AbstractCyclic multiredox centered systems are currently of great interest, with new compounds being reported and developments made in understanding their behavior. Efficient, elegant, and high-yielding (for macrocyclic species) synthetic routes to two novel alkynyl-conjugated multiple ferrocene- and biferrocene-containing cyclic compounds are presented. The electronic interactions between the individual ferrocene units have been investigated through electrochemistry, spectroelectrochemistry, density functional theory (DFT), and crystallography to understand the effect of cyclization on the electronic properties and structure.

AbstractCyclic multiredox centered systems are currently of great interest, with new compounds being reported and developments made in understanding their behavior. Efficient, elegant, and high-yielding (for macrocyclic species) synthetic routes to two novel alkynyl-conjugated multiple ferrocene- and biferrocene-containing cyclic compounds are presented. The electronic interactions between the individual ferrocene units have been investigated through electrochemistry, spectroelectrochemistry, density functional theory (DFT), and crystallography to understand the effect of cyclization on the electronic properties and structure.

Cu extraction from chalcopyrite ores is typically a slow process that involves aggressive chemical reagents with significant environmental impact. Ionic liquids (IL) have been proposed as a potentially more benign solution, but the sheer number of IL variants complicates the search for the most efficient solvent systems. Here, we present an automated electrochemical platform that allows for screening of 180 and more leaching samples in parallel with minimal solvent consumption. In a proof-of-concept study, we screen 25 samples with different IL and water contents, and find two orders of magnitude difference in leaching performance within this array. The best performing system is then applied in a tank leaching configuration, with real-time electrochemical monitoring of Cu evolution in solution. All electrochemical data is found to be in excellent agreement with off-line ICP-AES data.

We present a high-speed electrical detection scheme based on a custom-designed CMOS amplifier which allows the analysis of DNA translocation in glass nanopipettes on a microsecond timescale. Translocation of different DNA lengths in KCl electrolyte provides a scaling factor of the DNA translocation time equal to p = 1.22, which is different from values observed previously with nanopipettes in LiCl electrolyte or with nanopores. Based on a theoretical model involving electrophoresis, hydrodynamics and surface friction, we show that the experimentally observed range of p-values may be the result of, or at least be affected by DNA adsorption and friction between the DNA and the substrate surface.

•RuCl2(diphos)2 family extended to include coordinatively unsaturated PP3 ligands.•Pendant –PPh2 moieties available for further synthetic modification.•cis-RuCl2(PP3) can react with ethynylferrocene to form butenylnyl species.•Characterization includes solution voltammetry studies and X-ray crystallography.Trans-RuCl2(PP3)2 (1a) (PP3 = tris[2-(diphenylphosphino)ethyl]phosphine) was prepared by reaction of RuCl2(PPh3)3 with 2 eq. PP3. Through coordination of two potentially tetradentate ligands in a bidentate arrangement, four uncoordinated phosphine moieties remain readily available for subsequent reaction. This is demonstrated through their facile oxidation with hydrogen peroxide, providing trans-RuCl2(PP[PO]2)2 (1b) (PP[PO]2 = bis[2-(diphenylphosphine oxide)ethyl][2-(diphenylphosphino)ethyl]phosphine). Whilst chloride abstraction reactions from 1a appear slow (typical for trans dichlorides), cis-RuCl2(PP3) is shown to react rapidly with ethynylferrocene under ‘Dixneuf’ conditions (CH2Cl2, NaPF6, NEt3), providing the tri(hetero)metallic butenynyl complex [(PP3)Ru(η3–FcC3CHFc)]PF6 (2, Fc = ferrocenyl). The pendant groups of 1a-b offer great potential for future coordination studies (for example, to prepare mixed transition metal/lanthanide materials), whereby the facile synthetic route to 2 suggests a path towards examination of complex mixed-valence systems comprising multiple redox-active centres.Coordination of two potentially tetradentate ligands in a bidentate arrangement around a ruthenium centre can leave four uncoordinated phosphine moieties available for subsequent reaction, to give unusual binding motifs with ferrocenyl-alkynes.

