The electrical double layer on gold probed by electrokinetic and surface force measurements

Gold surfaces, obtained by vacuum deposition of 15-nm gold films on glass and silica wafers, were studied in aqueous solutions by streaming potential measurements and colloidal-probe AFM force measurements. In the force measurements both a bare and a gold-coated silica particle (6 microm in diameter) have been used as colloidal probes. From the streaming potential measurements we determined the zeta-potential of the gold surface, while from the force measurements the diffuse double-layer potential psi(d) was obtained by fitting the data to the DLVO theory or to the nonlinear Poisson-Boltzmann equation. Measured interactions were found to be entirely due to overlap of electric double layers with no indication of attractive Van der Waals forces. Results of both types of measurements are in good agreement. The double layer potential strongly depends on the pH, probably as a result of the presence of oxide species on the gold surface. Insight in the double layer potential of polarizable interfaces such as the gold/electrolyte solution interface is the first step for understanding the effect of externally applied potentials on the adsorption behavior of charged species.     

Adhesive properties of Staphylococcus epidermidis probed by atomic force microscopy

Mapping of the surface properties of Staphylococcus epidermidis and of biofilm forming bacteria in general is a key to understand their functions, particularly their adhesive properties. To gain a comprehensive view of the structural and chemical properties of S. epidermidis, four different strains (biofilm positive and biofilm negative strains) were analyzed using in situ atomic force microscopy (AFM). Force measurements performed using bare hydrophilic silicon nitride tips disclosed similar adhesive properties for each strain. However, use of hydrophobic tips showed that hydrophobic forces are not the driving forces for adhesion of the four strains. Rather, the observation of sawtooth force-distance patterns on the surface of biofilm positive strains documents the presence of modular proteins such as Aap that may mediate cell adhesion. Treatment of two biofilm positive strains with two chemical inhibitor compounds leads to a loss of adhesion, suggesting that AFM could be a valuable tool to screen for anti-adhesion molecules.     

Infrared reflexion-absorption measurements on emersed gold electrodes

Infrared reflexion-absorption spectroscopy was performed on gold electrodes which had been emersed from aqueous potassium cyanate electrolytes at different potentials. New information was, thereby, obtained on the process of emersion and the electrosorption of cyanate on gold.The polycrystalline gold electrodes could be emersed without any adhering solution (“hydrophobic”) only when traces of organic substances were present in the electrolyte. On such emersed electrodes the potential-dependent adsorption of cyanate was observed by the asymmetric stretching vibration at 2185 cm^?1, and by hot bands. At the potential of anodic gold oxide formation the surface concentration of cyanate was found to drop to near zero.     

Isoelectric point of fluorite by direct force measurements using atomic force microscopy

Interaction forces between a fluorite (CaF2) surface and colloidal silica were measured by atomic force microscopy (AFM) in 1 x 10(-3) M NaNO3 at different pH values. Forces between the silica colloid and fluorite flat were measured at a range of pH values above the isoelectric point (IEP) of silica so that the forces were mainly controlled by the fluorite surface charge. In this way, the IEP of the fluorite surface was deduced from AFM force curves at pH approximately 9.2. Experimental force versus separation distance curves were in good agreement with theoretical predictions based on long-range electrostatic interactions, allowing the potential of the fluorite surface to be estimated from the experimental force curves. AFM-deduced surface potentials were generally lower than the published zeta potentials obtained from electrokinetic methods for powdered samples. Differences in methodology, orientation of the fluorite, surface carbonation, and equilibration time all could have contributed to this difference.     

Structural properties of oligonucleotide monolayers on gold surfaces probed by fluorescence investigations

We present optical investigations on the conformation of oligonucleotide layers on Au surfaces. Our studies concentrate on the effect of varying surface coverage densities on the structural properties of layers of 12- and 24mer single-stranded DNA, tethered to the Au surface at one end while being labeled with a fluorescent marker at the opposing end. The distance-dependent energy transfer from the marker dye to the metal surface, which causes quenching of the observed fluorescence, is used to provide information on the orientation of the DNA strands relative to the surface. Variations in the oligonucleotide coverage density, as determined from electrochemical quantification, over 2 orders of magnitude are achieved by employing different preparation conditions. The observed enhancement in fluorescence intensity with increasing DNA coverage can be related to a model involving mutual steric interactions of oligonucleotides on the surface, as well as fluorescence quenching theory. Finally, the applicability of the presented concepts for investigations of heterogeneous monolayers is demonstrated by means of studying the coadsorption of mercaptohexanol onto DNA-modified Au surfaces.     

