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Molecular photocatalysis and photoelectrocatalysis have been widely used to conduct oxidation-reduction processes ranging from fuel generation to electroorganic synthesis. We recently showed that an electrostatic potential drop across the double layer contributes to the driving force for electron transfer (ET) between a dissolved reactant and a molecular catalyst immobilized directly on the electrode surface. In this article, we report voltage-driven molecular photoelectrocatalysis with a prevalent homogeneous water oxidation catalyst, (bpy)Cu (II), which was covalently attached to the carbon surface and exhibited photocatalytic activity. The strong potential dependence of the photooxidation current suggests that the electrostatic potential drop across the double layer contributes to the driving force for ET between a water molecule and the excited state of surface-bound (bpy)Cu (II). Scanning electrochemical microscopy (SECM) was used to analyze the products and determine the faradaic efficiencies for the generation of oxygen and hydrogen peroxide. Unlike electrocatalytic water oxidation by (bpy)Cu (II) in the dark, which produces only O 2 , the voltage-driven photooxidation includes an additional 2e - pathway generating H 2 O 2 . DFT calculations show that the applied voltage and the presence of light can alter the activation energy for the rate-determining water nucleophilic attack steps, thereby increasing the reaction rate of photo-oxidation of water and opening the 2e - pathway. These results suggest a new route for designing next-generation hybrid molecular photo(electro)catalysts for water oxidation and other processes.

Scanning electrochemical microscopy (SECM) has been extensively used for mapping electrocatalytic surface reactivity; however, most of the studies were carried out using micrometer-sized tips, and no quantitative kinetic experiments on the nanoscale have yet been reported to date. As the diffusion-limited current density at a nanometer-sized electrode is very high, an inner-sphere electron-transfer process occurring at a nanotip typically produces a kinetic current at any attainable overpotential. Here, we develop a theory for substrate generation/tip collection (SG/TC) and feedback modes of SECM with a kinetic tip current and use it to evaluate the rates of hydrogen and oxygen evolution reactions in a neutral aqueous solution from the current-distance curves. The possibility of using chemically modified nanotips for kinetic measurements is also demonstrated. The effect of the substrate size on the shape of the current-distance curves in SG/TC mode SECM experiments is discussed.

Particulate bismuth vanadate (BiVO 4 ) has attracted considerable interest as a promising photo(electro)catalyst for visible-light-driven water oxidation; however, overall water splitting (OWS) has been difficult to attain because its conduction band is too positive for efficient hydrogen evolution. Using photoscanning electrochemical microscopy (photo-SECM) with a chemically modified nanotip, we visualized for the first time the OWS at a single truncated bipyramidal microcrystal of phosphorus-doped BiVO 4 . The tip simultaneously served as a light guide to illuminate the photocatalyst and an electrochemical nanoprobe to observe and quantitatively measure local oxygen and hydrogen fluxes. The obtained current patterns for both O 2 and H 2 agree well with the accumulation of photogenerated electrons and holes on {010} basal and {110} lateral facets, respectively. The developed experimental approach is an important step toward nanoelectrochemical mapping of the activity of photocatalyst particles at the subfacet level.

Electrochemical resistive-pulse (ERP) sensing with conductive carbon nanopipettes (CNPs) has recently been developed and employed for the detection of single liposomes and biological vesicles, and for the analysis of redox molecules contained in such vesicles. However, the origins of different shapes of current transients produced by the translocation of single vesicles through the CNP remain poorly understood. Herein, we report extensive finite-element simulations of both portions of an ERP transient, the current blockage by a vesicle approaching and passing through the pipet orifice and the faradaic current spike due to oxidation/reduction of redox species released from a vesicle on the carbon surface, for different values of parameters defining the geometry and dynamics of the vesicle/CNP system. The effects of the pipet geometry, surface charge, transport, vesicle trajectory, and collision location on the shape of current transients are investigated. The possibility of quantitative analysis of experimental ERP transients produced by translocations of liposomes and extracellular vesicles by fitting them to simulated curves is demonstrated. The developed theory can enable a more reliable interpretation of complicated ERP signals and characterization of the size and contents of single biological and artificial vesicles.

Particulate photocatalysts for the overall water splitting (OWS) reaction offer promise as devices for hydrogen fuel generation. Even though such photocatalysts have been studied for nearly 5 decades, much of the understanding of their function is derived from observations of catalyst ensembles and macroscopic photoelectrodes. This is because the sub-micrometer size of most OWS photocatalysts makes spatially resolved measurements of their local reactivity very difficult. Here, we employ photo-scanning electrochemical microscopy (photo-SECM) to quantitatively measure hydrogen and oxygen evolution at individual OWS photocatalyst particles for the first time. Micrometer-sized Al-doped SrTiO 3 /Rh 2- y Cr y O 3 photocatalyst particles were immobilized on a glass substrate and interrogated with a chemically modified SECM nanotip. The tip simultaneously served as a light guide to illuminate the photocatalyst and as an electrochemical nanoprobe to observe oxygen and hydrogen fluxes from the OWS. Local O 2 and H 2 fluxes obtained from chopped light experiments and photo-SECM approach curves using a COMSOL Multiphysics finite-element model confirmed stoichiometric H 2 /O 2 evolution of 9.3/4.6 μmol cm -2 h -1 with no observable lag during chopped illumination cycles. Additionally, photoelectrochemical experiments on a single microcrystal attached to a nanoelectrode tip revealed a strong light intensity dependence of the OWS reaction. These results provide the first confirmation of OWS at single micrometer-sized photocatalyst particles. The developed experimental approach is an important step toward assessing the activity of photocatalyst particles at the nanometer scale.

