1 and from 1100 cm-1 to 1900 cm-1 and analysis of the additional peaks in all ranges confirmed complexes of different carbons structures in the film. Carbon films with more ordered C-C bonds grows during film formation at higher temperatures (>100o C) on the silicon surface with Cu particles, because atoms of copper penetrate into the deeper layers of silicon. The substrate temperature influenced the surface amorphisation because of active oxygen and hydrogen diffusion into deeper layers and formation of hydrogenated silicon carbon with fragments of amorphous carbon films becomes not significant.

Comparison imaging ellipsometry and its application to crystallization of indium oxide thin films

Null imaging ellipsometry was developed to show the net change in polarization state between two samples. This system has no moving parts for nulling and does not exhibit azimuthal dependence on the optical elements. Therefore, this system is fast in data acquisition and easy to operate. In addition, image uniformity can be improved through a normalization process. For a demonstration, this system was applied to annealing studies of indium oxide thin films. The results show that this system can visualize the temperature dependence of the annealing as well as the progress of the annealing over time.

Super-contrast-enhanced darkfield imaging of nano objects through null ellipsometry.

We rediscover the null ellipsometry principle for an outstanding image-contrast enhancement method for darkfield imaging. Simply by adding polarizers, compensators, and a photodiode sensor to a conventional darkfield imaging system and applying the null principle, Si nano-cylinder structures as small as D20nm (H20nm) on non-patterned wafer, and gap defects as small as 14.6nm and bridge defects as small as 21.9nm on 40nm line and 40nm space patterns (H40nm), which are invisible in conventional darkfield imaging, can be distinguished from scattered noise. To the best of our knowledge, no method has been successful for identifying such small non-metal (silicon) nanoscale objects with such low magnification (×20) optics.

Avidin-Biotin Cross-Linked Microgel Multilayers as Carriers for Antimicrobial Peptides.

Herein, we report on the formation of cross-linked antimicrobial peptide-loaded microgel multilayers. Poly(ethyl acrylate- co-methacrylic acid) microgels were synthesized and functionalized with biotin to enable the formation of microgel multilayers cross-linked with avidin. Microgel functionalization and avidin cross-linking were verified with infrared spectroscopy, dynamic light scattering, and z-potential measurements, while multilayer formation (up to four layers) was studied with null ellipsometry and quartz crystal microbalance with dissipation (QCM-D). Incorporation of the antimicrobial peptide KYE28 (KYEITTIHNLFRKLTHRLFRRNFGYTLR) into the microgel multilayers was achieved either in one shot after multilayer formation or through addition after each microgel layer deposition. The latter was found to strongly promote peptide incorporation. Further, antimicrobial properties of the peptide-loaded microgel multilayers against Escherichia coli were investigated and compared to those of a peptide-loaded microgel monolayer. Results showed a more pronounced suppression in bacterial viability in suspension for the microgel multilayers. Correspondingly, LIVE/DEAD staining showed promoted disruption of adhered bacteria for the KYE28-loaded multilayers. Taken together, cross-linked microgel multilayers thus show promise as high load surface coatings for antimicrobial peptides.

Sensitivity and accuracy of new ellipsometric technique for the characterization of ultrathin films

In this study the possibility for the improvement of accuracy of single wavelength measurements organized by multiple angle of incidence (MAI) null ellipsometry technique is shown. The “transparent film on transparent substrate” system in a range of 1.0–20.0 nm film thickness in the low-contrast region on the film-substrate surface was studied. The developed method allows an independent determination of the thickness and refractive index of an ultrathin transparent film under conditions of a strong correlation effect between these parameters.

New Development of Membrane Base Optoelectronic Devices.

