Raman micro-spectroscopy reveals the spatial distribution of fumarate in cells and tissues.

Aberrantly accumulated metabolites elicit intra- and inter-cellular pro-oncogenic cascades, yet current measurement methods require sample perturbation/disruption and lack spatio-temporal resolution, limiting our ability to fully characterize their function and distribution. Here, we show that Raman spectroscopy (RS) can directly detect fumarate in living cells in vivo and animal tissues ex vivo, and that RS can distinguish between Fumarate hydratase (Fh1)-deficient and Fh1-proficient cells based on fumarate concentration. Moreover, RS reveals the spatial compartmentalization of fumarate within cellular organelles in Fh1-deficient cells: consistent with disruptive methods, we observe the highest fumarate concentration (37 ± 19 mM) in mitochondria, where the TCA cycle operates, followed by the cytoplasm (24 ± 13 mM) and then the nucleus (9 ± 6 mM). Finally, we apply RS to tissues from an inducible mouse model of FH loss in the kidney, demonstrating RS can classify FH status. These results suggest RS could be adopted as a valuable tool for small molecule metabolic imaging, enabling in situ non-destructive evaluation of fumarate compartmentalization.

Spearheading a new era in complex colloid synthesis with TPM and other silanes.

Colloid science has recently grown substantially owing to the innovative use of silane coupling agents (SCAs), especially 3-trimethoxysilylpropyl methacrylate (TPM). SCAs were previously used mainly as modifying agents, but their ability to form droplets and condense onto pre-existing structures has enabled their use as a versatile and powerful tool to create novel anisotropic colloids with increasing complexity. In this Review, we highlight the advances in complex colloid synthesis facilitated by the use of TPM and show how this has driven remarkable new applications. The focus is on TPM as the current state-of-the-art in colloid science, but we also discuss other silanes and their potential to make an impact. We outline the remarkable properties of TPM colloids and their synthesis strategies, and discuss areas of soft matter science that have benefited from TPM and other SCAs.

Full Control of Plasmonic Nanocavities Using Gold Decahedra-on-Mirror Constructs with Monodisperse Facets.

Bottom-up assembly of nanoparticle-on-mirror (NPoM) nanocavities enables precise inter-metal gap control down to ≈ 0.4 nm for confining light to sub-nanometer scales, thereby opening opportunities for developing innovative nanophotonic devices. However limited understanding, prediction, and optimization of light coupling and the difficulty of controlling nanoparticle facet shapes restricts the use of such building blocks. Here, an ultraprecise symmetry-breaking plasmonic nanocavity based on gold nanodecahedra is presented, to form the nanodecahedron-on-mirror (NDoM) which shows highly consistent cavity modes and fields. By characterizing > 20 000 individual NDoMs, the variability of light in/output coupling is thoroughly explored and a set of robust higher-order plasmonic whispering gallery modes uniquely localized at the edges of the triangular facet in contact with the metallic substrate is found. Assisted by quasinormal mode simulations, systematic elaboration of NDoMs is proposed to give nanocavities with near hundred-fold enhanced radiative efficiencies. Such systematically designed and precisely-assembled metallic nanocavities will find broad application in nanophotonic devices, optomechanics, and surface science.

Boosting Optical Nanocavity Coupling by Retardation Matching to Dark Modes.

Plasmonic nanoantennas can focus light at nanometer length scales providing intense field enhancements. For the tightest optical confinements (0.5-5 nm) achieved in plasmonic gaps, the gap spacing, refractive index, and facet width play a dominant role in determining the optical properties making tuning through antenna shape challenging. We show here that controlling the surrounding refractive index instead allows both efficient frequency tuning and enhanced in-/output coupling through retardation matching as this allows dark modes to become optically active, improving widespread functionalities.

Tracking interfacial single-molecule pH and binding dynamics via vibrational spectroscopy.

Understanding single-molecule chemical dynamics of surface ligands is of critical importance to reveal their individual pathways and, hence, roles in catalysis, which ensemble measurements cannot see. Here, we use a cascaded nano-optics approach that provides sufficient enhancement to enable direct tracking of chemical trajectories of single surface-bound molecules via vibrational spectroscopy. Atomic protrusions are laser-induced within plasmonic nanojunctions to concentrate light to atomic length scales, optically isolating individual molecules. By stabilizing these atomic sites, we unveil single-molecule deprotonation and binding dynamics under ambient conditions. High-speed field-enhanced spectroscopy allows us to monitor chemical switching of a single carboxylic group between three discrete states. Combining this with theoretical calculation identifies reversible proton transfer dynamics (yielding effective single-molecule pH) and switching between molecule-metal coordination states, where the exact chemical pathway depends on the intitial protonation state. These findings open new domains to explore interfacial single-molecule mechanisms and optical manipulation of their reaction pathways.

