Bare Gold Surface Research Articles

Activity of lactoperoxidase when adsorbed on protein layers

Lactoperoxidase (LPO) is an enzyme, which is used as an antimicrobial agent in a number of applications, e.g., food technology. In the majority of applications LPO is added to a homogeneous product phase or immobilised on product surface. In the latter case, however, the measurements of LPO activity are seldom reported. In this paper we have assessed LPO enzymatic activity on bare and protein modified gold surfaces by means of electrochemistry. It was found that LPO rapidly adsorbs to bare gold surfaces resulting in an amount of LPO adsorbed of 2.9 mg/m 2. A lower amount of adsorbed LPO is obtained if the gold surface is exposed to bovine serum albumin, bovine or human mucin prior to LPO adsorption. The enzymatic activity of the adsorbed enzyme is in general preserved at the experimental conditions and varies only moderately when comparing bare gold and gold surface pretreated with the selected proteins. The measurement of LPO specific activity, however, indicate that it is about 1.5 times higher if LPO is adsorbed on gold surfaces containing a small amount of preadsorbed mucin in comparison to the LPO directly adsorbed on bare gold.

Effective CO Oxidation on Endohedral Gold-Cage Nanoclusters

Ab initio calculations are performed to study the CO oxidation on six endohedral gold-cage clusters (W@Au12, Nb@Au13, Zr@Au14, Sc@Au15, Ca@Au16, and Na@Au17). The calculations suggest that three clusters (Nb@Au13, Zr@Au14, and Sc@Au15) can be very effective nanocatalysts. The reaction energy barriers are lower than those associated with either bare gold clusters or gold surfaces and are comparable to those associated with the support Au clusters. The effective CO oxidation can be attributed to the charge transfer from the dopant to the gold cage, the low coordination number for gold atoms on the cages, as well as the fluxionality of the cage.

Dielectric-loaded surface plasmon-polariton waveguides at telecommunication wavelengths: Excitation and characterization

The excitation and propagation of strongly confined surface plasmon-polariton (SPP) waveguide modes, supported by 500-nm-wide and 550-nm-high dielectric ridges fabricated on smooth gold films, are investigated at telecommunication wavelengths using a scanning near-field optical microscope. Different tapering structures for coupling of SPPs, excited at bare gold surfaces, into dielectric-loaded SPP waveguide (DLSPPW) modes are considered. The DLSPPW mode confinement and propagation loss are characterized. The DLSPPW mode propagation along an S bend having the smallest curvature radius of 2.48μm is shown, demonstrating the potential of DLSPPW technology for the realization of high-density photonic integrated circuits.

Quartz crystal microbalance study of the interfacial nanobubbles

The applications of quartz crystal microbalance (QCM) to the study of surface and interfacial science such as adsorption have become progressively common and popular these days. In this work, QCM with dissipation monitoring was used to study the formation of nanobubbles on bare and thiol-coated gold surfaces. The nanobubbles were produced using one of the established solvent exchange protocols and the formation was first confirmed by the tapping mode atomic force microscopy (AFM). By QCM measurements, we found that the formation of nanobubbles on the hydrophobic crystal surfaces can yield easily detectable shifts of frequency and dissipation from those measured directly in water without the presence of nanobubbles. The direction of the shifts is consistent with the depletion of water by gases of lower density. We also found that the formation of nanobubbles is a fast process and can be finished in less than a minute. The response of QCM at several overtones showed that nanobubbles cannot be used to explain why the shift in the half bandwidth is sometimes smaller than the negative frequency shift at higher overtones when the QCM crystal is operated in liquids.

Potential-Induced Structure Changes of Oligopyridine Adlayers on Au(111) Electrodes

The structure of a bisterpyridine-like oligopyridine (abbreviated as 2,4'-BTP) monolayer on Au(111), adsorbed from an acetone solution, was studied by in situ scanning tunneling microscopy and cyclic voltammetry in aqueous 0.1 M H2SO4. Short-range ordered adsorption with an average distance between the individual molecules of about 2 nm was observed only at electrode potentials positive of +0.4 V vs SCE, whereas at more negative potentials, no order could be found. With the help of Cu underpotential deposition, a potential-induced, fast, and fully reversible structure transition within the organic monolayer was identified at about +0.4 V vs SCE. At negative potentials the molecules apparently cluster together and consequently current-potential curves resemble those for a bare gold surface, whereas for E>+0.4 V vs SCE the molecules are spread over the entire surface in a hexagonal, close-packed fashion. This may have interesting consequences for switching between different template structures.