Origin of the nonadhesive properties of fibrinogen matrices probed by force spectroscopy

The deposition of a multilayered fibrinogen matrix on various surfaces results in a dramatic reduction of integrin-mediated cell adhesion and outside-in signaling in platelets and leukocytes. The conversion of a highly adhesive, low-density fibrinogen substrate to the nonadhesive high-density fibrinogen matrix occurs within a very narrow range of fibrinogen coating concentrations. The molecular events responsible for this transition are not well understood. Herein, single-cell and molecular force spectroscopy were used to determine the early steps in the formation of nonadhesive fibrinogen substrates. We show that the adsorption of fibrinogen in the form of a molecular bilayer coincides with a several-fold reduction in the adhesion forces generated between the AFM tip and the substrate as well as between a cell and the substrate. The subsequent deposition of new layers at higher coating concentrations of fibrinogen results in a small additional decrease in adhesion forces. The poorly adhesive fibrinogen bilayer is more extensible under an applied tensile force than is the surface-bound fibrinogen monolayer. Following chemical cross-linking, the stabilized bilayer displays the mechanical and adhesive properties characteristic of a more adhesive fibrinogen monolayer. We propose that a greater compliance of the bi- and multilayer fibrinogen matrices has its origin in the interaction between the molecules forming the adjacent layers. Understanding the mechanical properties of nonadhesive fibrinogen matrices should be of importance in the therapeutic control of pathological thrombosis and in biomaterials science.     

Sensitive immunodetection through impedance measurements onto gold functionalized electrodes

This article deals with a direct electrochemical method of detecting antigens using new methods of functionalization of gold electrodes. Based on the reacting ability of gold with sulfhydryl groups, three protocols for the fixation of antibodies have been explored. They are based on either the self-assembling properties of functional thiols bearing long alkyl chains or the possibility of a direct coupling of antibody moieties. Coverage rates as high as 97% can be reached. The analysis of the electrochemical impedance behavior of such layers can lead to a sensitive method for the direct detection of the antibody/antigen interaction. The addition of a redox couple in the tested solution, acting as an amplifier, allowed detection limits for the antigens as low as a few picograms/milliliter to be reached.     

Effect of gold oxide in measurements of colloidal force

Atomic force microscopy, contact-angle, and spectroscopic ellipsometry measurements were employed to investigate the presence and properties of gold oxide on the surface of gold metal. It was found that, in agreement with available literature, unoxidized gold surfaces were hydrophobic, whereas oxidation rendered the surface highly hydrophilic. The oxide could be removed with ethanol or base but appeared to be stable over long periods in water or salt solutions between pH 3 and 7. After oxidation, the oxide layer thickness, determined using ellipsometry, was consistent with an approximate monolayer of Au-O bonds at the gold surface. The presence of gold oxide was found to alter significantly the electrical double-layer characteristics of the gold surface below pH 6 and may explain the apparent inconsistencies in observed force behavior where gold is employed as well as aiding in design of future microfluidic systems which incorporate gold as a coating.     

Determination of anisotropic surface characteristics of different phyllosilicates by direct force measurements

To fundamentally understand the electrokinetic behavior of clay minerals, it is necessary to study the anisotropic surface charge properties of clay surfaces. In this study, two 2:1 layer natural minerals, talc and muscovite, were chosen as representatives of magnesium and aluminum phyllosilicate minerals, respectively. The molecularly smooth basal planes of both platy minerals were obtained by cleavage along the basal planes, while suitable edge surfaces were prepared by an ultramicrotome cutting technique. Silicon nitride atomic force microscopy tip was used as a probe to study the interaction forces between the tip and clay basal/edge surfaces in aqueous solutions of various pH values. The measured interaction force profiles between the tip and clay basal/edge surfaces were fitted with the classical DLVO (Derjaguin-Landau-Verwey-Overbeek) theory, which allows direct determination of electrical surface potential of talc and muscovite surfaces. The surface potential of muscovite basal planes was found to be significantly more negative than the basal plane of talc, both being pH insensitive. In contrast, the surface potential of edge surfaces was highly pH-dependent, exhibiting a point of zero charge (PZC) at pH 7.5 and 8.1 for edges of muscovite and talc, respectively. The observed differences in surface potential of basal planes and edge surfaces for both talc and muscovite are closely related to their crystal structure and ionization characteristics. The protonation reactivity and the contribution of each surface group to the surface charging behavior are modeled using their protonation constants.     