p53 is an antitumor protein that plays an important role in apoptosis, preserving genomic stability and preventing angiogenesis, and it has been implicated in a large number of human cancers. For this reason it is an interesting target for both fundamental studies, such as the mechanism of interaction with DNA, and applications in biosensing. Here, we report a comprehensive study of label-free, full length p53 (flp53) and its interaction with engineered double-stranded DNA in vitro, at the single-molecule level, using atomic force microscopy (AFM) imaging and solid-state nanopore sensing. AFM data show that dimeric and tetrameric p53 bind to the DNA in a sequence-specific manner, confirming previously reported relative binding affinities. The statistical significance is tested using both the Grubbs test and stochastic simulations. For the first time, ultralow noise solid-state nanopore sensors are employed for the successful differentiation between bare DNA and p53/DNA complexes. Furthermore, translocation statistics reflect the binding affinities of different DNA sequences, in accordance with AFM data. Our results thus highlight the potential of solid-state nanopore sensors for single-molecule biosensing, especially when labeling is either not possible or at least not a viable option.

We have applied a new, robust and unsupervised approach to data collection, sorting and analysis that provides fresh insights into the nature of single-molecule junctions. Automation of tunneling current-distance (I(s)) spectroscopy facilitates the collection of very large data sets (up to 100 000 traces for a single experiment), enabling comprehensive statistical interrogations with respect to underlying tunneling characteristics, noise and junction formation probability (JFP). We frequently observe unusual low-to-high through-molecule conductance features with increasing electrode separation, in addition to numerous other “plateau” shapes, which may be related to changes in interfacial or molecular bridge structure. Furthermore, for the first time we use the JFP to characterize the homogeneity of functionalized surfaces at the nanoscale.

Solid-state nanopore devices with integrated electrodes are an important class of single-molecule biosensors, with potential applications in DNA, RNA, and protein detection and sequence analysis. Here we investigate solid-state nanopore sensors with an embedded gold film, fabricated using semiconductor processing techniques and focused ion beam milling. We characterize their geometric structure in three dimensions on the basis of experimental conductance studies and modeling as well as transmission electron microscopy imaging and tomography. We used electrodeposition to further shrink the pores to effective diameters below 10 nm and demonstrate how bipolar electrochemical coupling across the membrane can lead to significant contributions to the overall pore current and discuss its implications for nanopore sensing. Finally, we use metallized nanopores modified with homocysteine for the detection of insulin. We show that adsorption of the protein to the chemically modified nanopores slows down the translocation process to tens of milliseconds, which is orders of magnitude slower than expected for conventional electrophoretic transport.

We report the synthesis and full characterization of the entire haloferrocene (FcX) and 1,1′-dihaloferrocene (fcX2) series (X = I, Br, Cl, F; Fc = ferrocenyl, fc = ferrocene-1,1′-diyl). Finalization of this simple, yet intriguing set of compounds has been delayed by synthetic challenges associated with the incorporation of fluorine substituents. Successful preparation of fluoroferrocene (FcF) and 1,1′-difluoroferrocene (fcF2) were ultimately achieved using reactions between the appropriate lithiated ferrocene species and N-fluorobenzenesulfonimide (NFSI). The crude reaction products, in addition to those resulting from analogous preparations of chloroferrocene (FcCl) and 1,1′-dichloroferrocene (fcCl2), were utilized as model systems to probe the limits of a previously reported “oxidative purification” methodology. From this investigation and careful solution voltammetry studies, we find that the fluorinated derivatives exhibit the lowest redox potentials of each of the FcX and fcX2 series. This counterintuitive result is discussed with reference to the spectroscopic, structural, and first-principles calculations of these and related materials.

In the past two decades there has been a tremendous amount of research into the use of nanopores as single molecule sensors, which has been inspired by the Coulter counter and molecular transport across biological pores. Recently, the desire to increase structural resolution and analytical throughput has led to the integration of additional detection methods such as fluorescence spectroscopy. For structural information to be probed electronically high bandwidth measurements are crucial due to the high translocation velocity of molecules. The most commonly used solid-state nanopore sensors consist of a silicon nitride membrane and bulk silicon substrate. Unfortunately, the photoinduced noise associated with illumination of these platforms limits their applicability to high-bandwidth, high-laser-power synchronized optical and electronic measurements. Here we present a unique low-noise nanopore platform, composed of a predominately Pyrex substrate and silicon nitride membrane, for synchronized optical and electronic detection of biomolecules. Proof of principle experiments are conducted showing that the Pyrex substrates have substantially lowers ionic current noise arising from both laser illumination and platform capacitance. Furthermore, using confocal microscopy and a partially metallic pore we demonstrate high signal-to-noise synchronized optical and electronic detection of dsDNA.Keywords: fluorescence; nanopore; noise; single-molecule; zero-mode waveguide;