Nonspecific Protein Adsorption at the Single Molecule Level Studied by Atomic Force Microscopy

Liquid tapping atomic force microscopy was used to study the nonspecific adsorption of horse spleen ferritin at a bare gold surface at single molecule resolution. The majority of ferritin molecules adsorbed irreversible on gold surfaces in accordance with the random sequential adsorption (RSA) mechanism frequently used to describe irreversible adsorption processes. However, the time-resolved data also reveal events that go beyond the RSA model, i.e., lateral mobility and fragility of some molecules, resulting in desorption, chain formation, and subunit dissociation. Scanning effects of the AFM tip were observed, resulting in diminished protein coverage in the scanned area.

Inhibition of Escherichia coli Biofilm Formation by Self-Assembled Monolayers of Functional Alkanethiols on Gold

Bacterial biofilms cause serious problems, such as antibiotic resistance and medical device-related infections. To further understand bacterium-surface interactions and to develop efficient control strategies, self-assembled monolayers (SAMs) of alkanethiols presenting different functional groups on gold films were analyzed to determine their resistance to biofilm formation. Escherichia coli was labeled with green florescence protein, and its biofilm formation on SAM-modified surfaces was monitored by confocal laser scanning microscopy. The three-dimensional structures of biofilms were analyzed with the COMSTAT software to obtain information about biofilm thickness and surface coverage. SAMs presenting methyl, L-gulonamide (a sugar alcohol tethered with an amide bond), and tri(ethylene glycol) (TEG) groups were tested. Among these, the TEG-terminated SAM was the most resistant to E. coli biofilm formation; e.g., it repressed biofilm formation by E. coli DH5alpha by 99.5% +/- 0.1% for 1 day compared to the biofilm formation on a bare gold surface. When surfaces were patterned with regions consisting of methyl-terminated SAMs surrounded by TEG-terminated SAMs, E. coli formed biofilms only on methyl-terminated patterns. Addition of TEG as a free molecule to growth medium at concentrations of 0.1 and 1.0% also inhibited biofilm formation, while TEG at concentrations up to 1.5% did not have any noticeable effects on cell growth. The results of this study suggest that the reduction in biofilm formation on surfaces modified with TEG-terminated SAMs is a result of multiple factors, including the solvent structure at the interface, the chemorepellent nature of TEG, and the inhibitory effect of TEG on cell motility.

Adsorption Behavior of DNA-Wrapped Carbon Nanotubes on Self-Assembled Monolayer Surfaces

We have examined the adsorption of DNA-wrapped single-walled carbon nanotubes (DNA-SWNTs) on hydrophobic, hydrophilic, and charged surfaces of alkylthiol self-assembled monolayers (SAMs) on gold. Our goal is to understand how DNA-SWNTs interact with surfaces of varying chemical functionality. These samples were characterized using reflection absorption FTIR (RAIRS), X-ray photoelectron spectroscopy (XPS), and Raman spectroscopy. We have found that DNA-SWNTs preferentially adsorb to positively charged amine-terminated SAMs and to bare gold surfaces versus hydrophobic methyl-terminated or negatively charged carboxylic acid-terminated SAMs. Examination of the adsorption on gold of single-strand DNA (ssDNA) of the same sequence used to wrap the SWNTs suggests that the DNA wrapping plays a role in the adsorption behavior of DNA-SWNTs.

Adsorbed Gels versus Brushes: Viscoelastic Differences

It is of fundamental importance to be able to easily distinguish between the viscoelastic properties of a molecular gel (noncovalent cross-linked three-dimensional polymer structure) and a brush (polymer structure that emanates from a surface in three dimensions without cross-linking). This has relevance in biology and in designing surfaces with desired chemical and viscoelastic properties for nano and genomic technology applications. Agarose and thiol-tagged poly(ethylene glycol) were chosen as model systems, as they are known, on adsorption, to behave like a molecular gel and brush, respectively. Here, we focus on their viscoelastic differences using a quartz crystal microbalance with dissipation monitoring (QCM-D). Changes in resonance frequency and dissipation for three overtones using QCM-D were fitted with the Voigt viscoelastic model to calculate the shear viscosity and shear modulus for the adsorbed agarose gel and the PEG brush. At a surface coverage of 500 ng/cm2, the shear viscosities and shear moduli were 0.0025 +/- 0.0002 Pa-s and 2.0 +/- 0.17 x 105 Pa and 0.0010 +/- 0.0001 Pa-s and 5.0 +/- 0.3 x 104 Pa for the gel and brush, respectively. Thus, the adsorbed agarose gel layer was far more rigid than that of the covalently bound PEG brush due to its cross-linked network. Also, the diffusivity of agarose and PEG in solution was compared during adsorption onto a bare gold surface. The estimated value for the effective diffusivity of the PEG (without a thiol tag) and of the agarose gel was on the order of 10(-11) and 10(-15) m2/s, respectively. This low diffusivity for agarose supports the contention that it exists as a molecular gel with a H-bonded cross-linked network in aqueous solution. With the methods used here, it is relatively easy to distinguish the differences in viscoelastic properties between an adsorbed gel and brush.