Anisotropic orientation of horseradish peroxidase by reconstitution on a thiol-modified gold electrode

Horseradish peroxidase (HRP) was reconstituted on the surface of a gold electrode that was modified first with a hemin-carbon-chain-thiol derivative followed by addition of the apo protein to the contacting solution. To facilitate the reconstitution of the holo enzyme, the hemin needs to be immobilised on a carbon-chain spacer arm. To achieve this, an immobilisation protocol was developed that is based on the initial formation of a mixed self-assembled monolayer on the gold surface consisting of 3-carboxypropyl disulphide and an activated disulphide (3,3'-dithiodipropionic acid di-(N-succinimidyl ester)) followed by binding of a diaminoalkane to the activated disulphide. The hemin was then coupled to the second amino group of the diaminoalkane by means of a carbodiimide coupling reagent. Finally, the enzyme was reconstituted on the hemin-modified surface by immersion of the electrode in a solution containing apo-HRP. The advantage of this method is that the length of the spacer arm can be changed easily, because diaminoalkanes of different chain lengths are available. The electrochemistry of the hemin and the reconstituted HRP electrodes was studied by means of cyclic voltammetry and differential-pulse voltammetry. The catalytic ability for reduction of hydrogen peroxide was investigated for both direct and mediated electrochemistry with a soluble electron donor (ortho-phenylenediamine).     

Electrochemical properties of sulfur adsorbed on gold electrodes

The electrochemical properties of sulfur adsorbed on gold electrodes were studied in 10^?5M solutions of S^2? in 1 M NaOH. In general, ∵_S is less than a monolayer. At E=0.05 V only, a monolayer will be formed after long times. The sulfur layer is stable in the potential range between ?0.6 and +0.4 V. At lower potentials, sulfur can be desorbed cathodically (charge Q_red), but at higher potentials, where layers of gold oxide are formed, the sulfur is oxidized anodically (charge Q_ox). From the ratio Q_red·6/Q_ox=γ, the electrosorption valency γ=?2 is obtained. This means, that the sulfide ions are almost completely discharged during adsorption. The same layer can be formed by adsorption from polysulfide solutions, which can be explained by a break of the sulfur bond and adsorption of single sulfur atoms. The double layer capacity decreases during adsorption of sulfur indicating the formation of an insulating sulfur layer with a dielectric constant of about 2. The anodic adsorption of sulfide ions is limited by diffusion only. For longer polarisation times, the coverage is independent of time, i.e. place exchange reactions between Au and S can be excluded. The cathodic desorption as well as the anodic oxidation of the adsorbed sulfur are potential dependent charge transfer processes, as can be concluded from potentiodynamic measurements with various sweep rates.     

Electrocatalytic properties of Au(111)-Pd quasi-single-crystal film electrodes as probed by ATR-SEIRAS

Electrochemical and electrocatalytic properties of thin films Au(111-25 nm), which are quasi-single-crystal electrodes 25 nm thick made of gold with the (111) preferential orientation, and same electrodes modified with a monolayer (ML) of palladium are studied in 0.1 M solutions of HClO₄ and H₂SO₄ employing voltammetric techniques and surface enhanced infrared reflection absorption spectroscopy (ATR-SEIRAS). Spectroscopic experiments demonstrate strong adsorption of electrolyte species (H₂O, OH_ads, anions) on the Pd surface. The weak and reversible adsorption of CO on Au(111-25 nm) does not change the interfacial-water structure. Adsorption of CO on the Pd-modified film results in an irreversibly adsorbed CO adlayer stabilized by co-adsorbed isolated water species. Various electrooxidation mechanisms are discussed. Electrochemical and spectroscopic investigations on the adsorption and electrooxidation of HCOOH on bare and 1 ML Pd-Au(111-25 nm) electrodes reveal that electrooxidation proceeds in both cases via a direct or dehydrogenation pathway. This mechanism involves the formation of formate as intermediate, which is detected by in situ ATR-SEIRAS. The reactivity on Pd-modified surfaces is higher than on bare gold. The specifically adsorbed anions (sulfate/bisulfate) and the oxide formation on the substrate surface lower the reactivity for CO and HCOOH on both surfaces. Published in Russian in Elektrokhimiya, 2006, Vol. 42, No. 11, pp. 1312–1329. Based on the report delivered at the 8th International Frumkin Symposium “Kinetics of the Electrode Processes,” October 18–22, 2005, Moscow. The text was submitted by the authors in English.     

