Design and elaboration of original objects by and for electrochemistry
- Porous electrodes
- Chiral materials
- Surface gradients with tailored properties
- Janus particles
- Porous electrodes
Electrochemical signals are very often limited in terms of current or power output by the active surface area of the electrode. One way to circumvent such limitations is to employ porous electrodes. The activities within this topic are focused on the development of macro- and mesoporous electrodes with a high structural control. We follow an approach using either beads or supramolecular surfactant assemblies as templates during the electrodeposition of a wide range of materials, from different metals  or metal oxides  to metal organic frameworks and conducting polymers. This allows a perfect control of the organization of the pores and leads to electrodes with an active surface area that is two or three orders of magnitude higher compared to flat surfaces. The internal surface of these designer electrodes can be subsequently modified with catalysts, redox-mediators and enzymes for a variety of applications ranging from photocatalysis to (bio)sensors[6,7] and (bio)fuel cells.[8,9]
SEM view of a macro/mesoporous electrode
- Chiral materials
Chirality plays an extremely important role in biological systems, such as encountered for antigen-antibody interactions or enzyme-substrate recognition. A lot of effort has been made to mimic such natural systems by designing chemical analogous. We’ve developed an important expertise in the controlled generation of mesoporous metal structures which retain at their internal surface chiral information by imprinting the structure of chiral molecules in the walls of the mesopores. Our experiments show that such electrodes not only can distinguish between two enantiomers when they are used as electroanalytical tools [17, 18], but, most importantly, are also able to generate an enantiomeric excess of over 90%, when they are employed for electrosynthesis [19, 20]. Such a selectivity based on a purely geometric effect holds great promise to impact considerably not only the field of electrochemistry, but also materials science and physical chemistry in general. Our current efforts within this topic are focused on the optimization of the imprinting process, the efficiency of the molecular recognition and the range of materials which can be used as a matrix.
Scheme of the chiral recognition on an imprinted metal surface
- Surface gradients with tailored properties
The elaboration of surfaces with specific anisotropic features is a very hot contemporary topic of research. The different possible methods of fabrication need to break the symmetry of the initial isotropic surface. In this context, bipolar electrochemistry (BPE) can be employed for the controlled generation of directional physico-chemical gradients based on the polarization established alongside bipolar electrodes. For that, several metal layers exhibiting a variation of chemical composition, morphology and/or roughness are electrodeposited on conducting substrates with various geometries. Also, the formation of a tunable molecular gradient can be achieved by partial removal of a homogeneous layer under BPE conditions. The combination of electrode surface micro-structuration with such molecular gradients results in original wetting properties .
- Janus particles
Asymmetry is a very common feature of many systems, objects and molecules, that we encounter and use in our daily life. Actually, it is in a majority of cases the absolutely crucial ingredient for conferring a certain useful property to a system. In this context so-called Janus particles bearing an intrinsic asymmetric property are interesting model systems for breaking the symmetry at the level of micro- and nanoobjects. Various methods are described in the literature for synthesizing such particles, but most of them need an interface or a surface to break the symmetry. We explore alternative approaches allowing symmetry breaking in the bulk phase of a solution. One straight-forward way to achieve this is based on bipolar electrochemistry. Bipolar electrochemistry is a concept with a quite long history, but has only very recently revealed its virtues in many areas of chemistry, among others for the controlled surface modification at the micro- and nanoscale. We largely extended this concept in the frame of material science applications, especially by developing direct and indirect bipolar electrodeposition, eventually assisted with light, as a simple route to particles with a very sophisticated design [24, 25, 26].
In order to modify insulating objects, such as silica particles, bipolar electrochemistry cannot be used but we are exploring simultaneously alternative approaches. One of them is based on the concept that a removable mask temporarily protects a part of an object in a reactive medium. This is for example possible by carrying out an emulsion polymerization of styrene in the presence of silica particles. Parts of the silica particle will be covered by polymer, thus allowing in a second step the specific functionalization of the uncovered silica surface with a trialkoxysilane derivative. After dissolving the polymer, the unprotected zone of the silica surface can be specifically functionalized in order to introduce the final asymmetry.