Protein Nanopatterns and Biosensors Using Gold Binding Polypeptide as a Fusion Partner

An efficient strategy for immobilizing proteins on a gold surface was developed by employing the gold binding polypeptide (GBP) as a fusion partner. Using the enhanced green fluorescent protein (EGFP), severe acute respiratory syndrome coronavirus (SARS-CoV) envelope protein (SCVme), and core streptavidin (cSA) of Streptomyces avidinii as model proteins, specific immobilization of the GBP-fusion proteins onto the gold nanoparticles and generation of protein nanopatterns on the bare gold surface were demonstrated. The GBP-fused SCVme bound to gold nanoparticles successfully interacted with its antibody and showed changes in absorbance and color, allowing efficient diagnosis of SARS-CoV. The fusion proteins could be successfully immobilized on the gold surface by nanopatterning and microcontact printing as examined by atomic force microscopy and surface plasmon resonance analysis. The poly(dimethylsiloxane) microfluidic channels were created on the gold surface and were used for antigen-antibody and DNA-DNA interaction studies. Specific immobilization of GBP-EGFP fusion protein and its interaction with the antibody in the microchannels could be demonstrated. By immobilizing the DNA probe through the use of GBP-fused cSA, specific hybridization of the target DNA prepared from Salmonella could also be achieved. The GBP-fusion method allows immobilization of proteins onto the gold surface without surface modification and in bioactive forms suitable for studying protein-protein, DNA-DNA, and other biomolecular interaction studies. Furthermore, these studies can be carried out in a microfluidic system, which allows high-throughput analysis of biomolecular interactions.

Immunosensor with a controlled orientation of antibodies by using NeutrAvidin–protein A complex at immunoaffinity layer

The orientation of antibody was controlled by using NeutrAvidin-protein A complex on the gold surface of SPR biosensor. The surface density of receptor antibody (anti-hIgG) was compared by treatment of receptor antibody to the layer of avidin, NeutrAvidin, protein A, NeutrAvidin-protein A complex and bare gold surface of SPR biosensor. The ligand antibody (hIgG) was injected to each IA layer and the binding ratio of ligand antibody per unit receptor was estimated as a parameter of orientation control. The NeutrAvidin-protein A complex on gold surface of SPR biosensor showed the highest surface density of receptor antibody as well as the binding ratio of ligand antibody per receptor antibody. The NeutrAvidin-protein A complex was also prepared on biotin-labelled SAM, and the binding ratio of ligand per receptor was found to be significantly improved in comparison to the IA layer prepared by chemical coupling of receptor antibody to the SAM layer. The NeutrAvidin-protein A complex which showed the highest efficiency for the binding of ligand antibodies, was applied for the detection of a cancer marker called CEA. By using NeutrAvidin-protein A complex and sandwich assay for signal amplification, sensitivity was improved to be 1.5-fold higher than bare gold surface and the detection of CEA with the detection limit of 30 ng/ml was achieved.

Motion of redox molecules in solution monitored by the highly-sensitive EQCM technique

The behavior of redox molecules in solution that was not detected by electrochemical techniques was measured by a highly-sensitive electrochemical quartz crystal microbalance (EQCM) technique that has been improved in this study to obtain a high sensitivity of EQCM measurement in solution. The improved EQCM technique allowed to monitor the motion of a redox molecule, that is an access of the molecule to an electrode surface and repulsion from the surface during redox. An EQCM technique currently in use has measured adsorption of redox molecules on an electrode surface or polymerization on the surface caused by a chemical reaction following redox, which exhibits an enough large mass change response to detect with an EQCM measurement. However, access and repulsion of redox molecule, which is a slight motion of the molecule near on electrode surface, has not been detected and investigated by an EQCM technique, because the mass change response seems to be very small. In this study, the redox behavior of methyl viologen on a bare gold surface, pyridinethiol surface and methylpyridinethiol surface was investigated. Although the three electrodes give the same cyclic voltammogram of methyl viologen, the three are different in QCM response recorded at the same time as the voltammetry. Access/repulsion of methyl viologen within an electrical double layer was monitored by the highly-sensitive EQCM technique.