It is known that one factor that affects the operation of optoelectronic devices is the effective protection of the semiconductor materials against environmental conditions. The permeation of atmospheric oxygen and water molecules into the device structure induces degradation of the electrodes and the semiconductor. As a result, in this communication we report the fabrication of semiconductor membranes consisting of Magnesium Phthalocyanine-allene (MgPc-allene) particles dispersed in Nylon 11 films. These membranes combine polymer properties with organic semiconductors properties and also provide a barrier effect for the atmospheric gas molecules. They were prepared by high vacuum evaporation and followed by thermal relaxation technique. For the characterization of the obtained membranes, Fourier-transform infrared spectroscopy (FT-IR), scanning electron microscopy (SEM), and energy dispersive spectroscopy (EDS) were used to determine the chemical and microstructural properties. UV-ViS, null ellipsometry, and visible photoluminescence (PL) at room temperature were used to characterize the optoelectronic properties. These results were compared with those obtained for the organic semiconductors: MgPc-allene thin films. Additionally, semiconductor membranes devices have been prepared, and a study of the device electronic transport properties was conducted by measuring electrical current density-voltage (J-V) characteristics by four point probes with different wavelengths. The resistance properties against different environmental molecules are enhanced, maintaining their semiconductor functionality that makes them candidates for optoelectronic applications.

On the formation of inclusion complexes at the solid/liquid interface of anchored temperature-responsive PNIPAAM diblock copolymers with \u03b3-cyclodextrin

The thermal responsive behavior of adsorbed layers of diblock copolymers of poly(N-isopropylacrylamide) (PNIPAAM) and poly((3-acrylamidopropyl)trimethylammonium chloride) (PAMPTMA(+)) with γ-cyclodextrin (γ-CD) at the solid/liquid interface has been investigated using three in situ techniques: null ellipsometry, quartz–crystal microbalance with dissipation monitoring, and neutron reflectometry. The measurements provided information about the adsorbed amounts, the layer thickness, hydration and viscoelastic properties, and the interfacial structure and composition. The copolymers adsorb to silica with the cationic PAMPTMA(+) blocks sitting as anchors in a flat conformation and the PNIPAAM chains extending into the solution. The copolymer system alone exhibits reversible collapse above the lower critical solution temperature of PNIPAAM. The addition of γ-CD to pre-adsorbed copolymer layers results in a highly extended conformation as well as some loss of copolymer from the surface, which we discuss in terms of the formation of surface-invoked lateral steric repulsion of formed inclusion complexes.

Foundation of correlation ellipsometry.

An experimental strategy for the detection of fluctuation dynamics at interfaces based on a combination of photon correlation spectroscopy (PCS) with a nulling ellipsometry scheme is investigated theoretically. The intensity description of ellipsometry measurements is generalized to PCS time correlation functions. The nulling ellipsometry procedure is applied for every lag time t of the correlation functions, to extract the dynamics connected to the coherent signal which contains the interface dynamics. The classical ellipsometry parameters Δ and tanΨ are generalized to functions and tan [capital Psi, Greek, tilde]Q(t). A suitable Siegert relation is derived and employed to show that either field correlation functions or intensity correlation functions after baseline subtraction can be used as the starting point for the nulling ellipsometry procedure.

Nonlinear ellipsometry of Si(111) by second harmonic generation

In this paper we present experimental and simulated data of the second harmonic response for arbitrarily oriented linear input polarization and output polarization on Si(111) surfaces. In contrast to other nonlinear optical ellipsometry experiments (e.g. nonlinear optical null ellipsometry) we azimuthally rotate the sample for additional analysis of the crystal structure. In our study, we used the simplified bond hyperpolarizability model (SBHM) to simulate the observed angular shifts of the nonlinear peaks and the change of symmetry features related to a modification of the beam polarizations. Our findings help to identify the corresponding interface dipolar and bulk quadrupolar SHG sources.

Optical study of wedge-shaped films Part I Modeling

This paper, which is being published in two parts, provides a summary and analysis of the various effects that occur in ellipsometry and reflection spectroscopy studies of wedge-shaped thin films. In the first part of the paper, we calculate various optical characteristics of wedge-shaped films on substrates: the reflection coefficients and the spatial distribution of the light intensity entering a null-ellipsometer detector. We discuss whether existing theoretical approaches can be used in the study of wedge-shaped films and examine specific issues that arise when using null ellipsometry to determine the ellipsometric parameters (ψ and Δ) of such films. Part II of this paper will describe the results obtained through an experimental study of wedge-shaped films.

In Situ Potentiometry and Ellipsometry: A Promising Tool to Study Biofouling of Potentiometric Sensors.