The integration of heterogeneous electrocatalysis and molecular catalysis is a promising approach to designing new catalysts for the oxygen evolution reaction (OER) and other processes. We recently showed that the electrostatic potential drop across the double layer contributes to the driving force for electron transfer between a dissolved reactant and a molecular catalyst immobilized directly on the electrode surface. Here, we report high current densities and low onset potentials for water oxidation attained using a metal-free voltage-assisted molecular catalyst (TEMPO). Scanning electrochemical microscopy (SECM) was used to analyze the products and determine faradic efficiencies for the generation of H 2 O 2 and O 2 . The same catalyst was employed for efficient oxidations of butanol, ethanol, glycerol, and H 2 O 2 . DFT calculations show that the applied voltage alters the electrostatic potential drop between TEMPO and the reactant as well as chemical bonding between them, thereby increasing the reaction rate. These results suggest a new route for designing next-generation hybrid molecular/electrocatalysts for OER and alcohol oxidations.

Extracellular vesicles (EVs) released from biological cells have attracted considerable interest due to their potential for cancer diagnostics and important role in cell signaling. Most previously reported studies have been concerned with the detection of EVs in biofluids and analysis of proteins and nucleic acids they contain. Electrochemical resistive-pulse (ERP) sensing enables direct detection of single EVs released from a specific cell and analysis of reactive oxygen and nitrogen species in such vesicles. Here, we demonstrate the applicability of ERP sensing to distinguish between nontransformed and cancerous breast cell lines as well as between breast cancer cell lines with different metastatic potential. Another application of ERP sensing is in real-time monitoring of changes in a single cell induced by a chemical agent. This approach is potentially useful for evaluating the efficacy of therapeutic agents, including those that trigger breast cancer cell death by inducing intense oxidative stress.

Anion exchange membrane fuel cells are limited by the slow kinetics of alkaline hydrogen oxidation reaction (HOR). Here, we establish HOR catalytic activities of single-atom and diatomic sites as a function of *H and *OH binding energies to screen the optimal active sites for the HOR. As a result, the Ru-Ni diatomic one is identified as the best active center. Guided by the theoretical finding, we subsequently synthesize a catalyst with Ru-Ni diatomic sites supported on N-doped porous carbon, which exhibits excellent catalytic activity, CO tolerance, and stability for alkaline HOR and is also superior to single-site counterparts. In situ scanning electrochemical microscopy study validates the HOR activity resulting from the Ru-Ni diatomic sites. Furthermore, in situ x-ray absorption spectroscopy and computational studies unveil a synergistic interaction between Ru and Ni to promote the molecular H 2 dissociation and strengthen OH adsorption at the diatomic sites, and thus enhance the kinetics of HOR.

The use of scanning electrochemical microscopy (SECM) for nanoscale imaging of photoelectrochemical processes at semiconductor surfaces has recently been demonstrated. To illuminate a microscopic portion of the substrate surface facing the SECM probe, a glass-sealed, polished tip simultaneously served as a nanoelectrode and a light guide. One issue affecting nanoscale photo-SECM experiments is mechanical interactions of the rigid optical fiber with the tip motion controlled by the piezo-positioner. Here we report an improved experimental setup in which the tip is mechanically decoupled from the fiber and light is delivered to the back of the tip capillary using a complex lens system. The advantages of this approach are evident from the improved quality of the approach curves and photo-SECM images. The light intensity delivered from the optical fiber to the tip is not changed significantly by their decoupling.

Heterogeneous electrocatalysis and molecular redox catalysis have developed over several decades as two distinct ways to facilitate charge-transfer processes essential for energy conversion and storage. Whereas electrocatalytic reactions are driven by the applied voltage, molecular catalytic processes are driven by the difference between standard potentials of the catalyst and the reactant. Here, we demonstrate that the rate of electron transfer between a dissolved reactant and a molecular catalyst immobilized directly on the surface of a carbon nanoelectrode is governed by combination of chemical driving force and electrostatic potential drop across the double layer. DFT calculations show that varying the applied voltage alters the potential drop between the surface-bound and dissolved redox species. These results suggest a new route for designing next-generation hybrid molecular/electrocatalysts.

Scanning electrochemical microscopy (SECM) is a powerful tool for mapping surface reactivity. Electrochemical mapping of electrocatalytic processes at the nanoscale is, however, challenging because the surface of a nanoelectrode tip is easily fouled by impurities and/or deactivated by products and intermediates of innersphere surface reactions. To overcome this difficulty, we introduce new types of SECM nanotips based on bimolecular electron transfer between the dissolved electroactive species and a redox mediator attached to the surface of a carbon nanoelectrode. A tris(2,2'-bipyridine)ruthenium complex, Ru(bpy) 3 , that undergoes reversible oxidation/reduction reactions at both positive and negative potentials was used to prepare the SECM nanoprobes for mapping a wide range of electrocatalytic processes through oxidation of H 2 , reduction of O 2 , and both oxidation and reduction of H 2 O 2 at the tip. In addition to high-resolution reactivity mapping and localized kinetic measurements, chemically modified nanoelectrodes can serve as nanosensors for a number of important analytes such as reactive oxygen and nitrogen species and neurotransmitters.