Nanosecond Imaging of Microboiling Behavior on Pulsed-Heated Au Films Modified with Hydrophilic and Hydrophobic Self-Assembled Monolayers

Fast transient microboiling has been characterized on modified gold microheaters using a novel laser strobe microscopy technique. Microheater surfaces of different hydrophobicity were prepared using self-assembled monolayers of hexadecane thiol (hydrophobic) and 16-mercaptohexadecanol (hydrophilic) as well as the naturally hydrophilic bare gold surface. The microheater was immersed in a pool of water, and a 5-micros voltage pulse to the heater was applied, causing superheating of the water and nucleation of a vapor bubble on the heater surface. Light from a pulsed Nd:Yag laser was configured to illuminate and image the sample through a microscope assembly. The timing of the short duration (7.5 ns) laser flash was varied with respect to the voltage pulse applied to the heater to create a series of images illuminated by the flash of the laser. These images were correlated with the transient resistance change of the heater both during and after the voltage pulse. It was found that hydrophobic surfaces produced a bubble that nucleated at an earlier time, grew more slowly to a smaller maximum size, and collapsed more rapidly than bubbles formed on hydrophilic surfaces.

Mapping molecular orientation of pentacene on patterned Au surface

With a photoemission electron microscope, we investigated the orientation of pentacene films deposited on Au surface that has been patterned with self-assembled monolayers (SAMs). Organic thiols X(CH 2) nSH with X = COOH or CH 3 and an aromatic thiol; served as alignment layers to control the orientation of pentacene molecules thermally deposited on top of the SAM. According to the PEEM images and near-edge X-ray absorption fine structure spectra, a SAM exposing a non-polar methyl group, a terphenyl group or a polar carboxyl group all induced perpendicular alignment of the pentacene moiety, in contrast with parallel alignment on a bare gold surface.

Impedance Sensing of DNA Binding Drugs Using Gold Substrates Modified with Gold Nanoparticles

Interfacial interactions between immobilized DNA probes and DNA-specific sequence binding drugs were investigated using impedance spectroscopy toward the development of a novel biosensing scheme. The impedance measurements are based on the charge-transfer kinetics of the [Fe(CN)6]3-/4- redox couple. Compared to bare gold surfaces, the immobilization of DNA and then the DNA-drug interaction on electrode surfaces altered the capacitance and the interfacial electron resistance and thus diminished the charge-transfer kinetics by reducing the active area of the electrode or by preventing the redox species from approaching the electrode. Electrochemical deposition of gold nanoparticles on a gold electrode surface showed significant improvement in sensitivity. DNA-capped gold nanoparticles on electrodes act as selective sensing interfaces with tunable sensitivity due to higher amounts of DNA probes and the concentric orientation of the DNA self-assembled monolayer. The specificity of the interactions of two classical minor groove binders, mythramycin, a G-C specific-DNA binding anticancer drug, netropsin, an A-T specific-DNA binding drug and an intercalator, nogalamycin on AT-rich DNA-modified substrate and GC-rich DNA-modified substrate are compared. Using gold nanoparticle-deposited substrates, impedance spectroscopy resulted in a 20-40-fold increase in the detection limit. Arrays of deposited gold nanoparticles on gold electrodes offered a convenient tool to subtly control probe immobilization to ensure suitably adsorbed DNA orientation and accessibility of other binding molecules.

Azurin Self-Assembled Monolayers Characterized by Coupling Electrical Impedance Spectroscopy and Spectroscopic Ellipsometry

We present a combined study of azurin monolayers deposited on both bare and hexadecanethiol-covered gold surfaces carried out by coupling two independent techniques, in situ electrical impedance spectroscopy (EIS) and ex situ spectroscopic ellipsometry (SE). The combination of the two techniques which probe different sample properties in different environmental conditions results in a cross-consistent characterization of the optical and electrical properties of the protein layers. Moreover, it provides a reliable estimate of the layer thickness. Azurin self-assembles on bare gold forming a monolayer with a thickness of 3.2 ± 0.2 nm, a value very close to the crystallographic height of the cylindrically shaped protein. When azurin is deposited on hexadecanethiol-covered gold, a protein layer thickness of 2.7 ± 0.2 nm is obtained. This indicates that azurin forms a less dense layer when it is deposited on thiol-covered gold compared to the adsorption on bare gold. Both EIS and SE provide a direct estimate of the dielectric constant of the protein layer in the hydrated and dry states, respectively. In the case of the composite azurin/hexadecanethiol system, the combined EIS and SE analysis proves to be a valuable tool for the correct interpretation of the EIS data and allows for an estimate of the azurin and hexadecanethiol conductivity ratio.

