Sterically Enhanced Control of Enzyme-Assisted DNA Assembly.

Traditional methods for the assembly of functionalised DNA structures, involving enzyme restriction and modification, present difficulties when working with small DNA fragments (<100 bp), in part due to a lack of control over enzymatic action during the DNA modification process. This limits the design flexibility and range of accessible DNA structures. Here, we show that these limitations can be overcome by introducing chemical modifications into the DNA that spatially restrict enzymatic activity. This approach, sterically controlled nuclease enhanced (SCoNE) DNA assembly, thereby circumvents the size limitations of conventional Gibson assembly (GA) and allows the preparation of well-defined, functionalised DNA structures with multiple probes for specific analytes, such as IL-6, procalcitonin (PCT), and a biotin reporter group. Notably, when using the same starting materials, conventional GA under typical conditions fails. We demonstrate successful analyte capture based on standard and modified sandwich ELISA and also show how the inclusion of biotin probes provides additional functionality for product isolation.

Quantum Interference and Contact Effects in the Thermoelectric Performance of Anthracene-Based Molecules.

We report on the single-molecule electronic and thermoelectric properties of strategically chosen anthracene-based molecules with anchor groups capable of binding to noble metal substrates, such as gold and platinum. Specifically, we study the effect of different anchor groups, as well as quantum interference, on the electric conductance and the thermopower of gold/single-molecule/gold junctions and generally find good agreement between theory and experiments. All molecular junctions display transport characteristics consistent with coherent transport and a Fermi alignment approximately in the middle of the highest occupied molecular orbital/lowest unoccupied molecular orbital gap. Single-molecule results are in agreement with previously reported thin-film data, further supporting the notion that molecular design considerations may be translated from the single- to many-molecule devices. For combinations of anchor groups where one binds significantly more strongly to the electrodes than the other, the stronger anchor group appears to dominate the thermoelectric behavior of the molecular junction. For other combinations, the choice of electrode material can determine the sign and magnitude of the thermopower. This finding has important implications for the design of thermoelectric generator devices, where both n- and p-type conductors are required for thermoelectric current generation.

Programmed Self-Assembly of Single Colloidal Gyroids for Chiral Photonic Crystals.

Gyroid structures are of extensive interest because they provide a rich platform for chiroptics as well as topological photonics. While the double-gyroid morphology as a bicontinuous structure is not uncommon in self-assembled soft materials, direct self-assembly of single-network gyroids has proven elusive. Here, an enantiomorphic pair of single-gyroid crystals comprising colloidal spheres is presented, and two distinct routes are demonstrated for programmed self-assembly of each single colloidal gyroid enantiomorph from rationally designed patchy spheres. The designer colloidal patchy spheres, which closely hew to their synthetic feasibility, are chiral, having either two staggered rectangular patches at opposite poles or four circular patches arranged in a well-defined geometry. The single colloidal gyroid, as well as its inverse structure, is shown to support a wide complete photonic bandgap in addition to exhibiting rich chiroptical properties, making them attractive chiral photonic crystals. The versatility of this single colloidal gyroid, the bottom-up routes devised here in silico, and the robustness of the design space for the chiral colloidal patchy spheres together make a strong case for single colloidal gyroids to supersede colloidal diamond, as a target for programmed self-assembly, in the quest for photonic crystals operating at optical frequencies.

Unsupervised classification of voltammetric data beyond principal component analysis.

In this study, we evaluate different apoproaches to unsupervised classification of cyclic voltammetric data, including Principal Component Analysis (PCA), t-distributed Stochastic Neighbour Embedding (t-SNE), Uniform Manifold Approximation and Projection (UMAP) as well as neural networks. To this end, we exploit a form of transfer learning, based on feature extraction in an image recognition network, VGG-16, in combination with PCA, t-SNE or UMAP. Overall, we find that t-SNE performs best when applied directly to numerical data (noise-free case) or to features (in the presence of noise), followed by UMAP and then PCA.

Assembly, structure and thermoelectric properties of 1,1'-dialkynylferrocene 'hinges'.

Dialkynylferrocenes exhibit attractive electronic and rotational features that make them ideal candidates for use in molecular electronic applications. However previous works have primarily focussed on single-molecule studies, with limited opportunities to translate these features into devices. In this report, we utilise a variety of techniques to examine both the geometric and electronic structure of a range of 1,1'-dialkynylferrocene molecules, as either single-molecules, or as self-assembled monolayers. Previous single molecule studies have shown that similar molecules can adopt an 'open' conformation. However, in this work, DFT calculations, STM-BJ experiments and AFM imaging reveal that these molecules prefer to occupy a 'hairpin' conformation, where both alkynes point towards the metal surface. Interestingly we find that only one of the terminal anchor groups binds to the surface, though both the presence and nature of the second alkyne affect the thermoelectric properties of these systems. First, the secondary alkyne acts to affect the position of the frontier molecular orbitals, leading to increases in the Seebeck coefficient. Secondly, theoretical calculations suggested that rotating the secondary alkyne away from the surface acts to modulate thermoelectric properties. This work represents the first of its kind to examine the assembly of dialkynylferrocenes, providing valuable information about both their structure and electronic properties, as well as unveiling new ways in which both of these properties can be controlled.