Another strategy leading to Janus objects explored by us is based on the use of colloidal microgel particles which can adsorb at an oil-water interface and thus stabilize so-called Pickering emulsions. The microgels which are decorating the surface of the oil droplets intrinsically experience two different chemical environments and thus can be modified selectively on one side. The so-obtained asymmetric objects can carry out, among others, interesting functions such as light emission, catalysis and controlled propulsion.
Scheme of light-assisted bipolar electrodeposition of metals on semiconductor micro- and nanoparticles
For controlled propulsion asymmetry is the crucial ingredient and we have explored various concepts leading to so-called microswimmers. In all cases bipolar electrochemistry plays the key role as it is by definition well-suited for symmetry breaking.
Essentially there are two different ways of using bipolar electrochemistry for generating controlled motion:
(1) Janus particles generated by bipolar electrodeposition containing a magnetic or a catalytic extremity can be used as synthetic motors [27, 28]. We have shown that carbon microtubes with an electrochemically generated Pt tip on one side can move in H2O2 solutions. The motion was caused by the release of O2 bubbles, due to the catalytic decomposition of H2O2 at the Pt surface.
(2) The asymmetric reactivity, offered by bipolar electrochemistry, can also be used directly to generate motion. The first example of a translational motion was based on a deposition/dissolution mechanism. In this work, a Zn dendrite was placed in capillary that was previously filled with a zinc sulfate solution. When the dendrite is acting as a bipolar electrode, its anodic pole is consumed by the oxidation, while deposition occurs at its cathodic pole, leading to its self-regeneration. This results in an apparent locomotion of the object. Another example is based on asymmetric bubble production, due to water electrolysis at the reactive poles of a spherical bipolar electrode. Because the quantity of produced H2 at the cathodic pole is twice as much as the amount of generated O2 at the anodic pole, the resulting force rolls the bead in a controlled way.[30, 31] A possibility to enhance the propulsion speed and to better control its direction is to quench one of the bubble producing reactions, by adding to the solution a sacrificial molecule (which is easier to oxidize or to reduce than water), thus allowing the bubble production to occur only at one bead pole. A completely different type of motion triggered by bipolar asymmetric reactivity is exemplified with conducting polymer objects. The oxidation and reduction occurring at the two extremities of such objects when they are exposed to the electric field leads to differential swelling and shrinking due to the uptake and release of counter ions. This ultimately results in a deformation of the polymer and can be transformed into controlled motion [32,33].
Deformation and motion of a polypyrrole object triggered by bipolar electrochemistry
Studying the synergy between photons and electrons
- Electrogenerated chemiluminescence
- Electrochemical microscopy and photoelectrochemistry
- Coupling electrochemistry and fluorescence microscopy
- Nanostructured optical fibers
- Electrogenerated chimiluminescence
Electrogenerated ChemiLuminescence (ECL) is the light emission of a luminophore resulting from an initial electrochemical step. It combines thus intimately electrochemistry and photochemistry. Due to its intrinsic characteristics, ECL is a powerful analytical technique and it is widely commercialized for immunoassays and clinical diagnostics by several companies. Our goal is to decipher the ECL mechanistic pathways and, based on this fundamental knowledge, to propose original (bio)analytical applications. For this, we develop a holistic approach at the differetn scales to push back the limits of ECL:
(1)at the molecular scale: discovery and study of new efficient ECL luminophores [1,2,3]
(2) at the nanometer scale: stimuli-responsive hydrogel nanomaterials (nanoparticles and thin films)[4,5,6] and nanoelectrodes[7,8] are exploited to miniaturize analytical systems
(3) at the micrometer scale: immunoassays,[8,9] analytical (bio)swimmers and 3D ECL,[10,11,12] and cell imaging
(4) at the macroscale: to develop integrated (bio)analytical ECL-based platforms[14,15]
Potential applications : bioassays, cell microscopy, ECL imaging, ultrasensitive detection
From left to right : ECL luminophores, stimuli-responsive NPs, bead-based ECL immunoassays, cell imaging, (bio)swimmers developed by the synergy between ECL and bipolar electrochemistry, microelectrode arrays.
