Scanning electrochemical microscopy. 46. Shielding effects on reversible and quasireversible reactions

An approximate theory for the feedback mode of the scanning electrochemical microscope (SECM) is developed to interpret the effects of substrate shielding on an ultramicroelectrode tip during a recording of iT versus d curves (approach curves) for reversible and quasireversible kinetics at a substrate surface. The resulting expressions for the tip current, iT, show a good fit to more accurate SECM simulations as well as to the experimental response of a reversible and quasireversible reaction. SECM shielding experiments thus give an interesting new insight into SECM approach curves over electrodes at different potentials, which suggest possible applications to measuring heterogeneous kinetics for fast reactions and diffusion coefficient determination.     

Scanning electrochemical microscopy. 55. Fabrication and characterization of micropipet probes

The fabrication and characterization of novel micropipet probes for use in scanning electrochemical microscopy (SECM) are described. These can be used to dispense small (pL) amounts of a solution while monitoring the electrochemical response at a substrate and at a ring electrode tip on the micropipet probe. The probes were constructed by insulating gold-coated borosilicate micropipets with electrophoretic paint and exposing a ring electrode at the tip by heat treatment. Characterization of the probes was performed using scanning electron microscopy, cyclic voltammetry, and SECM approach curve experiments. Routine construction of tips with diameters of the order of 3 microm was possible using this technique. The probes exhibited stable steady-state currents and positive and negative feedback approach curves that agreed with those predicted by theory. Demonstrative SECM imaging experiments were performed using a picodispenser to continuously dispense an electroactive solution (ferrocenemethanol) to the SECM cell while the probe was located within a few micrometers of a Pt substrate surface. Oxidation of the dispensed electroactive solution was performed at the substrate, and feedback currents were measured at the probe tip by holding the gold ring at a reducing potential. This mode of tip-dispensing SECM was used to obtain images of a platinum substrate electrode while monitoring both the substrate current and the feedback current at the probe.     

Fabrication and characterization of probes for combined scanning electrochemical/optical microscopy experiments

A technique that combines scanning electrochemical microscopy (SECM) and optical microscopy (OM) was implemented with a new probe tip. The tip for scanning electrochemicaVoptical microscopy (SECM/OM) was constructed by insulating a typical gold-coated near-field scanning optical microscopy tip using electrophoretic anodic paint. Once fabricated, the tip was characterized by steady-state cyclic voltammetry, as well as optical and electrochemical approach experiments. This tip generated a stable steady-state current and well-defined SECM approach curves for both conductive and insulating substrates. Durable tips whose geometry was a ring with < 1 microm as outer ring radius could be consistently fabricated. Simultaneous electrochemical and optical images of an interdigitated array electrode were obtained with a resolution on the micrometer scale, demonstrating good performance of the tip as both an optical and an electrochemical probe for imaging microstructures. The SECM feedback current measurements were successfully employed to determine tip-substrate distances for imaging.     

Microfluidic push-pull probe for scanning electrochemical microscopy

This paper presents a microfluidic push-pull probe for scanning electrochemical microscopy (SECM) consisting of a working microelectrode, an integrated counter/reference electrode and two microchannels for pushing and pulling an electrolyte solution to and away from a substrate. With such a configuration, a droplet of a permanently renewed redox mediator solution is maintained just at the probe tip to carry out SECM measurements on initially dry substrates or in microenvironments. For SECM imaging purposes, the probe fabricated in a soft polymer material is used in a contact regime. SECM images of various gold-on-glass samples demonstrate the proof-of-concept of a push-pull probe for local surface activity characterization with high spatial resolution even on vertically oriented substrates. Finite element computations were performed to guide the improvement of the probe sensitivity.     