In situ potentiometry and null ellipsometry was combined and used as a tool to follow the kinetics of biofouling of ion-selective electrodes (ISEs). The study was performed using custom-made solid-contact K(+)-ISEs consisting of a gold surface with immobilized 6-(ferrocenyl)hexanethiol as ion-to-electron transducer that was coated with a potassium-selective plasticized polymer membrane. The electrode potential and the ellipsometric signal (corresponding to the amount of adsorbed protein) were recorded simultaneously during adsorption of bovine serum albumin (BSA) at the surface of the K(+)-ISEs. This in situ method may become useful in developing sensors with minimized biofouling.

Adsorption of the intrinsically disordered saliva protein histatin 5 to silica surfaces. A Monte Carlo simulation and ellipsometry study

HypothesisThe adsorption of histatin 5 to hydrophilic silica surfaces is governed by electrostatic attractive forces between the positive protein and the negative surface. Hence pH and ionic strength control the adsorbed amount, which can be described by coarse-grained Monte Carlo simulations accounting for electrostatic forces and charge regulation of the protein. ExperimentsThe amount of histatin 5 adsorbed to hydrophilic silica surfaces at different pH and ionic strengths was measured using null ellipsometry. The results were compared with coarse-grained Monte Carlo simulations of a single histatin 5 molecule and a surface with a fixed, smeared charge set according to experimental values for silica. The Langmuir isotherm was used to calculate the surface coverage from the simulation results. The effect of charge regulation of the protein was investigated. FindingsEven though electrostatic attractive forces are important for the investigated system, a non-electrostatic short-ranged attraction with a strength of about 2.9kBT per amino acid was needed in the simulations to get surface coverages close to experimental values. The importance of electrostatics increases with increasing pH. Charge regulation of the protein affected the results from the simulations only at high surface charge and low ionic strength.

Ellipsometric study of molecular orientations of Thermomyces lanuginosus lipase at the air–water interface by simultaneous determination of refractive index and thickness

Ellipsometric studies of very thin organic films suffer from the low refractive index contrast between layer and bulk substrate. We demonstrate that null ellipsometry can not only provide detailed information about the adsorption kinetics and surface excess values, but in addition on layer thicknesses with submonolayer resolution of a lipase from Thermomyces lanuginosus at the air–water interface. While measuring very close to the Brewster angle, refractive indices and layer-thicknesses can both be determined with a precision that is sufficiently high to make conclusions on the density and orientation of the molecules at the interface. The orientation was found to be concentration- and pH value-dependent. At the isoelectric point, the lipase was almost vertically oriented with respect to the surface, while for pure distilled water and low lipase concentration a rather horizontal alignment was found. Further experiments, varying the size of the interfacial area in a Langmuir trough, confirm the different layer structures.

Ellipsometry method application in optics of inhomogeneous media

Physico-mathematical methods of modeling and measurements of polarization optical characteristics of “inhomogeneous surface layer – inhomogeneous medium” reflective system have been improved based on generalization of the polarized light reflection theory in the Born approximation from inhomogeneous media interface. Theoretical and experimental justification for simultaneous application of multi-angular and scanning ellipsometry methods is given for gradient optics elements in inhomogeneous layer presence on the surface of the optical detail. The measurement of the main ellipsometric parameters – linear reduced polarization azimuth Ψand phase difference Δbetween mutually orthogonal p– and s– components of a reflected polarized light beam – was carried out on a dual-channel installation, in which the scheme of measurements of polarization parameters Ψand Δby balanced (null) ellipsometry method is used in the first channel; the photometric scheme of measurements of parameters Ψand Δby Fourier method of output detection is used in the second channel. Such separation of channels gives the possibility for measuring of polarization parameters of reflected light beam separately and, thereby, improving the accuracy of ellipsometric measurements in relation to observable physical quantity of measurements object. The values of refractive index n(r) and absorption index k(r) were calculated by axial section of selfoc on the basis of measurements of polarization parameters Ψand Δwith angle of incidence of the light beam φ=45о within the limits of scanning ellipsometry method and a model of geometrically flat interface of homogeneous media. Empirical data indicated the model incorrectness of the geometrically flat interface for polarization measurements of refractive index distributionn(r) and absorption index k(r) in gradient optical engineering elements. For the reflective system “inhomogeneous surface layer – inhomogeneous medium” the refractive index n(r) change by axial section of an element r is proportional to phase difference Δ(r) between mutually orthogonal components of a reflected light beam. At the same time the linear reduced polarization azimuth Ψ(r) in the section gradient optical element varies slightly. The main proportions for calculation of refractive index nсand thickness dсof superficial layer in each local point r of selfoc are adduced on the basis of the ellipsometry equation, obtained within the framework of the polarized light reflection Born theory. Empirical data of experimental researches make it possible to give quantitative assessment of surface processing quality for optical engineering elements. Elaborated ellipsometric techniques are recommended in technological production of optical base assemblies with gradient and fiber optics elements.