Multi-component self-assembled molecular-electronic films: towards new high-performance thermoelectric systems.

The thermoelectric properties of parallel arrays of organic molecules on a surface offer the potential for large-area, flexible, solution processed, energy harvesting thin-films, whose room-temperature transport properties are controlled by quantum interference (QI). Recently, it has been demonstrated that constructive QI (CQI) can be translated from single molecules to self-assembled monolayers (SAMs), boosting both electrical conductivities and Seebeck coefficients. However, these CQI-enhanced systems are limited by rigid coupling of the component molecules to metallic electrodes, preventing the introduction of additional layers which would be advantageous for their further development. These rigid couplings also limit our ability to suppress the transport of phonons through these systems, which could act to boost their thermoelectric output, without comprising on their impressive electronic features. Here, through a combined experimental and theoretical study, we show that cross-plane thermoelectricity in SAMs can be enhanced by incorporating extra molecular layers. We utilize a bottom-up approach to assemble multi-component thin-films that combine a rigid, highly conductive 'sticky'-linker, formed from alkynyl-functionalised anthracenes, and a 'slippery'-linker consisting of a functionalized metalloporphyrin. Starting from an anthracene-based SAM, we demonstrate that subsequent addition of either a porphyrin layer or a graphene layer increases the Seebeck coefficient, and addition of both porphyrin and graphene leads to a further boost in their Seebeck coefficients. This demonstration of Seebeck-enhanced multi-component SAMs is the first of its kind and presents a new strategy towards the design of thin-film thermoelectric materials.

Taming the thermodiffusion of alkali halide solutions in silica nanopores.

Thermal fields give rise to thermal coupling phenomena, such as mass and charge fluxes, which are useful in energy recovery applications and nanofluidic devices for pumping, mixing or desalination. Here we use state of the art non-equilibrium molecular simulations to quantify the thermodiffusion of alkali halide solutions, LiCl and NaCl, confined in silica nanopores, targeting diameters of the order of those found in mesoporous silica nanostructures. We show that nanoconfinement modifies the thermodiffusion behaviour of the solution. Under confinement conditions, the solutions become more thermophilic, with a preference to accumulate at hot sources, or thermoneutral, with the thermodiffusion being inhibited. Our work highlights the importance of nanoconfinement in thermodiffusion and outlines strategies to tune mass transport at the nanoscale, using thermal fields.

Scale-Up of Room-Temperature Constructive Quantum Interference from Single Molecules to Self-Assembled Molecular-Electronic Films.

The realization of self-assembled molecular-electronic films, whose room-temperature transport properties are controlled by quantum interference (QI), is an essential step in the scale-up of QI effects from single molecules to parallel arrays of molecules. Recently, the effect of destructive QI (DQI) on the electrical conductance of self-assembled monolayers (SAMs) has been investigated. Here, through a combined experimental and theoretical investigation, we demonstrate chemical control of different forms of constructive QI (CQI) in cross-plane transport through SAMs and assess its influence on cross-plane thermoelectricity in SAMs. It is known that the electrical conductance of single molecules can be controlled in a deterministic manner, by chemically varying their connectivity to external electrodes. Here, by employing synthetic methodologies to vary the connectivity of terminal anchor groups around aromatic anthracene cores, and by forming SAMs of the resulting molecules, we clearly demonstrate that this signature of CQI can be translated into SAM-on-gold molecular films. We show that the conductance of vertical molecular junctions formed from anthracene-based molecules with two different connectivities differ by a factor of approximately 16, in agreement with theoretical predictions for their conductance ratio based on CQI effects within the core. We also demonstrate that for molecules with thioether anchor groups, the Seebeck coefficient of such films is connectivity dependent and with an appropriate choice of connectivity can be boosted by ∼50%. This demonstration of QI and its influence on thermoelectricity in SAMs represents a critical step toward functional ultra-thin-film devices for future thermoelectric and molecular-scale electronics applications.

Rapid Fragmentation during Seeded Lysozyme Aggregation Revealed at the Single Molecule Level.