Scanning electrochemical microscopy 50. Kinetic study of electrode reactions by the tip generation-substrate collection mode

A scanning electrochemical microscopy (SECM) methodology for localized quantitative kinetic studies of electrode reactions based on the tip generation-substrate collection (TG-SC) operation mode is presented. This approach does not use the mediator feedback required in typical kinetic SECM experiments. The reactant is galvanostatically electrogenerated on a tip placed in proximity to the substrate. It diffuses through the tip-substrate gap and undergoes the reaction of interest on the substrate surface. The substrate current is monitored with time until it reaches an apparent steady-state value. The process was digitally simulated using an explicit finite difference method, for an irreversible first-order electrode reaction at the substrate. Transient responses, steady-state polarization curves, and TG-SC approach curves can be used to obtain substrate kinetics. The effects of the experimental parameters were analyzed. The possibility of easily changing the experimental conditions with the SECM is an attractive approach to obtain independent evidence that can be used for a strict test of reaction mechanisms. The technique was applied for a preliminary simplified kinetic examination of the oxygen reduction reaction in phosphoric acid.     

Scanning electrochemical microscopy of living cells. 4. Mechanistic study of charge transfer reactions in human breast cells

Scanning electrochemical microscopy (SECM) has recently been employed for probing the redox properties of individual mammalian cells. It was shown that intracellular redox activity can be probed noninvasively by measuring the rate of mediator regeneration by the cell. Depending on the properties of the mediator species (e.g., formal potential, ionic charge, and hydrophobicity), different steps can limit the rate of the mediator regeneration reaction. This paper describes the evaluation of several factors that determine the rates of different steps of the process. These include intracellular concentration of redox centers, mixed redox potential inside the cell, and the rate of membrane permeation by mediator species. The kinetic analysis has been carried out to clarify the origins of different rates of the overall charge-transfer reaction in different cell types and with different redox mediators. The results can be used to facilitate differentiation between different types of cells, for example, normal and metastatic breast cells, on the basis of differences in redox responses.     

Use of catechol as selective redox mediator in scanning electrochemical microscopy investigations

The use of catechols, and more specifically of dopamine, as a specific redox mediator for scanning electrochemical microscopy (SECM) investigations was evaluated in the challenging situation of an ultrathin layer deposited on a conductive substrate (carbon materials). Experiments show that dopamine is a well-adapted redox system for SECM in feedback mode and in unbiased conditions. Used as a redox mediator, catechol permits the investigations of modified surfaces without an electrical connection of the sample thanks to fast charge transfer kinetics but with a surface selectivity that does not exist in classical outer-sphere redox mediators. The interest of catechol in SECM as a sensitive redox mediator is exemplified by monitoring several modification steps of an ultrathin (<1 nm) hierarchically porous organic monolayer deposited on carbon substrates. For quantitative analysis, the SECM approach curves using dopamine could simply be characterized with an irreversible electron transfer kinetics model in a large range of pH.     

Surface reactivity from electrochemical lithography: illustration in the steady-state reductive etching of perfluorinated surfaces

The scanning electrochemical microscope (SECM) in the lithographic mode is used to assess quantitatively, from both theoretical and experimental points of view, the kinetics of irreversible transformation of electroactive molecular moieties immobilized on a surface as self-assembled monolayers (SAMs). The SECM tip allows the generation of an etchant that transforms the surface locally and irreversibly. The resulting surface patterning is detectable by different surface analyses. The quantification of the surface transformation kinetics is deduced from the evolution of the pattern dimensions with the etching time. The special case of slow etching kinetics is presented; it is predicted that the pattern evolution follows the expansion of the etchant at the substrate surface. The case of a chemically unstable etchant is considered. The model is then tested by inspecting the slow reductive patterning of a perfluorinated SAM. Good agreement is found with different independent SECM interrogation modes, depending on the insulating or conducting nature of the covered substrate. The surface transformation measurements are also compared to the reduction of solutions of perfluoroalkanes. The three-orders-of-magnitude-slower electron transfer observed at the immobilized molecules likely describes the large reorganization associated with the generation of a perfluoroalkyl-centered radical anion.     