Herein, we describe the integration of two glass nanopores into a segmented flow microfluidic device with a view on enhancing the functionality of label free, single molecule nanopore sensors. Within a robust and mechanically stable platform, individual droplet compositions are distinguished before single molecule translocations from the droplet are detected electrochemically via the Coulter principle. This result is highly significant, combining the sensitivity of single molecule methods and their ability to overcome the clouding of the ensemble average with the “isolated microreactor” benefits of droplet microfluidics. Furthermore, devices as presented here provide the platform for the development of systems where the injection and extraction of single molecules allow droplet composition to be controlled at the molecular level.

Single-stranded DNA (ssDNA) binding protein plays an important role in the DNA replication process in a wide range of organisms. It binds to ssDNA to prevent premature reannealing and to protect it from degradation. Current understanding of SSB/ssDNA interaction points to a complex mechanism, including SSB motion along the DNA strand. We report on the first characterization of this interaction at the single-molecule level using solid-state nanopore sensors, namely without any labeling or surface immobilization. Our results show that the presence of SSB on the ssDNA can control the speed of nanopore translocation, presumably due to strong interactions between SSB and the nanopore surface. This enables nanopore-based detection of ssDNA fragments as short as 37 nt, which is normally very difficult with solid-state nanopore sensors, due to constraints in noise and bandwidth. Notably, this fragment is considerably shorter than the 65 nt binding motif, typically required for SSB binding at high salt concentrations. The nonspecificity of SSB binding to ssDNA further suggests that this approach could be used for fragment sizing of short ssDNA.

A simple and versatile method for the direct fabrication of tunneling electrodes with controllable gap distance by using electron-beam-induced deposition (EBID) is presented. We show that tunneling nanogaps smaller than the minimum feature size realizable by conventional EBID can be achieved with a standard scanning electron microscope. These gaps can easily be embedded in nanopores with high accuracy. The controllability of this fabrication method and the nanogap geometry was verified by SEM and TEM imaging. Furthermore, tunneling spectroscopy in a group of solvents with different barrier heights was used to determine the nanogap functionality. Ultimately, the presented fabrication method can be further applied for the fabrication of arrays of nanogap/nanopores or nanogap electrodes with tunable electrode materials. Additionally, this method can also offer direct fabrication of nanoscale electrode systems with tunable spacing for redox cycling and plasmonic applications, which represents an important step in the development of tunneling nanopore structures and in enhancing the capabilities of nanopore sensors.Keywords: label-free detection; nanopore sequencing; nanopores; single-molecule detection; tunneling gaps

A systematic study into the Sonogashira cross-coupling of 1,1′-diiodoferrocene (fcI2) confirms that the Pd(0)–P(tBu)3 system provides a remarkable rate increase over Pd(0)–(PPh3)2. Attempts to couple 4-ethynylphenylthioacetate (2) with fcI2 instead produced a novel cyclic trimer of the former, from syn addition of S–Ac across CC.

We report the large scale syntheses and ‘oxidative purification’ of fcI2, fcBr2 and FcBr (fc = ferrocene-1,1′-diyl, Fc = ferrocenyl). These valuable starting materials are typically laborious to separate via conventional techniques, but can be readily isolated by taking advantage of their increased E1/2 relative to FcH/FcX contaminants. Our work extends this methodology towards a generic tool for the separation of redox active mixtures.