Protein aggregation is associated with neurodegenerative disorders such as Alzheimer's and Parkinson's diseases. The poorly understood pathogenic mechanism of amyloid diseases makes early stage diagnostics or therapeutic intervention a challenge. Seeded polymerization that reduces the duration of the lag phase and accelerates fibril growth is a widespread model to study amyloid formation. Seeding effects are hypothesized to be important in the "infectivity" of amyloids and are linked to the development of systemic amyloidosis in vivo. The exact mechanism of seeding is unclear yet critical to illuminating the propagation of amyloids. Here we report on the lateral and axial fragmentation of seed fibrils in the presence of lysozyme monomers at short time scales, followed by the generation of oligomers and growth of fibrils.

Single-Molecule Analysis with Solid-State Nanopores.

Solid-state nanopores and nanopipettes are an exciting class of single-molecule sensors that has grown enormously over the last two decades. They offer a platform for testing fundamental concepts of stochasticity and transport at the nanoscale, for studying single-molecule biophysics and, increasingly, also for new analytical applications and in biomedical sensing. This review covers some fundamental aspects underpinning sensor operation and transport and, at the same time, it aims to put these into context as an analytical technique. It highlights new and recent developments and discusses some of the challenges lying ahead.

Assisted delivery of anti-tumour platinum drugs using DNA-coiling gold nanoparticles bearing lumophores and intercalators: towards a new generation of multimodal nanocarriers with enhanced action.

New gold and lipoic based nanocarriers for the delivery of platinum(ii) and platinum(iv) drugs are developed, which allow enhanced loading of the drug on the surface of the nanocarriers and release in a pH-dependent fashion, with superior release at lower pHs which are associated with many tumours. The conjugate nanoparticles and their conjugates enter cells rapidly (within 3 hours). They tend to cluster in vesicles and are also observed by light and electron microscopies in the cytoplasm, endoplasmic reticulum and nucleus. We further incorporate aminoanthraquinone units that are both fluorophores and DNA intercalators. This results in nanocarriers that after drug release will remain surface decorated with DNA-binders challenging the conventional design of the nanocarrier as an inert component. The outcome is nanocarriers that themselves have distinctive, remarkable and unusual DNA binding properties being able to bind and wrap DNA (despite their anionic charge) and provide enhanced cytotoxic activity beyond that conferred by the platinum agents they release. DNA coiling is usually associated with polycations which can disrupt cell membranes; anionic nanoparticles that can cause novel and dramatic effects on DNA may have fascinating potential for new approaches to in-cell nucleic acid recognition. Our findings have implications for the understanding and interpretation of the biological activities of nanoparticles used to deliver other DNA-binding drugs including clinical drug doxorubicin and its formulations.

Cross-plane conductance through a graphene/molecular monolayer/Au sandwich.

The functionalities offered by single-molecule electrical junctions are yet to be translated into monolayer or few-layer molecular films, where making effective and reproducible electrical contact is one of the challenging bottlenecks. Here we take a significant step in this direction by demonstrating that excellent electrical contact can be made with a monolayer biphenyl-4,4'-dithiol (BPDT) molecular film, sandwiched between gold and graphene electrodes. This sandwich device structure is advantageous, because the current flows through the molecules to the gold substrate in a 'cross-plane' manner, perpendicular to the plane of graphene, yielding high-conductance devices. We elucidate the nature of the cross-plane graphene/molecule/Au transport using quantum transport calculations and introduce a simple analytical model, which captures generic features of the current-voltage characteristic. Asymmetry in junction properties results from the disparity in electrode electrical properties, the alignment of the BPDT HOMO-LUMO energy levels and the specific characteristics of the graphene electrode. The experimental observation of scalability of junction properties within the junction area, in combination with a theoretical description of the transmission probability of the thiol-graphene contact, demonstrates that between 10% and 100% of the molecules make contact with the electrodes, which is several orders of magnitude greater than that achieved to date in the literature.

A Redox-Activated G-Quadruplex DNA Binder Based on a Platinum(IV)-Salphen Complex.

There has been increasing interest in the development of small molecules that can selectively bind to G-quadruplex DNA structures. The latter have been associated with a number of key biological processes and therefore are proposed to be potential targets for drug development. Herein, we report the first example of a reduction-activated G-quadruplex DNA binder. We show that a new octahedral platinum(IV)-salphen complex does not interact with DNA in aqueous media at pH 7.4; however, upon addition of bioreductants such as ascorbic acid or glutathione, the compound is readily reduced to the corresponding square planar platinum(II) complex. In contrast to the parent platinum(IV) complex, the in situ generated platinum(II) complex has good affinity for G-quadruplex DNA.

Controlling the Dynamic Instability of Capped Metal Nanoparticles on Metallic Surfaces.