Probing heterogeneous electron transfer at an unbiased conductor by scanning electrochemical microscopy in the feedback mode

The theory of the feedback mode of scanning electrochemical microscopy is extended for probing heterogeneous electron transfer at an unbiased conductor. A steady-state SECM diffusion problem with a pair of disk ultramicroelectrodes as a tip and a substrate is solved numerically. The potential of the unbiased substrate is such that the net current flow across the substrate/solution interface is zero. For a reversible substrate reaction, the potential and the corresponding tip current depend on SECM geometries with respective to the tip radius including not only the tip-substrate distance and the substrate radius but also the thickness of the insulating sheath surrounding the tip. A larger feedback current is obtained using a probe with a thinner insulating sheath, enabling identification of a smaller unbiased substrate with a radius that is approximately as small as the tip radius. An intrinsically slow reaction at an unbiased substrate as driven by a SECM probe can be quasi-reversible. The standard rate constant of the substrate reaction can be determined from the feedback tip current when the SECM geometries are known. The numerical simulations are extended to an SECM line scan above an unbiased substrate to demonstrate a "dip" in the steady-state tip current above the substrate center. The theoretical predictions are confirmed experimentally for reversible and quasi-reversible reactions at an unbiased disk substrate using disk probes with different tip radii and outer radii.     

Fabrication of nanometer-sized electrodes and tips for scanning electrochemical microscopy.

A novel method for fabrication of nanometer-sized electrodes and tips suitable for scanning electrochemical microscopy (SECM) is reported. A fine etched Pt wire is coated with polyimide, which was produced by polymerization on the Pt surface initiated by heat. This method can prepare electrodes with effective radii varying from a few to hundreds of nanometers. Scanning electron microscopy, cyclic voltammetry, and SECM were used to characterize these electrodes. Well-defined steady-state voltammograms could be obtained in aqueous or in 1,2-dichloroethane solutions. This method produced the nanoelectrodes with exposed Pt on the apex, and they can also be employed as the nanotips for SECM investigations. Different sizes of Pt nanotips made by this method were employed to evaluate the kinetics of the redox reaction of Ru(NH3)6(3+) on the surface of a large Pt electrode by SECM, and the standard rate constant kappa0 of this system was calculated from the best fit of the SECM approach curve. This result is similar to the values obtained by analysis of the obtained voltammetric data.     

Scanning electrochemical microscopy. 58. Application of a micropipet-supported ITIES tip to detect Ag+ and study its effect on fibroblast cells

We report the use of a micropipet-supported ITIES (interface between two immiscible electrolyte solutions, also called a liquid/liquid (L/L) or water/oil (W/O) interface) as a scanning electrochemical microscopy (SECM) tip to detect silver ion and explore Ag+ toxicity in living cells. A 1,2-dichloroethane solution containing a commercially available calixarene-based Ag+ ionophore (IV) was injected into a micrometer-size glass pipet to construct an Ag+-selective SECM tip. The local Ag+ concentration, down to the micromolar level, in the vicinity of living fibroblast cells, was monitored by SECM approach curves and through imaging of the uptake and efflux of Ag+ by living fibroblast cells in real time. The results show that several stages of interaction between Ag+ and fibroblast cells exist. Since a number of biological processes of cells are involved with non-redox-active ions, the work presented here provides a new way to explore cell metabolism, drug delivery, and toxicity assessment by SECM.     

Polished nanopipets: new probes for high-resolution scanning electrochemical microscopy

Nanometer-sized pipets pulled from glass or quartz capillaries have been extensively used as probes for scanning electrochemical microscopy (SECM) and scanning ion conductance microscopy (SICM). A small separation distance between such a probe and the sample, which is required for high-resolution SECM measurements, may be hard to attain because of considerable roughness of the pipet tip. In this Letter, we report the preparation and characterization of polished nanopipet SECM probes with a much smoother tip edge. Using polished pipets, quantitative SECM measurements were performed at extremely short tip/substrate distances (e.g., d ≈ 1 nm).     