Scanning electrochemical microscopy (SECM) is a powerful technique for mapping surface reactivity and investigating heterogeneous processes on the nanoscale. Despite significant advances in high-resolution SECM and photo-SECM imaging, they cannot provide atomic scale structural information about surfaces. By correlating the SECM images with atomic scale structural and bonding information obtained by transmission electron microscopy (TEM) techniques with one-to-one correspondence, one can elucidate the nature of the active sites and understand the origins of heterogeneous surface reactivity. To enable multitechnique imaging of the same nanoscale portion of the electrode surface, we develop a methodology for using a TEM finder grid as a conductive support in SECM and photo-SECM experiments. In this paper, we present the results of our first nanoscale SECM and photo-SECM experiments on carbon TEM grids, including imaging of semiconductor nanorods.

Carbon nanotube yarn microelectrodes (CNTYMEs) are an alternative to carbon-fiber microelectrodes (CFMEs) with interesting electrochemical properties because analyte is momentarily trapped in cavities between the CNTs. Here, we compare fast-scan cyclic voltammetry (FSCV) detection of catecholamines, including dopamine, norepinephrine, and epinephrine, at CNTYMEs, CFMEs, as well as cavity carbon nanopipette electrodes (CNPEs). At CFMEs, current decreases dramatically at high FSCV repetition frequencies. At CNTYMEs, current is almost independent of FSCV repetition frequency because the analytes are trapped in the crevices between CNTs, and thus the electrode acts like a thin-layer cell. At CFMEs, small cyclization product peaks are observed due to an intramolecular cyclization reaction to form leucocatecholamine, which is electroactive, and these peaks are largest for the secondary amine epinephrine. At CNTYMEs, more of the leucocatecholamine cyclization product is detected for all catecholamines because of the enhanced trapping effects, particularly at higher repetition rates where the reaction occurs more frequently and more product is accumulated. For epinephrine, the secondary peaks have larger currents than the primary oxidation peaks at 100 Hz, and similar trends are observed with faster scan rates and 500 Hz repetition frequencies. Finally, we examined CNPEs, which also momentarily trap neurotransmitters. Similar to CNTYMEs, at CNPEs, catecholamines have robust cyclization peaks, particularly at high repetition rates. Thus, CNTYMEs and CNPEs have thin layer cell behavior that facilitates high temporal resolution measurements, but catecholamines CVs are complicated by cyclization reactions. However, those additional peaks could be useful in discriminating the analytes, particularly epinephrine and norepinephrine., Competing Interests: The authors declare no conflicts of interest. Declaration of interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

In this work, open carbon nanopipettes (CNPs) with radius between 50 and 600 nm were used to control translocation of different-sized vesicles through the pipette orifice followed by nanoelectrochemical analysis. Vesicle impact electrochemical cytometry (VIEC) was used to determine the number of catecholamine molecules expelled from single vesicles onto an inner-wall carbon surface, where the duration of transmitter release was quantified and correlated to the vesicle size all in the same nanotip. This in turn allowed us to both size and count molecules for vesicles in a living cell. Here, small and sharp open CNPs were employed to carry out intracellular VIEC with minimal invasion and high sensitivity. Our findings with VIEC reveal that the vesicular content increases with vesicle size. The release kinetics of vesicular transmitters and dense core size have the same relation with the vesicle size, implying that the vesicular dense core size determines the speed of each release event. This direct correlation unravels one of the complexities of exocytosis.

The continuing interest in nanoscale research has spurred the development of nanosensors for liquid phase measurements. These include nanopore-based sensors typically employed for detecting nanoscale objects, such as nanoparticles, vesicles and biomolecules, and electrochemical nanosensors suitable for identification and quantitative analysis of redox active molecules. In this Perspective, we discuss conductive nanopipettes (CNP) that can combine the advantages of single entity sensitivity of nanopore detection with high selectivity and capacity for quantitative analysis offered by electrochemical sensors. Additionally, the small physical size and needle-like shape of a CNP enables its use as a tip in the scanning electrochemical microscope (SECM), thus, facilitating precise positioning and localized measurements in biological systems., Competing Interests: The authors declare that they have no conflict of interest., (This journal is © The Royal Society of Chemistry.)

Electrochemical monitoring of catalytically amplified collisions of individual metal nanoparticles (NP) with ultramicroelectrodes (UME) has been extensively used to study electrocatalysis, mass-transport, and charge-transfer processes at the single NP level. More recently, photoelectrochemical collision experiments were carried out with semiconductive NPs. Here, we introduce two new types of light-controlled nanoimpact experiments. The first experiment involves localized photodeposition of catalyst (Pt) on TiO 2 NPs with a glass-sheathed carbon fiber simultaneously serving as the light guide and collector UME. The collisions of in situ prepared Pt@TiO 2 NPs with the carbon surface produced blips of water oxidation current, while the activity of pristine TiO 2 NPs was too low to yield measurable signal. In another experiment, collisions of catalytic (Ir oxide) NPs with the semiconductor (Nb doped n-type TiO 2 rutile single crystal) electrode are monitored by measuring the photocurrent of water oxidation.

Resistive-pulse sensing is a technique widely used to detect single nanoscopic entities such as nanoparticles and large molecules that can block the ion current flow through a nanopore or a nanopipette. Although the species of interest, e.g., antibodies, DNA, and biological vesicles, are typically produced by living cells, so far, they have only been detected in the bulk solution since no localized resistive-pulse sensing in biological systems has yet been reported. In this report, we used a nanopipette as a scanning ion conductance microscopy (SICM) tip to carry out resistive-pulse experiments both inside immobilized living cells and near their surfaces. The characteristic changes in the ion current that occur when the pipet punctures the cell membrane are used to monitor its insertion into the cell cytoplasm. Following the penetration, cellular vesicles (phagosomes, lysosomes, and/or phagolysosomes) were detected inside a RAW 264.7 macrophage. Much smaller pipettes were used to selectively detect 10 nm Au nanoparticles in the macrophage cytoplasm. The in situ resistive-pulse detection of extracellular vesicles released by metastatic human breast cells (MDA-MB-231) is also demonstrated. Electrochemical resistive-pulse experiments were carried out by inserting a conductive carbon nanopipette into a macrophage cell to sample single vesicles and measure reactive oxygen and nitrogen species (ROS/RNS) contained inside them.