Electrochemical microscopy and photoelectrochemistry
Scanning Electrochemical Microscopy (SECM) has been first described in the early 90’s and is nowadays widely used to analyze interfacial phenomenon. The principle of SECM is to use a microelectrode, that can move in the three directions of space to study a substrate. The electrochemical response of the tip is disturbed by the substrate, providing information about its nature and its properties. SECM recently find applications in the field of solar energy, by coupling the microelectrode with an optical fiber, it is called scanning photoelectrochemical microscopy (SPECM). We used the SPECM to rapidly screen the photoelectrochemical properties of TiO2 spots prepared in different conditions. We simultaneous measured the water oxidation photocurrent and the reduction current of the produced oxygen. By placing the optoelectrode at a constant distance and by scanning the surface in the X-Y direction, an image was obtained, allowing to observe the most efficient photocatalyst.
Potential applications : Chemical surface patterning, rapid screening of photocatalysts
a) Scheme of local generation and screening of TiO2 by SECM; b) Photoelectrochemical image of different type of TiO2 generated by local anodization.
Coupling electrochemistry and fluorescence microscopy
Nowadays, the coupling between electrochemistry and fluorescence microscopy is gaining a considerable interest. The collection of both analytical signals provides simultaneously a space- and time-resolved information, thus enabling the detection of spatial or temporal heterogeneities of electrode processes. We have implemented fluorescence confocal laser scanning microscopy (FCLSM) under electrochemical control to map in situ various reaction layers in 3D (xy plane at different z positions with respect to the electrode surface). The main objectives are: (1) the study of the (electro)chemical reactivity of biorelevant fluorogenic dyes used as analytical probes in medicinal/clinical diagnostics and (2) the control and/or improvement of the coverage homogeneity of modified electrodes used as biosensing platforms.
Fluorescence intensity mapping along the z-direction perpendicular to the electrode surface collected by confocal laser scanning microscopy for an extinction (left) or activation of fluorescence (right) upon electron transfer
Nanostructured optical fibers for imaging and analysis
Optical fiber bundles are well-established tools in analytical chemistry for sensing and imaging. For instance, we fabricated high density nanoprobe arrays[1,2] for DNA detection and for remote imaging of the human skin (corneocytes) by wet etching optical fiber bundles. Such bundles are composed of thousands of micrometric optical fibers arranged in a coherent optical way (for example: 6000 optical fibers with a 3µm-core diameter assembled in a 350µm bundle). The bundles are micro/nanostructured by well-controlled wet etching with regular 3D patterns such as nano-tips, micro/nano-wells, micro-pillars, micro/nano-beads or more complex shapes[3,4] can also be achieved. The resulting optical nanostructures keep the initial architecture of the bundle and therefore its intrinsic imaging properties. Remote imaging and detection based on fluorescence, ECL, Surface-Enhanced Raman Scattering (SERS), Fluorescence Correlation Spectroscopy (FCS), Surface Plasmon Resonance (SPR) were demonstrated through the bundle itself.[5,6] We also developed with an industrial partner (L’Oréal) a remote imaging technique of living corneocytes directly on the human skin.
a) SERS with etched nanotip. b) Remote imaging of corneocytes on the forearm. c) Fluorescence image of the DNA spots electropolymerized on the nanotip array.
Development of sensors and analytical methods
- Electrochemical and optical microsystems to study mitochondria
- Biosensing and bioenergy
- Sensing in complex liquid media
- Particles for drug delivery
Electrochemical and optical microsystems to study mitochondria
Mitochondria play major roles into diverse metabolic pathways (oxidative phosphorylation, calcium handling, redox signalling) and when defective, they lead to severe pathologies (myopathies, neurological disorders…). Consequently, novel methodologies are required to decipher mitochondrial activities and provide diagnosis tools. We are developing microsystems integrating electrochemical or optical sensors, or both combined in a single device. Electrochemical systems are based on microelectrode or nanoelectrode arrays. These allow to monitor the oxygen consumption or H2O2 production by mitochondria in response to OXPHOS activators and inhibitors. Optical systems are based on glass, PDMS and SiO2 substrates with microwell structures in which small populations of mitochondria are deposited. Mitochondria are monitored individually by fluorescence microscopy based on NADH, calcium or membrane potential dies. Overall, these different microsystems offer unprecedented resolution to assess single mitochondria activities, hetereogenity and dysfunctions in physio-pathological processes.
Left: Optical and electrochemical microsystems developed for the monitoring of mitochondrial activities. Right: example of simultaneous detections of oxygen consumption and hydrogen peroxide release by isolated mitochondria.