Application of scanning electrochemical microscope in the study of corrosion of metals

Scanning electrochemical microscope (SECM) has become a very useful and powerful technique for probing a variety of electrochemical reactions in corrosion process due to its high spatial resolution and electrochemical sensitivity to characterize the topography and redox activities of the metal/electrolyte solution interface. Its capability for the direct identification of chemical species in localized corrosion processes with high spatial resolution would be more advantageous compared to other local probe techniques with only morphological characterization. In this review, the applications of the SECM in the study of early stages of localized corrosion, electroactive defect sites in passive films, local initiation of pits, degradation of coating properties on steels, and some combined methods through SECM integrated with other techniques have been summarized and commented. Finally, the optimization for SECM’s experiment design and operation as well as foreseeable application range has been proposed.     

Scanning electrochemical microscopy of living cells. 3. Rhodobacter sphaeroides.

The scanning electrochemical microscope (SECM) was used to probe the redox activity of individual purple bacteria (Rhodobacter sphaeroides). The approaches developed in our previous studies of mammalian cells were expanded to measure the rates and investigate the pathway of transmembrane charge transfer in bacteria. The two groups of redox mediators (i.e., hydrophilic and hydrophobic redox species) were used to shuttle the electrons between the SECM tip electrode in solution and the redox centers inside the cell. The analysis of the dependencies of the measured rate constant on formal potential and concentration of mediator species in solution yielded information about the permeability of the outer cell membrane to different ionic species and intracellular redox properties. The maps of redox reactivity of the cell surface were obtained with a micrometer or submicrometer spatial resolution.     

Alternating current impedance imaging of high-resistance membrane pores using a scanning electrochemical microscope. Application of membrane electrical shunts to increase measurement sensitivity and image contrast

Whether an individual pore in a porous membrane can be imaged using scanning electrochemical microscopy (SECM), operated in ac impedance mode, is determined by the magnitude of the change in the total impedance of the imaging system as the SECM tip is scanned over the pore. In instances when the SECM tip resistance is small relative to the internal pore resistance, the total impedance changes by a negligible amount, rendering the pore invisible during impedance imaging. A simple solution to this problem is to introduce a low-impedance electrical shunt (i.e., a salt bridge) across the membrane. This principle is demonstrated by imaging polycarbonate membranes (6-12-microm thickness) containing between 1 and 2000 conical-shaped pores (60-nm- and 2.5-microm-diameter openings) using an approximately 1-microm-radius Pt tip. Theory and experiments show that image contrast (the change in ac current measured as the probe is scanned over the pore) is inversely proportional to the total resistance of the membrane and can be increased by a factor of approximately 50x by introducing a low-resistance electrical shunt across the membrane. Remarkably, SECM images of membranes containing a single high-resistance (approximately 1 G Omega) pore can only be imaged by short-circuiting the membrane. Image contrast also becomes independent of membrane resistance when an electrical shunt is used, allowing for more quantitative comparisons of the features in ac impedance images of different membranes.     

Integrated scanning Kelvin probe-scanning electrochemical microscope system: development and first applications

The integration of a scanning Kelvin probe (SKP) and a scanning electrochemical microscope (SECM) into a single SKP-SECM setup, the concept of the proposed system, its technical realization, and first applications are presented and discussed in detail. A preloaded piezo actuator placed in a grounded stainless steel case was used as the driving mechanism for oscillation of a Pt disk electrode as conventionally used in SECM when the system was operated in the SKP mode. Thus, the same tip is recording the contact potential difference (CPD) during SKP scanning and is used as a working electrode for SECM imaging in the redox-competition mode (RC-SECM). The detection of the local CPD is established by amplification of the displacement current at an ultralow noise operational amplifier and its compensation by application of a variable backing potential (V(b)) in the external circuit. The control of the tip-to-sample distance is performed by applying an additional alternating voltage with a much lower frequency than the oscillation frequency of the Kelvin probe. The main advantage of the SKP-SECM system is that it allows constant distance measurements of the CPD in air under ambient conditions and in the redox-competition mode of the SECM in the electrolyte of choice over the same sample area without replacement of the sample or exchange of the working electrode. The performance of the system was evaluated using a test sample made by sputtering thin Pt and W films on an oxidized silicon wafer. The obtained values of the CPD correlate well with known data, and the electrochemical activity for oxygen reduction is as expected higher over Pt than W.     