Antibody immobilisation on the metal and silicon surfaces. The use of self-assembled layers and specific receptors

The use of Staphylococcal protein A and lectins as intermediate immobilising agents allows operators to orient antibodies (Ab) towards the solution due to the presence of a specific binding sites of immunoglobulin (Ig) molecules. Antibodies of different species of animals have unequal affinities to individual lectins. The effective thickness of immobilised Ab's depends on the type of substrates used and increases in the following sequence: bare gold or silicon surface, the surface treated with self-assembled polyelectrolytes (PESA) or with protein A or some lectins deposited on the preliminary formed polyelectrolyte layer. The glycolysated protein of jp51 may be selectively immobilised from the mixture of retroviral proteins (p24 and jp51), if it is necessary to distinguish infected animals from preliminarily immunised ones by means of a vaccine based on p24 protein. It was shown that the use of Staphylococcal protein A, instead of some lectins as intermediate layer for the Ab immobilisation, does not lead to a more sensitive determination of such low-weight toxins as 2,4-dichlorophenoxyacetic acid (2,4-D). The above-mentioned results were obtained with surface plasmon resonance (SPR) technique.

Adsorption of enantiomeric poly(lactide)s on surface-grafted poly(L-lactide).

We show that resistance of densely grafted polymer layers to adsorption of chemically identical free chains, which is known to be caused by entropic expulsion of free chains from the grafted layer, can be suppressed using the grafted and free chains of opposite stereoconfiguration. Specifically, we study adsorption of poly(L-lactide) (PLLA) and its enantiomer poly(D-lactide) (PDLA) onto layers of surface-grafted PLLA in acetonitrile and chloroform by infrared spectroscopy (IR). The grafted layers with thicknesses ranging from 7 to 35 nm are produced by ring-opening polymerization of L-lactide from hydroxyl end-groups of a self-assembled monolayer on gold. The IR data indicate that adsorption on the bare gold surface is the same for the L- and D-form of the polymer. However, covering the gold with the surface-grafted PLLA produces a significant decline in the adsorption of free PLLA and, by contrast, a strong enhancement in the adsorption of free PDLA. In addition, the IR data indicate that the adsorbed PDLA chains are stereocomplexed with the grafted PLLA chains. Thus, entropic expulsion of free chains from the grafted layer, which is responsible for the resistance of surface-grafted PLLA to adsorption of free PLLA, is suppressed in the case of free PDLA by stereocomplexation between the grafted and free chains.

Voltammetric and in situ FTIRS study on CN − and Au(CN) −x complexes at the polycrystalline gold surface in citrate medium

The adsorption and reactivity of citrate anions, cyanide anions, and the Au(I) and Au(III) cyanocomplexes (KAu(CN) 2 and KAu(CN) 4, respectively) on gold polycrystalline electrodes have been investigated by means of cyclic voltammetry and in situ FTIR spectroscopy in pH 7 sodium citrate medium. It was found that citrate anion adsorbs reversibly through the carboxylate groups from 0.4 V (RHE) on bare gold surfaces. However, the onset of this adsorption process is shifted by 600 mV in the positive direction when adsorbed cyanide species are present at the electrode surface. The Au(I) cyanocomplex produces a film on the gold electrode that inhibits the oxidation of both citrate anions and the gold surface. The reduction of the Au(III) cyanocomplex gives rise to the formation of the Au(I) cyanocomplex at potentials less negative than those required for gold deposition. Upon reduction, both cyanocomplexes release adsorbed cyanide on the gold electrode, which can be subsequently oxidized to cyanate at higher potentials.

Controlled deposition of single DNA molecules on bare gold electrodes

We report a new approach to a directly controlled deposition of long deoxyribonucleic acid (DNA) molecules on a bare gold electrode array predefined on a positively charged substrate surface by electrostatic interaction without modification of gold electrodes and DNA molecules. The method involves depositing amino-terminated self-assembled monolayer on a specific region on a SiO 2/Si substrate, and subsequently defining gold electrode array on the modified region by mask pattern transfer technique, as well as transferring a droplet of DNA solution onto the patterned substrate. Atomic force microscopy observations demonstrate that DNA molecules are significantly less stable on the bare gold surface than that on the aminothiolated gold surface. However, DNA can stably immobilize on the bare gold electrode array predefined on the positively charged substrate surface. The noncovalent binding of DNA onto the bare gold electrodes was shown to be robust enough to withstand ultrasonically washing. The approach represents a simple route to positioning of single DNA molecules on predefined structures, and can be potentially applied to other charged one-dimensional nanostructures.