Biosensing and Bioenergy
Different electrode architectures are developed and used for biosensing and biofuel cells. The challenge is miniaturization while reaching high analytical performances. For biosensors, we design electrochemical and combined electrochemical/optical sensors, which dimensions and structuration range from micro- to nanoscale. These ones are adapted to the size of the biological entity under study (tissue, single cell, membrane, isolated DNA). Individual microelectrodes are placed next to a neuro-secretory or cardiac cell to measure local fluxes of catecholamines, oxygen or hydrogen peroxide. This approach is combined to optical sensing in micro-optoelectrodes (see topic 2) to observe the biological objects at the sensor tip while detecting locally redox activities. For biofuel cells, the principle is to draw energy from physiological fluids to power biomedical devices. We develop micro-biofuel cells, in which the two electrodes are modified with enzymes: at the anode glucose is oxidized while oxygen is reduced at the cathode. Recently we have integrated such electrodes into an electronic chip designed to be powered by this miniaturized energy source. This work suggests a number of biomedical applications including skin-implanted glucose sensors.
Left: Electrochemical detection by a microelectrode on a single cardiac cell; Right: SEM image of a coaxial electrochemical cell.
Sensing in complex liquids
Our development of electrochemical sensors for small molecules involved in redox processes of living systems has led to diverse applications based on measurements in complex aqueous media, such as blood, skin hydrolipidic film or beverages. In particular, we are developing electrodes sensitive to antioxidants or reducing species (ascorbic acid, anthocyans …). Such species have been measured in usual beverages and especially in wines. Also, sulfur dioxide in its dissolved and chemically unbound forms is detected in wines since this species is the main antioxidant and antifungal chemical compound used in wine making. Measuring sulfites in such complex matrices is an analytical challenge and of high interest for wine industry. Dedicated modified electrodes and detection protocols have been developed and used successfully to monitor sulfite and pH in wines along different phases of wine making. This work is currently pursued in order to monitor other reducing agents used as food preservatives.
Development of methodologies and sensors for measuring reducing compounds in complex media, such as wine.
Particles for drug delivery
Hydrogels are biocompatible materials due to their high hydration degree. These porous structures can be used as drug delivery vehicles. Controlled released can be achieved by application of a stimulus or by degradation of the polymer matrix. In this purpose, we develop functionalized polysaccharide-based hydrogels and nanogels for the encapsulation of proteins and peptides as well as hydrophobic hormones.
Enzyme-triggered release of progesterone-loaded oil nanodroplets from a hyaluronic acid microgel
Synthesis and assembly of functional particles
- Responsive nanogels for sensing and drug delivery
- Capsules, emulsions and interfaces
- Artificial cells
- Self-assembly of polymers
- Films and materials based on colloidal particles
- Responsive nanogels for sensing and drug delivery
Nanogels and microgels are colloidal particles made of swollen cross-linked polymers. In aqueous solution, these soft particles behave as biomimetic responsive materials, which undergo reversible volume phase transitions in response to changes in their environment. Our group develops different elaboration techniques such as batch synthesis, miniemulsion or microfluidic approach to prepare original architectures based on synthetic or bio-based polymers. Our main objectives are:
1) the development of (bio)sensors, where the swelling degree of nanogels depends on the recognition of the (bio)molecule. Modulation of the swelling degree induces a change in a physical signal (color, luminescence , electrochemical signal ;
2) the release of an active molecule that has been previously encapsulated in the porous structure; they can be used as smart drug delivery systems ;
3) coupling both sensing and delivery properties lead to closed-loop drug delivery systems. An example is glucose-responsive nanogels, to be used in closed-loop delivery of insulin for type 1 diabetes .On a fundamental point of view, we study the effect of volume phase transition on electrochemical processes.
Representation of a responsive nanogel and examples of nanogel structures :
a) simple nanogel with dangling chains; b) Core-shell nanogel with silica core; c) Hollow nanogel.
Capsules, emulsions and interfaces
Microgels/Nanogels adsorb at liquid interfaces and can be used as stabilizers for emulsionsand foams. Because they are responsive, the resulting formulations can be destabilized on demand, upon application of a stimulus. Moreover, nanogels are soft particles, which appear to be deformed at the interface, being either flattened or compressed. We have shown that the deformability is an important criterion to control formulation stability and its rheological properties. In this topic, we are interested in two aspects:
(1) understanding the role of nanogel structure on their interfacial properties and the consequences on emulsion properties;
(2) preparing new materials, in particular capsules, from these assemblies. These capsules may be purely organic or hybrid organic/inorganic (gold, silica), responsive, and have a controlled thickness and porosity.