Combined scanning electrochemical/optical microscopy with shear force and current feedback

A technique that combines scanning electrochemical microscopy (SECM) and scanning optical microscopy (OM) was developed. Simultaneous scanning electrochemical/optical microscopy (SECM/OM) was performed by a special probe tip, which consists of an optical fiber core for light passage, surrounded by a gold ring electrode, and an outermost electrophoretic insulating sheath, with the tip attached to a tuning fork. To regulate the tip-substrate distance, either the shear force or the SECM tip current was employed as the feedback signal. The application of a quartz crystal tuning fork (32.768 kHz) for sensing shear force allowed simultaneous topographic, along with SECM and optical imaging in a constant-force mode. The capability of this technique was confirmed by obtaining simultaneously, for the first time, topographic, electrochemical, and optical images of an interdigitated array electrode. Current feedback from SECM also provided simultaneous electrochemical and optical images of relatively soft samples, such as a polycarbonate membrane filter and living diatoms in a constant-current mode. This mode should be useful in mapping the biochemical activity of a living cell.     

Kinetics of electron-transfer reactions at nanoelectrodes

The kinetics of several fast heterogeneous electron-transfer reactions were investigated by steady-state voltammetry at nanoelectrodes and scanning electrochemical microscopy (SECM). The disk-type, polished Pt nanoelectrodes (3.7-400-nm radius) were characterized by a combination of voltammetry, scanning electron microscopy, and SECM. A number of experimental curves were obtained at the same nanoelectrode to attain the accuracy and reproducibility similar to those reported previously for micrometer-sized probes. A new analytical approximation was developed and used for analysis of steady-state tip voltammograms. The self-consistent kinetic parameter values with the uncertainty margin of approximately 10% were obtained for electrodes of different radii and for a wide range of the SECM tip/substrate separation distances. The determined standard rate constants are compared to those previously measured at the electrodes of different dimensions, and the correlation between the heterogeneous and self-exchange rate constants is discussed.     

Scanning electrochemical microscopy of quinoprotein glucose dehydrogenase

The activity of immobilized glucose dehydrogenase (GDH), a typical PQQ-dependent quinoprotein, was studied qualitatively and quantitatively by scanning electrochemical microscopy (SECM). PQQ-dependent GDH is of interest because of its high activity and independence of dissolved oxygen in catalyzing the transfer of electrons from glucose to an electron mediator. Biotinylated glucose dehydrogenase was bound to streptavidin-coated paramagnetic beads (surface concentration > or = 1.8 x 10(-11) mol cm(-2)) which were deposited as microscopic microspots on a hydrophobic surface. The catalytic activity of immobilized GDH was mapped in SECM feedback mode and generation-collection mode using ferrocenemethanol, ferrocenecarboxylic acid, p-aminophenol, and ferricyanide as electron mediators, respectively. The apparent steady-state kinetics of catalysis were measured under conditions of high d-glucose concentration using the theory developed for the SECM feedback and generation collection (GC) modes. In feedback mode, curves of the kinetically controlled substrate current against normalized distance were plotted, and it was found that GDH catalysis follows pseudo-first-order kinetics. In GC mode detection, the catalysis follows zero-order kinetics in the presence of high concentration of both substrates for GDH. The turnover rate obtained for immobilized GDH is lower than that of native GDH but much higher than that generally observed for glucose oxidase.