Interfacial Behavior and Activity of Laccase and Bilirubin Oxidase on Bare Gold Surfaces

Two blue multicopper oxidases (MCOs) (viz. Trametes hirsuta laccase (ThLc) and Myrothecium verrucaria bilirubin oxidase (MvBOx)) were immobilized on bare polycrystalline gold (Au) surfaces by direct adsorption from both dilute and concentrated enzyme solutions. The adsorption was studied in situ by means of null ellipsometry. Moreover, both enzyme-modified and bare Au electrodes were investigated in detail by atomic force microscopy (AFM) as well as electrochemically. When adsorbed from dilute solutions (0.125 and 0.25 mg mL⁻¹ in the cases of ThLc and MvBOx, respectively), the amounts of enzyme per unit area were determined to be ca. 1.7 and 4.8 pmol cm⁻², whereas the protein film thicknesses were determined to be 29 and 30 Å for ThLc and MvBOx, respectively. A well-pronounced bioelectrocatalytic reduction of molecular oxygen (O₂) was observed on MvBOx/Au biocathodes, whereas this was not the case for ThLc-modified Au electrodes (i.e., adsorbed ThLc was catalytically inactive). The initially observed apparent k(cat)(app) values for adsorbed MvBOx and the enzyme in solution were found to be very close to each other (viz. 54 and 58 s⁻¹, respectively (pH 7.4, 25 °C)). However, after 3 h of operation of MvBOx/Au biocathodes, kcatapp dropped to 23 s⁻¹. On the basis of the experimental results, conformational changes of the enzymes (in all likelihood, their flattening on the Au surface) were suggested to explain the deactivation of MCOs on the bare Au electrodes.

Ellipsometry with polarisation analysis at cryogenic temperatures inside a vacuum chamber

In this paper we describe a new variant of null ellipsometry to determine thicknesses and optical properties of thin films on a substrate at cryogenic temperatures. In the PCSA arrangement of ellipsometry the polarizer and the compensator are placed before the substrate and the analyzer after it. Usually, in the null ellipsometry the polarizer and the analyzer are rotated to find the searched minimum in intensity. In our variant we rotate the polarizer and the compensator instead, both being placed in the incoming beam before the substrate. Therefore the polarisation analysis of the reflected beam can be realized by an analyzer at fixed orientation. We developed this method for investigations of thin cryogenic films inside a vacuum chamber where the analyzer and detector had to be placed inside the cold shield at a temperature of T ≈ 90 K close to the substrate. All other optical components were installed at the incoming beam line outside the vacuum chamber, including all components which need to be rotated during the measurements. Our null ellipsometry variant has been tested with condensed krypton films on a highly oriented pyrolytic graphite substrate (HOPG) at a temperature of T ≈ 25 K. We show that it is possible to determine the indices of refraction of condensed krypton and of the HOPG substrate as well as thickness of krypton films with reasonable accuracy.

Monitoring the microstructure of nanosized palladium layers obtained via thermal and VUV stimulated MOCVD

Nanosized palladium layers on silicon (100) substrates were obtained employing MOCVD method under VUV and thermal stimulations in hydrogen medium, with the use of Pd(hfaс)2 as a precursor. The deposition temperature under thermal stimulation amounted to 230°C, whereas with the VUV (145nm) stimulation this value was equal to 100°C, within the time interval ranging from 1 to 30min. With the use of monochromatic null ellipsometry and scanning electron microscopy techniques, initial stages of palladium nanolayer growth were studied. It has been established that the growth of Pd films exhibits a pronounced cyclical character. The analysis of electron microscopy images of growing film surfaces was based on the Voronoi–Delaunay method with the introduction of parameters such as peak, valley and minimum spanning tree (MST). It was demonstrated that the parameters used directly reflect the process of film growth, whereby two stages of the process have been distinguished.