Resistive-pulse sensing with biological or solid-state nanopores and nanopipettes has been widely employed in detecting single molecules and nanoparticles. The analytical signal in such experiments is the change in ionic current caused by the molecule/particle translocation through the pipet orifice. This paper describes a new version of the resistive-pulse technique based on the use of carbon nanopipettes (CNP). The measured current is produced by electrochemical oxidation/reduction of redox molecules at the carbon surface and responds to the particle translocation. In addition to counting single entities, this technique enables qualitative and quantitative analysis of the electroactive material they contain. Using liposomes as a model system, we demonstrate the capacity of CNPs for (1) conventional resistive-pulse sensing of single liposomes, (2) electrochemical resistive-pulse sensing, and (3) electrochemical identification and quantitation of redox species (e.g., ferrocyanide, dopamine, and nitrite) contained in a single liposome. The small physical size of a CNP suggests the possibility of single-entity measurements in biological systems.

High-resolution scanning electrochemical microscopy (SECM) is a powerful technique for mapping surface topography and reactivity on the nanoscale and investigating heterogeneous processes at the level of single nanoparticles. The ability to fabricate ultrasmall nanoelectrode tips is critical for the progress in nano-SECM. Despite long-term efforts to improve previously developed procedures, the preparation and characterization of disk-type polished tips with the radius <∼25 nm remains challenging and unpredictable. One of the problems is that the geometry of such tips is hard to characterize by either SEM or atomic force microscopy (AFM) that has been employed for examination of somewhat larger nanoelectrodes. Herein, we report a new approach to more predictable and reproducible two-step fabrication of ultrasmall (≤10 nm radius) polished Pt electrodes assisted by transmission electron microscopy (TEM) imaging. Both voltammetric and SECM responses of the prepared nanoelectrodes are consistent with the size and geometry extracted from TEM images. These tips can be used to attain sub-10 nm spatial resolution of SECM imaging and kinetic studies.

Carbon fiber micro- and nanoelectrodes have been extensively used to measure dopamine and other neurotransmitters in biological systems. Although the radii of some reported probes were ≪1 μm, the lengths of the exposed carbon were typically on the micrometer scale, thus limiting the spatial resolution of electroanalytical measurements. Recent attempts to determine neurotransmitters in single cells and vesicles have provided additional impetus for decreasing the probe dimensions. Here, we report two types of dopamine sensors based on carbon nanopipets (CNP) prepared by chemical vapor deposition of carbon into prepulled quartz capillaries. These include 10-200 nm radius CNPs with a cavity near the orifice and CNPs with an open path in the middle, in which the volume of sampled solution can be controlled by the applied pressure. Because of the relatively large surface area of carbon exposed to solution inside the pipet, both types of sensors yielded well-shaped voltammograms of dopamine down to ca. 1 nM concentrations, and the unprecedented voltammetric response to 100 pM dopamine was obtained with open CNPs. TEM tomography and numerical simulations were used to model CNP responses. The effect of dopamine adsorption on the CNP detection limit is discussed along with the possibilities of measuring other physiologically important analytes (e.g., serotonin) and eliminating anionic and electrochemically irreversible interferences (e.g., ascorbic acid).

Scanning electrochemical microscopy (SECM) has previously been employed in probing photoelectrochemical processes at semiconductor surfaces. However, the spatial resolution of these studies has not yet matched the nanoscale SECM resolution attained without substrate illumination. Herein, we introduce nanoscale photo-SECM with a glass-sealed, polished tip simultaneously serving as a nanoelectrode and a light guide to produce a microscopic light spot on the substrate surface. The advantages of this approach are demonstrated by comparing current transients obtained using through-tip and global illumination of the sample. The spot of light on the substrate surface facing the nanotip was sufficiently bright to measure the diffusion-controlled positive feedback current in good agreement with the theory. We employed this approach for high-resolution photoelectrochemical mapping of ferrocenemethanol oxidation and oxygen evolution reactions at the Nb:TiO 2 rutile (110) single crystal surface. The images obtained using 40-50 nm radius tips showed only minor and random variations in photoelectrochemical reactivity for both processes, pointing to essentially uniform distribution of the Nb dopant over the TiO 2 surface and no measurable segregation on the ∼ 50 nm scale.

The catalytic activity of low-dimensional electrocatalysts is highly dependent on their local atomic structures, particularly those less-coordinated sites found at edges and corners; therefore, a direct probe of the electrocatalytic current at specified local sites with true nanoscopic resolution has become critically important. Despite the growing availability of operando imaging tools, to date it has not been possible to measure the electrocatalytic activities from individual material edges and directly correlate those with the local structural defects. Herein, we show the possibility of using feedback and generation/collection modes of operation of the scanning electrochemical microscope (SECM) to independently image the topography and local electrocatalytic activity with 15-nm spatial resolution. We employed this operando microscopy technique to map out the oxygen evolution activity of a semi-2D nickel oxide nanosheet. The improved resolution and sensitivity enables us to distinguish the higher activities of the materials' edges from that of the fully coordinated surfaces in operando The combination of spatially resolved electrochemical information with state-of-the-art electron tomography, that unravels the 3D complexity of the edges, and ab initio calculations allows us to reveal the intricate coordination dependent activity along individual edges of the semi-2D material that is not achievable by other methods. The comparison of the simulated line scans to the experimental data suggests that the catalytic current density at the nanosheet edge is ∼200 times higher than that at the NiO basal plane., Competing Interests: The authors declare no conflict of interest.