Small metal nanoparticles (NPs) with core-sizes ranging from 1 to 3 nm constitute a bridge between molecules and colloids with unique electronic, catalytic, and other properties. Many applications entail immobilization onto solid supports, but while NP behavior in solution is well studied, the effect of the interaction between NPs and the substrate surface is understood less. Here, we follow the structural evolution of thiolated monolayer-protected AuNPs on Au(111) substrates at the single-particle level in real-time using high-resolution in situ scanning tunneling microscopy. We show how the reactivity of the substrate affects the stability of the immobilized NPs and how their structural identity can be preserved. Entropically driven redistribution of the NP's protective capping layer is an important element in the disintegration process and at the same time rather generic. Our findings may thus have wider implications on the design and optimization of functional surfaces involving NPs, made of materials other than Au.

Single Molecule Trapping and Sensing Using Dual Nanopores Separated by a Zeptoliter Nanobridge.

There is a growing realization, especially within the diagnostic and therapeutic community, that the amount of information enclosed in a single molecule can not only enable a better understanding of biophysical pathways, but also offer exceptional value for early stage biomarker detection of disease onset. To this end, numerous single molecule strategies have been proposed, and in terms of label-free routes, nanopore sensing has emerged as one of the most promising methods. However, being able to finely control molecular transport in terms of transport rate, resolution, and signal-to-noise ratio (SNR) is essential to take full advantage of the technology benefits. Here we propose a novel solution to these challenges based on a method that allows biomolecules to be individually confined into a zeptoliter nanoscale droplet bridging two adjacent nanopores (nanobridge) with a 20 nm separation. Molecules that undergo confinement in the nanobridge are slowed down by up to 3 orders of magnitude compared to conventional nanopores. This leads to a dramatic improvement in the SNR, resolution, sensitivity, and limit of detection. The strategy implemented is universal and as highlighted in this manuscript can be used for the detection of dsDNA, RNA, ssDNA, and proteins.

Deep learning for single-molecule science.

Exploring and making predictions based on single-molecule data can be challenging, not only due to the sheer size of the datasets, but also because a priori knowledge about the signal characteristics is typically limited and poor signal-to-noise ratio. For example, hypothesis-driven data exploration, informed by an expectation of the signal characteristics, can lead to interpretation bias or loss of information. Equally, even when the different data categories are known, e.g., the four bases in DNA sequencing, it is often difficult to know how to make best use of the available information content. The latest developments in machine learning (ML), so-called deep learning (DL) offer interesting, new avenues to address such challenges. In some applications, such as speech and image recognition, DL has been able to outperform conventional ML strategies and even human performance. However, to date DL has not been applied much in single-molecule science, presumably in part because relatively little is known about the 'internal workings' of such DL tools within single-molecule science as a field. In this Tutorial, we make an attempt to illustrate in a step-by-step guide how one of those, a convolutional neural network (CNN), may be used for base calling in DNA sequencing applications. We compare it with a SVM as a more conventional ML method, and discuss some of the strengths and weaknesses of the approach. In particular, a 'deep' neural network has many features of a 'black box', which has important implications on how we look at and interpret data.

Insulated molecular wires: inhibiting orthogonal contacts in metal complex based molecular junctions.

Metal complexes are receiving increased attention as molecular wires in fundamental studies of the transport properties of metal|molecule|metal junctions. In this context we report the single-molecule conductance of a systematic series of d8 square-planar platinum(ii) trans-bis(alkynyl) complexes with terminal trimethylsilylethynyl (C[triple bond, length as m-dash]CSiMe3) contacting groups, e.g. trans-Pt{C[triple bond, length as m-dash]CC6H4C[triple bond, length as m-dash]CSiMe3}2(PR3)2 (R = Ph or Et), using a combination of scanning tunneling microscopy (STM) experiments in solution and theoretical calculations using density functional theory and non-equilibrium Green's function formalism. The measured conductance values of the complexes (ca. 3-5 × 10-5G0) are commensurate with similarly structured all-organic oligo(phenylene ethynylene) and oligo(yne) compounds. Based on conductance and break-off distance data, we demonstrate that a PPh3 supporting ligand in the platinum complexes can provide an alternative contact point for the STM tip in the molecular junctions, orthogonal to the terminal C[triple bond, length as m-dash]CSiMe3 group. The attachment of hexyloxy side chains to the diethynylbenzene ligands, e.g. trans-Pt{C[triple bond, length as m-dash]CC6H2(Ohex)2C[triple bond, length as m-dash]CSiMe3}2(PPh3)2 (Ohex = OC6H13), hinders contact of the STM tip to the PPh3 groups and effectively insulates the molecule, allowing the conductance through the full length of the backbone to be reliably measured. The use of trialkylphosphine (PEt3), rather than triarylphosphine (PPh3), ancillary ligands at platinum also eliminates these orthogonal contacts. These results have significant implications for the future design of organometallic complexes for studies in molecular junctions.