Combining electrochemistry and direct force measurements: from the control of surface properties towards applications

Direct force measurements contributed in the last years much to our understanding of the diffuse double layer of charged interfaces in electrolyte solutions. Such measurements have been performed with the atomic force microscope or the surface force apparatus. This review gives an overview over the recent studies based on force measurements with electrode surfaces. Not only bare metal electrodes but also electrodes modified by different organic layers, including electroactive films, have been studied by these techniques. Direct force measurements indicate that further effects besides classical Gouy–Chapman–Stern theory have to be taken into consideration in order to describe the force profiles. In addition to the long-range forces also the adhesion between surfaces can be tuned by potentiostatic control. New single-molecule techniques based on the atomic force microscope allow to probe the extension of polymer strands or their desorption from solid interfaces. In combination with electrochemistry, it became now possible to tune the desorption behavior of polymer strands or to measure the electromechanical coupling of motors from single strands of electroactive polymers.     

Probing surface charge potentials of clay basal planes and edges by direct force measurements

The dispersion and gelation of clay suspensions have major impact on a number of industries, such as ceramic and composite materials processing, paper making, cement production, and consumer product formulation. To fundamentally understand controlling mechanisms of clay dispersion and gelation, it is necessary to study anisotropic surface charge properties and colloidal interactions of clay particles. In this study, a colloidal probe technique was employed to study the interaction forces between a silica probe and clay basal plane/edge surfaces. A muscovite mica was used as a representative of 2:1 phyllosilicate clay minerals. The muscovite basal plane was prepared by cleavage, while the edge surface was obtained by a microtome cutting technique. Direct force measurements demonstrated the anisotropic surface charge properties of the basal plane and edge surface. For the basal plane, the long-range forces were monotonically repulsive within pH 6-10 and the measured forces were pH-independent, thereby confirming that clay basal planes have permanent surface charge from isomorphic substitution of lattice elements. The measured interaction forces were fitted well with the classical DLVO theory. The surface potentials of muscovite basal plane derived from the measured force profiles were in good agreement with those reported in the literature. In the case of edge surfaces, the measured forces were monotonically repulsive at pH 10, decreasing with pH, and changed to be attractive at pH 5.6, strongly suggesting that the charge on the clay edge surfaces is pH-dependent. The measured force profiles could not be reasonably fitted with the classical DLVO theory, even with very small surface potential values, unless the surface roughness was considered. The surface element integration (SEI) method was used to calculate the DLVO forces to account for the surface roughness. The surface potentials of the muscovite edges were derived by fitting the measured force profiles with the surface element integrated DLVO model. The point of zero charge of the muscovite edge surface was estimated to be pH 7-8.     

Configuration of triporphyrin ethers probed by fluorescence measurements.

The fluorescence spectra and lifetimes of triporphyrin ethers derived from hematoporphyrin, mesoporphyrin and protoporphyrin were examined, together with relative hydrophobicities estimated from reverse-phase high performance liquid chromatography (HPLC) elution times. The following data suggest a molecular arrangement with two of the three rings in a "folded configuration". The trimers display a greater fluorescence yield (phi f) than the corresponding diporphyrin ethers which contain only the two folded rings. The fluorescence lifetime data for the trimers are consistent with signals from both a folded ring pair (7-8 ns) and a free ring (14 ns). Reverse-phase HPLC studies indicate that the trimers are intermediate in hydrophobicity between the monomers and dimers. Preliminary data suggest that, for certain peripheral substitutions, the trimer configuration is superior to the dimer for photodynamic therapy.     