Ion transport controlled by electrostatic interactions is an important phenomenon in biological and artificial membranes, channels, and nanopores. Here, we employ carbon-coated nanopipets (CNPs) for studying permselective electrochemistry in a conductive nanopore. A significant accumulation (up to 2000-fold) of cationic redox species and anion depletion inside a CNP by diffuse-layer and surface-charge effects in a solution of low ionic strength were observed as well as the shift of the voltammetric midpeak potential. Finite-element simulations of electrostatic effects on CNP voltammograms show permselective ion transport in a single conducting nanopore and semiquantitatively explain our experimental data. The reported results are potentially useful for improving sensitivity and selectivity of CNP sensors for ionic analytes.

Microelectrodes are typically used for neurotransmitter detection, but nanoelectrodes are not because there is a trade-off between spatial resolution and sensitivity that is dependent on surface area. Cavity carbon-nanopipette electrodes (CNPEs), with tip diameters of a few hundred nanometers, have been developed for nanoscale electrochemistry. Here, we characterize the electrochemical performance of CNPEs with fast-scan cyclic voltammetry (FSCV) for the first time. Dopamine detection using cavity CNPEs, with a depth equivalent to a few radii, is compared with that using open-tube CNPEs, an essentially infinite geometry. Open-tube CNPEs have very slow temporal responses that change over time as the liquid rises in the CNPE. However, a cavity CNPE has a fast temporal response to a bolus of dopamine that is not different from that of a traditional carbon-fiber microelectrode. Cavity CNPEs, with tip diameters of 200-400 nm, have high currents because the small cavity traps and increases the local dopamine concentration. The trapping also leads to an FSCV frequency-independent response and the appearance of cyclization peaks that are normally observed only with large concentrations of dopamine. CNPEs have high dopamine selectivity over ascorbic acid (AA) because of the repulsion of AA by the negative electric field at the holding potential and the irreversible redox reaction. In mouse-brain slices, cavity CNPEs detected exogenously applied dopamine, showing they do not clog in tissue. Thus, cavity CNPEs are promising neurochemical sensors that provide spatial resolution on the scale of hundreds of nanometers, which is useful for small model organisms or for locations near specific cells.

The release of reactive oxygen and nitrogen species (ROS/RNS) by macrophages undergoing phagocytosis is crucial for the efficiency of the immune system. In this work, platinized carbon nanoelectrodes were used to detect, characterize, and quantify for the first time the intracellular production rates of the four primary ROS/RNS (i.e., H 2 O 2 , ONOO - , NO • , and NO 2 - ) inside single phagolysosomes of living RAW 264.7 murine macrophages stimulated by interferon-γ and lipopolysaccharide (IFN-γ/LPS) to mimic an in vivo inflammatory activation. The time-dependent concentrations of the four primary ROS/RNS in individual phagolysosomes monitored using a four-step chronoamperometric method evidenced a high variability of their production rates. This intrinsic variability unravels the complexity of phagocytosis.

Fast and cost-efficient detection and identification of bacteria in food and water samples and biological fluids is an important challenge in bioanalytical chemistry. It was shown recently that bacteria can be detected by measuring the decrease in the diffusion current to the ultramicroelectrode caused by cell collisions with its surface. To add selectivity to the bacteria detection, herein we show the possibility of collision experiments with the signal produced by electrochemical activity of bacterial cells reducing (or oxidizing) redox species. The mediator oxidation/reduction rate can be used to identify different types of bacteria based on their specific redox activities. Here we report the analysis of electrochemical collision transients produced by two kinds of bacteria, Escherichia coli and Stenotrophomonas maltophilia. The effects of the charge and redox activity of bacterial cells on collision events are discussed. The current transients due to live cell collisions were compared to those produced by bacteria killed either by heavy metal ions (cobalt) or by an antibiotic (colistin). This approach is potentially useful for evaluating the effectiveness of antimicrobial agents. Finite-element simulations were carried out to model collision transients.

This paper is concerned with long-distance interactions between an unbiased metal nanoparticle (NP) and a nanoelectrode employed as a tip in the scanning electrochemical microscope (SECM). A NP immobilized on the inert substrate acts as a bipolar electrode, producing positive SECM feedback. The tip current magnitude depends strongly on the ratio of the particle and tip radii and the heterogeneous charge-transfer kinetics. The onset of electron tunneling was observed at very short separation distances (<2-3 nm) at which the NP behaves as a part of the tip electrode. The rate constant of the electron-transfer (ET) or electrocatalytic reaction at the NP can be extracted from either feedback or tunneling current. The tunneling mode of SECM can be used to investigate heterogeneous reactions occurring at a single NP without making an ohmic contact with it. This technique can also help elucidate nanoparticle/electrode interactions in various electrochemical systems ranging from NPs immobilized on the electrode surface to nanoimpact collision events.