Solid-state nanopores with integrated electrodes have interesting prospects in next-generation single-molecule biosensing and sequencing. These include “gated” nanopores with a single electrode integrated into the membrane, as well as two-electrode designs, such as a transversal tunneling junction. Here we report the first comprehensive analysis of current flow in a three-electrode device as a model for this class of sensors. As a new feature, we observe apparent rectification in the pore current that is rooted in the current distribution of the cell, rather than the geometry or electrostatics of the pore. We benchmark our results against a recently developed theoretical model and define operational parameters for nanopore/electrode structures. Our findings thus facilitate the rational design of such sensor devices.Keywords: charge transfer resistance; electrode cross-coupling; ionic current rectification; metallic nanopores; multielectrode arrangement

The syntheses and electrochemical/optical properties of some branched and linear 1,1′-substituted ferrocene complexes for molecular electronics are described. Metal centers were extended (and where relevant, connected) by arylethynyl spacers functionalized with m-pyridyl, tert-butylthiol (StBu), and trimethylsilyl (TMS) moieties. Such systems provide two well-defined molecular pathways for electron transfer and hold interesting prospects for the study of new charge transport processes, such as quantum interference, local gating, and correlated hopping events.

Studying electron transport through immobilized proteins at the single-molecule level has been of interest for more than two decades, with a view on the fundamentals of charge transport in condensed media and applications in bioelectronics. Scanning tunneling microscopy (STM) is a powerful tool in this context, because, at least in principle, it should be possible to address individual proteins on an electrode surface reproducibly with single-protein precision. As reported in this issue of ACS Nano, MacDonald and colleagues have now achieved this for the first time at room temperature for covalently immobilized cytochrome b562, combining imaging and tunneling spectroscopy in a custom-built, ultralow drift STM, with single-protein precision. Using site-directed mutagenesis, cysteines introduced in specific locations in the amino acid sequence of the protein allowed the team to investigate conduction along different directions through the protein, namely along its short and long axes.

We report on the fabrication and characterization of a DNA nanopore detector with integrated tunneling electrodes. Functional tunneling devices were identified by tunneling spectroscopy in different solvents and then used in proof-of-principle experiments demonstrating, for the first time, concurrent tunneling detection and ionic current detection of DNA molecules in a nanopore platform. This is an important step toward ultrafast DNA sequencing by tunneling.

Nanopore-based single-molecule sensors have become an important class of analytical devices that have in some cases already reached the market place. Traditionally operated in a two-electrode configuration, devices with three or more electrodes have emerged recently, for example with a view on switching the transport properties of the nanopore or even tunneling-based detection of analytes with the ultimate goal of inexpensive and ultrafast DNA sequencing. How do these additional electrodes affect the current distribution in the cell and hence the sensor performance? This is significantly less clear and thus in focus here. We use impedance modeling of a prototypical three-electrode nanopore sensor and show that, depending on the conditions, standard experimental device characterization is severely affected by the presence of the third electrode. On the other hand, the simulations also provide guidelines on how to avoid such complications, identify “safe” operating conditions, and design criteria for optimized nanopore sensors.Keywords: nanopore; sensing; sequencing; single-molecule; solid-state

Biological and solid-state nanopores have recently attracted much interest as ultrafast DNA fragment sizing and sequencing devices. Their potential however goes far beyond DNA sequencing. In particular, nanopores offer perspectives of single-molecule (bio)sensing at physiologically relevant concentrations, which is key for studying protein/protein or protein/DNA interactions. Integration of electrode structures into solid-state nanopore devices moreover enables control and fast switching of the pore properties, e.g. for active control of biopolymer transport through the nanopore. We present some of recent work in this area, namely the fabrication and characterization of nanopore/electrode architectures for single-(bio)molecule sensing. Specifically, we introduce a new technique to fabricate ultra-small metal nanopores with diameters smaller than 20 nm based on ion current feedback (ICF) controlled electrodeposition. It offers precise control of the pore conductance, is easily multiplexed, and can be extended to a wide range of different metals.

We report the large scale syntheses and ‘oxidative purification’ of fcI2, fcBr2 and FcBr (fc = ferrocene-1,1′-diyl, Fc = ferrocenyl). These valuable starting materials are typically laborious to separate via conventional techniques, but can be readily isolated by taking advantage of their increased E1/2 relative to FcH/FcX contaminants. Our work extends this methodology towards a generic tool for the separation of redox active mixtures.

A systematic study into the Sonogashira cross-coupling of 1,1′-diiodoferrocene (fcI2) confirms that the Pd(0)–P(tBu)3 system provides a remarkable rate increase over Pd(0)–(PPh3)2. Attempts to couple 4-ethynylphenylthioacetate (2) with fcI2 instead produced a novel cyclic trimer of the former, from syn addition of S–Ac across CC.