Applications: Responsive emulsions and foams for drug delivery, coatings.
Representation of a drop stabilized by nanogels (a) and confocal microscopy image (b); Close view of the drop surface covered with compressed nanogels (obtained by cryoSEM) (c).
The building of an "artificial cell" has become a major goal of biophysical chemistry. Within this field, our objective is to develop biomimetic microreactors for the study of biochemical or biological processes devoted to redox reactions, in particular to the synthesis of Reactive Oxygen - Nitrogen Species (ROS, RNS). By confining reactions within a cell-sized compartment (10-100 µm diameter), reactive species (H2O2, NO•, O2•-) can be produced and analyzed in situ with a quantitative and kinetic resolutions. Giant unilamellar vesicles (GUVs) made of phospholipids and fatty acids are used as a biomimetic reactor for the monitoring of glucose oxidase (GOX), NADPH oxidase (NOX) and NO-Synthase activities. Fluorescence microscopy allows individual vesicle observation and the monitoring of reactions triggered by microinjection. Then, released species can be detected by electrochemistry in order to decipher on the enzymatic kinetics and pathways (ex. NO-Synthases). A single ultramicroelectrode is placed at the vicinity of the microreactor membrane and species diffusing through lipidic bilayer, including H2O2 and NO• are measured. Current developments target autonomous microreactors with controlled transport of substrates and products of biological reactions.
Principle of a biomimetic microeactor wherein biochemical and biological reactions can be achieved
and analyzed in order to build up an artificial cell..
Controlling the self-assembly of elementary brick such as polymer chains is a power tool for the conception of smart materials. Our group focuses on the design of complex stimuli sensitive polymeric assemblies like self-healing hydrogels or coacervates, allowing the sequestration and/or the delivery of chemical compounds. Two main approaches are explored by introducing attractive interactions such as:
- Dynamic covalent coupling by using boronate/diol interactions 
- Electrostatic coupling with the design of coacervates 
Scheme of polymeric self-assembly
Films and materials based on colloidal particles
As a prerequisite for generating more complex architectures with tailored properties, it is a key point to master the assembly of particles on various substrates. Among many techniques, our lab has a very strong expertise in Langmuir-Blodgett and electrophoretic deposition processes. These techniques are powerful tools to generate colloidal assemblies of a very precise thickness over a large range of substrates, whatever their size (from the micro to macroscale) and geometry (planar, cylindrical or more complex ones). In a second step, it is possible to use these colloidal assemblies as molds for obtaining replicas made of metals, conductive or insulating polymers.
Polymeric materials with embedded layers of beads or pores with a controlled size.
Neso Sojic, Stéphane Arbault, Dodzi Zigah, Valérie Ravaine, Alexander Kuhn, Laurent Bouffier, Bertrand Goudeau, Patrick Garrigue
This thematic is focused on the development of new tools and methods for bioanalytical chemistry and nano-imaging by combining optics, luminescence and electrochemistry. Our activities in this field are organized as follow:
1. Optical Tools for Imaging & Enhanced Coupled Methodologies
2. Electrogenerated Chemiluminescence Imaging
3. Electrochemical Imaging
High density nanoprobe arrays for DNA detection and for SERS nano-imaging are fabricated by wet etching optical fiber bundles. The resulting optical nanostructures keep the initial architecture of the bundle and therefore its intrinsic imaging properties. Remote SERS imaging was demonstrated through the bundle itself in collaboration within the ISM (group of molecular spectroscopy). We also developed with an industrial partner a remote imaging technique of living corneocytes directly on the human skin (L’Oréal).
a) SERS with etched nanotip. b) Remote imaging of corneocytes on the forearm. c) Fluorescnece image of the DNA spots electropolymerized on the nanotip array.
Understanding mitochondrial metabolic status at the single organelle level is of major interest for several bio-medical fields, since there is a genetic and metabolic heterogeneity within the mitochondrial pool-network of each cell. In this context, we developed microwell arrays for fluorescence microscopy of single isolated mitochondria. We are working on the multiparametric monitoring of mitochondrial metabolic state using fluorescent dyes or endogenous fluorescent metabolites. Our goal is focused on the understanding of the kinetic relationships between mitochondrial membrane potential fluctuations and reactive oxygen and nitrogen species formation. We combine also such fluorescence microscopy approaches with miniaturized electrochemical sensors (see also the webpage Electrochemistry of Biosystems).