Polymeric gold thiolates, [Au(I)SR] n , are common synthetic intermediate precursors of gold nanoclusters and larger nanoparticles. The size and dispersity of the precursors strongly influence the properties of the synthesis products. Evaluating the size of the precursors is not straightforward because they are irregularly shaped (nonspherical) and hard to isolate from solution. Herein, we propose an effective method for determining the number of Au atoms in polymeric thiolate particles from current transients resulting from single precursor collisions, where individual [Au(I)SR] n species are electrochemically reduced at the collector ultramicroelectrode. The developed approach can lead to a better control over the mean size and dispersity of colloidal metal nanoclusters and nanoparticles.

Electrochemical experiments at individual nanoparticles (NPs) can provide new insights into their structure-activity relationships. By using small nanoelectrodes as tips in a scanning electrochemical microscope (SECM), we recently imaged individual surface-bound 10-50 nm metal NPs. Herein, we introduce a new mode of SECM operation based on tunneling between the tip and a nanoparticle immobilized on the insulating surface. The obtained current vs. distance curves show the transition from the conventional feedback response to electron tunneling between the tip and the NP at separation distances of less than about 3 nm. In addition to high-resolution imaging of the NP topography, the tunneling mode enables measurement of the heterogeneous kinetics at a single NP without making an ohmic contact with it. The developed method should be useful for studying the effects of nanoparticle size and geometry on electrocatalytic activity in real-world applications., (© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

We demonstrate the feasibility of attaining femtomolar limits of quantitation in electroanalysis. The method employed is based on electrocatalytic amplification, where small quantities of metal deposit performed on a carbon electrode causes a large increase in the observed current, for example, for the oxidation of water. We show calibration curves at the femtomolar level for cobalt, nickel, and lead ions on carbon ultramicroelectrodes (CUMEs), ca. 500 nm radii. The CUME was biased at a potential where the ion would deposit as the metal oxide, MOx, and a high concentration of species that is oxidized at the deposit is present in solution. Blips were observed in the amperometric i-t response, and their frequency scaled linearly with the concentration of ions at the femtomolar level. From these results, the limits of quantitation for cobalt, nickel, and lead ions were reported at 10 s of femtomolar level for the first time.

The loss of Pt during the oxygen reduction reaction (ORR) affects the performance and economic viability of fuel cells and sensors. Our group previously observed the dissolution of Pt nanoelectrodes at moderately negative potentials during the ORR. Here we report a more detailed study of this process and identify its product. The nanoporous Pt surface formed during the ORR was visualized by AFM and high-resolution SEM, which also showed ∼5 nm sized Pt particles on the glass surface surrounding the electrode. The release of these nanoparticles into the solution was confirmed by monitoring their catalytically amplified collisions with a Hg-coated microelectrode used as the tip in the scanning electrochemical microscope (SECM).

The production of reactive oxygen and nitrogen species (ROS and RNS) in human cells is implicated in various diseases, including cancer. Micrometer-sized electrodes coated with Pt black and platinized Pt nanoelectrodes have previously been used for the detection of primary ROS and RNS in biological systems. In this Article, we report the development of platinized carbon nanoelectrodes with well-characterized geometry and use them as scanning electrochemical microscopy (SECM) tips to measure ROS and RNS inside noncancerous and metastatic human breast cells. By performing time-dependent quantitative amperometric measurements at different potentials, the relative concentrations of four key ROS/RNS in the cell cytoplasm and their dynamics were determined and used to elucidate the chemical origins and production rates of ROS/RNS in nontransformed and metastatic human breast cells.

Coating the inner wall of a quartz nanopipet with a thin layer of carbon yields a nanopore with tunable surface charge and chemical state for resistive-pulse and rectification sensing. Herein we report the experimental study and modeling of the electron-transfer gated ion transport processes in carbon nanopipets. The potential of the unbiased carbon layer can be tuned by adding very low (sub-nM) concentrations of redox species to the solution via bipolar electrochemistry. The potential of the carbon layer determines the electrical double-layer structure that, in turn, affects the ionic transport processes. The ion current rectification decreased when redox species with a relatively positive formal potential (e.g., Fe(CN) 6 3/4- ) were added to the solution and increased upon adding redox species with a negative formal potential (e.g., Ru(NH 3 ) 6 3/2+ ). Additionally, the ion current displays high sensitivity to redox species, suggesting the possibility of trace-level analysis.

The properties of nanoparticles (NPs) are determined by their size and geometric structures. A reliable determination of NP dimension is critical for understanding their physical and chemical properties, but sizing ultrasmall particles on the order of nanometer (nm) scale in the solution is still challenging. Here, we report the size measurement of PtNP at nanometer resolution by in situ scanning electrochemical microscopy (SECM), performed with the electrochemical generation and removal of H 2 bubble at a reasonably small distance between tip and substrate electrodes in 200 or 500 mM HClO 4 solution. A series of different PtNPs or nanoclusters were electrodeposited and in situ measured in the solution, proving the concept of sizing ultrasmall particles using tip generation/substrate collection mode of SECM. This technique could be also used for investigations of other supported ultrasmall metal nanocluster systems.

Recent theoretical and experimental studies revealed strong effects of the electrical double layer (EDL) on mass transfer at nanometer-sized electrodes and in electrochemical nanogaps. Although the EDL effect is much stronger in weakly supported media, it can significantly influence the kinetics of electron-transfer processes involving multicharged ionic redox species, even at high concentrations of supporting electrolyte. We measured the kinetics of Fe(CN) 6 4- oxidation in 1 M KCl solution at the Pt nanoelectrode used as a tip in the scanning electrochemical microscope. The apparent standard rate constant values extracted from tip voltammograms without double-layer correction increased markedly with the decreasing separation distance between the tip and substrate electrodes. The same steady-state voltammograms were fitted to the theory including the EDL effect and yielded the rate constant essentially independent of the separation distance.