Surface ionization state and nanoscale chemical composition of UV-irradiated poly(dimethylsiloxane) probed by chemical force microscopy, force titration, and electrokinetic measurements

The surface chemistry and ionization state of cross-linked poly(dimethylsiloxane) (PDMS) exposed to UV/ozone were studied as a function of treatment time. Various complementary and independent experimental techniques were utilized, which yielded information on the macroscopic as well as the nanometric scale. The average chemical composition of the PDMS surface was quantitatively investigated by time-of-flight secondary ion mass spectrometry (ToF-SIMS). It was found that the top 1-2 nm surface layer was dominated by silanol groups (-SiOH) for which the concentration increased with increasing treatment dose. The lateral distributions of the silanol groups were analyzed on the nanometer scale by means of atomic force microscopy (AFM) with chemically functionalized tip probes in aqueous buffer solutions at varying pHs. Spatially dependent pull-off force curves (also called "force volume" imaging) indicated the presence of strong chemical heterogeneity of the probed surface. This heterogeneity took the form of patches of silanol functionalities with high local concentration surrounded by a matrix of predominantly hydrophobic domains at low pH. The average pull-off forces for the entire surface scanned were significantly reduced for pH values larger than a characteristic pK(a) constant (in the range between 4.5 and 5.5). The extent of the decrease in the pull-off force and the particular value of pK(a) were found to be a function of treatment time and to differ from the commonly reported values for silanol functional groups on a homogeneous silica surface. These dependences were ascribed to the evoking of a protonation/deprotonation process of the surface silanol groups which was sensitive to the hydrophobic/hydrophilic balance of their close molecular environment. Intermolecular hydrogen bonding may also account for the shifts in the surface pK(a). Furthermore, depending on the nature of the electrolyte, a third effect related to double layer composition, as determined by specific ion adsorption, was quantitatively analyzed by streaming potential measurements in the presence of sodium chloride and phosphate electrolytes.     

Adsorption of 4-aminopyridine on Co and Ag electrodes probed by SERS

The adsorption of 4-aminopyridine (4-AP) on Co and Ag electrodes in acid or alkaline solutions of KCl and KI electrolyte salts were monitored by the Surface-enhanced Raman Spectroscopy (SERS) technique. The SERS intensity for the Ag electrode was in 2 orders of magnitude higher than for the Co electrode, due to the enhancement of the Raman cross-section on Ag by the surface-plasmon excitation. In acidic chloride medium (pH 4), the SERS results for Ag electrodes indicate that the protonated form of 4-AP (4-APH⁺) adsorbs in the potential range of −0.1 to −0.6 V (Ag|AgCl|KCl sat) through hydrogen-bonding between 4-APH⁺ and Cl⁻ adsorbed on the electrode surface; at more negative potentials the neutral form 4-AP is the predominant adsorbed species. For Co electrode in the same medium, only bands due to neutral 4-AP were observed in the spectra at −0.8 and −0.9 V. For more negative potentials bands assigned to both 4-AP and 4-AP surface complex are observed, with the lasts being enhanced, as the potentials are turned more negative. In alkaline chloride medium (pH 13), for less negative potentials the bands assigned to free 4-AP were observed in the spectra of both Ag and Co surfaces. For more negative potentials, only bands assigned to the 4-AP surface complex were observed. For 0.1 mol L⁻¹ KI acidic or alkaline solutions, bands assigned to 4-AP and 4-APH⁺ were observed in a wider potential range than in chloride solutions. An adsorption scheme of 4-AP on Ag and Co is proposed for acidic and alkaline solutions.     

High-affinity tags fused to s-layer proteins probed by atomic force microscopy

Two-dimensional, crystalline bacterial cell surface layers, termed S-layers, are one of the most commonly observed cell surface structures of prokaryotic organisms. In the present study, genetically modified S-layer protein SbpA of Bacillus sphaericus CCM 2177 carrying the short affinity peptide Strep-tag I or Strep-tag II at the C terminus was used to generate a 2D crystalline monomolecular protein lattice on a silicon surface. Because of the genetic modification, the 2D crystals were addressable via Strep-tag through streptavidin molecules. Atomic force microscopy (AFM) was used to investigate the topography of the single-molecules array and the functionality of the fused Strep-tags. In high-resolution imaging under near-physiological conditions, structural details such as protein alignment and spacing were resolved. By applying molecular recognition force microscopy, the Strep-tag moieties were proven to be fully functional and accessible. For this purpose, streptavidin molecules were tethered to AFM tips via approximately 8-nm-long flexible polyethylene glycol (PEG) linkers. These functionalized tips showed specific interactions with 2D protein crystals containing either the Strep-tag I or Strep-tag II, with similar energetic and kinetic behavior in both cases.