Monitoring of NADH fluorescence variations in individual mitochondria (from Saccharomyces cerevisiae) under different stages
Stage 1: resting state of mitochondria; Stage 2: mitochondria energized by injecting ethanol 1% in the well; Stage 3: mitochondrial respiration inibited by injecting Antimycin A, a usual inhibitor of complex III in the respiratory chain.
Electrogenerated ChemiLuminescence (ECL) is the luminescence emitted by a luminophore resulting from an initial electron transfer reaction occurring at the electrode surface. In other words, it combines intimately electrochemistry and photochemistry. Immunoassays based on ECL are widely commercialized. We developed a novel photopatterning method for immobilisation of ECL ultrathin films. ECL hydrogel films were immobilised in form of uniform photopatterns which size, shape and thickness were modulated depending on the fabrication parameters. The design and implementation of a new class of sensing microarrays were also developed. ECL was used as a readout mechanism to detect multiple antigens simultaneously. The method enables multiplexed assays because all the individual sensing beads in the array are simultaneously imaged by ECL.
a) ECL images of different patterns of nanometer-thin films obtained on gold electrode. b) Platform for multiplexed sandwich immunoassays: antigens detection with ECL bead-based microarray.
Particularly fascinating nanomaterials are stimuli-responsive hydrogel particles, or microgels. The properties of such so-called “smart” microgels are modulated by an external stimulus, which triggers expansion or contraction of the polymer network, at the origin of sensing capabilities (see the webpage Chemical Sensors for Biology). Through a rational choice of model ECL and microgel systems, we studied electrochemistry and ECL of thermo-responsive microgels. We demonstrated an unexpected enhancement of the ECL signal which occurs at the swell-collapse transition of the microgel particles.
Carbon nanotubes (CNTs) have emerged recently as a powerful analytical tool with a wide range of electronic and mechanic properties. For instance, in the context of electrochemical biosensors, CNTs are used to structure electrodes and increase the corresponding active surface. We are now developing an original approach based on tubes which are selectively modified on one side by bipolar electrochemistry, in order to generated unsymmetrical systems with promising analytical potential and improved detection capability. In order to locally modify this carbon tube an original approach using bipolar electrochemistry and diazonium salts reduction is used. The electroreduction of diazionium salts allow us to immobilize an organic layer using a covalent bonding. This organic layer can be used to develop analytical system.
a) Schematic representation of the thermo-responsvie ECL microgels in the swollen (left) and collapsed (right) states; b) Transmission electron microscopy image of dried microgels; c) Asymmetric light-emitting swimmer. The synergetic reduction of H2O at the cathodic pole and oxidation of the ECL reagents at the anodic pole induces simultaneous motion and light emission of the bead in a capillary.
Bipolar electrochemistry, that usually has been employed in our group to modify objects in an asymmetric way (see paragraph on Janus particles on the webpage Smart and Dynamic Particles) can also be applied to the simultaneous bubble production and ECL generation on both poles of a conducting bead. It leads to the first example of a propulsion mechanism for a swimmer that is coupled with a chemical light source. In this case ECL provides a direct monitoring of the motion, which could be very useful for localizing micromotors. Changing the chemistry occurring at the surface of the moving object allows tuning the emitted colour, for example, using the classic luminol/H2O2 reaction leads to a blue light emitting swimmer.
Scanning Electrochemical Microscopy (SECM) has been first described in the early 90’s and is nowadays widely used to study interfacial phenomenon. By scanning an ultramicroelectrode (UME) above a surface, a three-dimensional image can be obtained. SECM can be used in order to obtain a topographic or/and chemical imaging, for example it is possible to observe the location of enzyme sites in a membrane, or to identify areas with different reactivity on a surface. For example, we studied a polypyrrole layer deposited on gold electrode by bipolar electrochemistry. Thanks to the SECM we proved that this layer presented various reactivity depending of its deposition potential.
a) Ultramicroelectrode and its diffusion layer close to the substrate; b) Substrate: polypyrrole on gold; c) SECM image, the high current (in red) corresponding to the gold layer.