Investigating the collisions of individual metal nanoparticles (NPs) with electrodes can provide new insights into their electrocatalytic behavior, mass transport, and interactions with surfaces. Here we report a new experimental setup for studying NP collisions based on the use of carbon nanopipettes to enable monitoring multiple collision events involving the same NP captured inside the pipet cavity. A patch clamp amplifier capable of measuring pA-range currents on the microsecond time scale with a very low noise and stable background was used to record the collision transients. The analysis of current transients produced by oxidation of hydrogen peroxide at one IrO x NP provided information about the origins of deactivation of catalytic NPs and the effects of various experimental conditions on the collision dynamics. High-resolution TEM of carbon pipettes was used to attain better understanding of the NP capture and collisions.

Along with more prevalent solid-state nanopores, glass or quartz nanopipettes have found applications in resistive-pulse and rectification sensing. Their advantages include the ease of fabrication, small physical size and needle-like geometry, rendering them useful for local measurements in small spaces and delivery of nanoparticles/biomolecules. Carbon nanopipettes fabricated by depositing a thin carbon layer on the inner wall of a quartz pipette provide additional means for detecting electroactive species and fine-tuning the current rectification properties. In this paper, we discuss the fundamentals of resistive-pulse sensing with nanopipettes and our recent studies of current rectification in carbon pipettes.

Steady-state voltammetry at nanoelectrodes and scanning electrochemical microscopy (SECM) have recently been used to measure kinetics of several rapid heterogeneous electron transfer (ET) reactions. One problem with those experiments was that the dependence of the shape of the steady-state voltammogram on kinetic parameters becomes weak when the reaction rate approaches the diffusion limit. The possibility to fit the same experimental voltammogram using different combinations of the standard rate constant, transfer coefficient, and standard potential results in significant uncertainties in extracted parameter values. In this article, the reliability of the kinetic analysis was improved by obtaining steady-state voltammograms with both oxidized and reduced forms of redox species initially present in solution. Additional improvements were attained by characterizing the nanoelectrode geometry with the atomic force microscope and using water with a very low level of organic contaminants (TOC ≤ 1 ppb). This approach was used to re-evaluate the ET rate constants measured for several electroactive species, including ferrocene, ferrocenemethanol, 7,7,8,8-tetracyanoquinodimethane (TCNQ), and ferrocyanide at Pt electrodes. The obtained standard rate constants are higher than the values measured earlier at Pt and Au nanoelectrodes but comparable to those obtained in recent nanogap/SECM experiments.

Electrochemistry at metal nanoparticles (NPs) is of significant current interest because of its applications in catalysis, energy conversion and storage, and sensors. The electrocatalytic activity of NPs depends strongly on their size, shape, and surface attachment. The use of a large number of particles in most reported kinetic experiments obscured the effects of these factors because of polydispersity and different NP orientations. Recent efforts to probe electrochemistry at single NPs included recording of the catalytically amplified current produced by random collisions of particles with the electrode surface, immobilizing an NP on the surface of a small electrode, and delivering individual NPs to electrode surfaces. Although the signals recorded in such experiments were produced by single NPs, the characterization issues and problems with separating an individual particle from other NPs present in the system made it difficult to obtain spatially and/or temporally resolved information about heterogeneous processes occurring at a specific NP. To carry out electrochemical experiments involving only one NP and characterize such an NP in situ, one needs nanoelectrochemical tools with the characteristic dimension smaller than or comparable to those of the particle of interest. This Account presents fundamentals of two complementary approaches to studying NP electrochemistry, i.e., probing single immobilized NPs with the tip of a scanning electrochemical microscope (SECM) and monitoring the collisions between one catalytic NP and a carbon nanopipette. The former technique can provide spatially resolved information about NP geometry and measure its electron transfer properties and catalytic activity under steady-state conditions. The emphasis here is on the extraction of quantitative physicochemical information from nanoelectrochemical data. By employing a polished disk-type nanoelectrode as an SECM tip, one can characterize a specific nanoparticle in situ and then use the same NP for kinetic experiments. A new mode of SECM operation based on tunneling between the tip and nanoparticle can be used to image the NP topography with a lateral resolution of ∼1 nm. An alternative approach employs carbon nanoprobes produced by chemical vapor deposition of carbon into quartz nanopipettes. One metal NP is captured inside the carbon nanocavity to probe the dynamics of its interactions with the electrode surface on the microsecond time scale. The use of high-resolution transmission electron microscopy is essential for interpreting the results of single-NP collision experiments. A brief discussion of the nanoelectrochemical methodology, recent advances, and future directions is included.

Electroreduction of diazonium salts is a widely used technique for grafting organic films on various surfaces. In this paper, scanning electrochemical microscopy (SECM) was used for high-resolution characterization of a thiolated aryl multilayer film obtained by electrografting of thiophenol diazonium on highly ordered pyrolytic graphite (HOPG). The blocking properties of the film were evaluated, and the origins of incomplete surface passivation were elucidated by comparing current-distance curves and surface reactivity maps obtained with nanometer- and micrometer-sized tips. In this way, one can distinguish between different pathways of charge transport in the film, e.g., pinhole defects versus rate-limiting charge transfer through the film. Pd nanocubes were anchored to the film by thiol groups and imaged by SECM. The applicability of SECM to in situ visualization of the geometry of non-spherical nanoparticles has been demonstrated.

We report on imaging of local electric field on an electrode surface with plasmonic electrochemical impedance microscopy (P-EIM). The local electric field is created by putting an electrode inside a micropipet positioned over the electrode and applying a voltage between the two electrodes. We show that the distribution of the surface charge as well as the local electric field at the electrode surface can be imaged with P-EIM. The spatial distribution and the dependence of the local charge density and electric field on the distance between the micropipet and the surface are measured, and the results are compared with the finite element calculations. The work also demonstrates the possibility of integrating plasmonic imaging with scanning ion conductance microscopy (SICM) and other scanning probe microscopies.

Nanoscale scanning electrochemical microscopy (SECM) has emerged as a powerful electrochemical method that enables the study of interfacial reactions with unprecedentedly high spatial and kinetic resolution. In this work, we develop carbon nanoprobes with high electrochemical reactivity and well-controlled size and geometry based on chemical vapor deposition of carbon in quartz nanopipets. Carbon-filled nanopipets are milled by focused ion beam (FIB) technology to yield a flat disk tip with a thin quartz sheath as confirmed by transmission electron microscopy. The extremely high electroactivity of FIB-milled carbon nanotips is quantified by enormously high standard electron-transfer rate constants of ≥10 cm/s for Ru(NH 3 ) 6 3+ . The tip size and geometry are characterized in electrolyte solutions by SECM approach curve measurements not only to determine inner and outer tip radii of down to ~27 and ~38 nm, respectively, but also to ensure the absence of a conductive carbon layer on the outer wall. In addition, FIB-milled carbon nanotips reveal the limited conductivity of ~100 nm-thick gold films under nanoscale mass-transport conditions. Importantly, carbon nanotips must be protected from electrostatic damage to enable reliable and quantitative nanoelectrochemical measurements.

Molecular grafting of diazonium is a widely employed surface modification technique. Local electrografting of this species is a promising approach to surface doping and related properties tailoring. The instability of diazonium cation complicates this process, so that this species was generated in situ in many reported studies. In this Article, we report the egress transfer of aryl diazonium cation across the liquid/liquid interface supported at the nanopipette tip that can be used for controlled delivery this species to the external aqueous phase for local substrate patterning. An aryl diazonium salt was prepared with weakly coordinating and lipophilic tetrakis(pentafluorophenyl)borate anion stable as a solid and soluble in low polarity media. The chemically stable solution of this salt in 1,2-dichloroethane can be used as "diazonium ink". The ink-filled nanopipette was employed as a tip in the scanning electrochemical microscope (SECM) for surface patterning with the spatial resolution controlled by the pipette orifice radius and a few nanometers film thickness. The submicrometer-size grafted spots produced on the HOPG surface were located and imaged with the atomic force microscope (AFM).

Experiments at individual metal nanoparticles (NPs) can provide important information about their electrochemical and catalytic properties. The scanning electrochemical microscope (SECM) equipped with a nanometer-sized tip was recently used to image single 10 or 20 nm gold particles and quantitatively investigate electrochemical reactions occurring at their surfaces. In this Article, the theory is developed for SECM current vs distance curves obtained with a disk-shaped tip approaching a comparably sized, surface-bound conductive or insulating spherical NP. The possibility of evaluating the size of a surface-bound particle by fitting the experimental current-distance curve to the theory is shown for NPs and tips of different radii. The effects of the NP being partially buried into an insulating layer and the imperfect positioning of the tip with respect to the NP center are considered. The collection efficiency is calculated for redox species generated at the nanoparticle surface and collected at the tip.

Quartz nanopipettes have recently been employed for resistive-pulse sensing of Au nanoparticles (AuNP) and nanoparticles with bound antibodies. The analytical signal in such experiments is the change in ionic current caused by the nanoparticle translocation through the pipette orifice. This paper describes resistive-pulse detection of cancer biomarker (Vascular Endothelial Growth Factor-C, VEGF-C) through the use of antibody-modified AuNPs and nanopipettes. The main challenge was to differentiate between AuNPs with attached antibodies for VEGF-C and antigen-conjugated particles. The zeta-potentials of these types of particles are not very different, and, therefore, carefully chosen pipettes with well-characterized geometry were necessary for selective detection of VEGF-C.

Development of better catalysts for the oxygen reduction reaction (ORR) and other electrocatalytic processes requires detailed knowledge of reaction pathways and intermediate species. Here we report a new methodology for detecting charged reactive intermediates and its application to the mechanistic analysis of ORR. A nanopipette filled with an organic phase that is immiscible with the external aqueous solution was used as a tip in the scanning electrochemical microscope to detect and identify a short-lived superoxide (O2(●-)) intermediate and to determine the rate of its generation at the catalytic Pt substrate and its lifetime in neutral aqueous solution. The voltammogram of the O2(●-) anion transfer to the organic phase provides a unique signature for unambiguous identification of superoxide. The extremely short attainable separation distance between the pipette tip and substrate surface (∼1 nm) makes this technique suitable for detecting and identifying charged intermediates of catalytic processes with a lifetime of a few nanoseconds.

Unlike macroscopic and micrometer-sized solid electrodes whose surface can be reproducibly cleaned by mechanical polishing, cleaning the nanoelectrode surface is challenging because of its small size and extreme fragility. Even very gentle polishing typically changes the nanoelectrode size and geometry, thus, complicating the replication of nanoelectrochemical experiments. In this letter, we show the possibility of cleaning nanoelectrode surfaces nondestructively by using an air plasma cleaner. The effects of plasma cleaning have been investigated by atomic force microscopy (AFM) imaging, voltammetry, and scanning electrochemical microscopy (SECM). A related issue, the removal of an insoluble organic film from the nanoelectrode by plasma cleaning, is also discussed.