Contact angle as a powerful tool in anisotropic colloid synthesis.

Nucleation and growth is a technique widely used to prepare colloids, in which droplets are adsorbed onto substrate particles. Changing the contact angle of the substrates can greatly alter the morphology of the product particles. Here, we investigate the nucleation and growth of 3-methacryloxypropyltrimethoxysilane (MPTMS) both onto Stöber spheres and onto (cross-linked) MPTMS* spheres. The former results in 'snowman' particles with a cap-shaped MPTMS* compartment, and we show that their morphology is highly controllable via the MPTMS content in the reaction mixture. The contact angle of the MPTMS* compartment decreases with droplet diameter, suggesting that this wetting process is affected not only by surface tension but also by line tension. In contrast to Stöber spheres, MPTMS* substrate particles yield highly reproducible and tuneable 'engulfed-sphere' colloids with an internal reference axis (but a homogeneous mass distribution). These engulfed-sphere particles can be fully index-matched for confocal microscopy on account of their homogeneous refractive index. Suitable index-matching mixtures of polar and of low-polar media are presented, where cyclohexyl iodide (CHI) is introduced as a new medium for colloids of high refractive index. Finally, the index-matched engulfed-sphere colloids are self-assembled into (close-packed and long-range) plastic phases, and the particles' rotational diffusion inside the crystal phases is tracked via confocal microscopy.

Cascaded nanooptics to probe microsecond atomic-scale phenomena.

Plasmonic nanostructures can focus light far below the diffraction limit, and the nearly thousandfold field enhancements obtained routinely enable few- and single-molecule detection. However, for processes happening on the molecular scale to be tracked with any relevant time resolution, the emission strengths need to be well beyond what current plasmonic devices provide. Here, we develop hybrid nanostructures incorporating both refractive and plasmonic optics, by creating SiO2 nanospheres fused to plasmonic nanojunctions. Drastic improvements in Raman efficiencies are consistently achieved, with (single-wavelength) emissions reaching 107 counts⋅mW-1⋅s-1 and 5 × 105 counts∙mW-1∙s-1∙molecule-1, for enhancement factors >1011 We demonstrate that such high efficiencies indeed enable tracking of single gold atoms and molecules with 17-µs time resolution, more than a thousandfold improvement over conventional high-performance plasmonic devices. Moreover, the obtained (integrated) megahertz count rates rival (even exceed) those of luminescent sources such as single-dye molecules and quantum dots, without bleaching or blinking.

Light-Induced Coalescence of Plasmonic Dimers and Clusters.

The properties of nanoplasmonic structures depend strongly on their geometry, creating the need for high-precision control and characterization. Here, by exploiting the low activation energy of gold atoms on nanoparticle surfaces, we show how laser irradiation reshapes nanoparticle dimers. Time-course dark-field microspectroscopy allows this process to be studied in detail for individual nanostructures. Three regimes are identified: facet growth, formation of a conductive bridge between particles, and bridge growth. Electromagnetic simulations confirm the growth dynamics and allow measurement of bridge diameter, found to be highly reproducible and also self-limiting. Correlations in spectral resonances for the initial and final states give insight into the energy barriers for bridge growth. Dark-field microscopy shows that coalescence of multiple gaps in nanoparticle clusters can be digitally triggered, with each gap closing after discrete increases in irradiation power. Such control is important for light-induced nanowire formation or trimming of electronic and optoelectronic devices.

Multivalent Patchy Colloids for Quantitative 3D Self-Assembly Studies.

We report methods to synthesize sub-micron- and micron-sized patchy silica particles with fluorescently labeled hemispherical titania protrusions, as well as routes to efficiently characterize these particles and self-assemble these particles into non-close-packed structures. The synthesis methods expand upon earlier work in the literature, in which silica particles packed in a colloidal crystal were surface-patterned with a silane coupling agent. Here, hemispherical amorphous titania protrusions were successfully labeled with fluorescent dyes, allowing for imaging by confocal microscopy and super-resolution techniques. Confocal microscopy was exploited to experimentally determine the numbers of protrusions per particle over large numbers of particles for good statistical significance, and these distributions were compared to simulations predicting the number of patches as a function of core particle polydispersity and maximum separation between the particle surfaces. We self-assembled these patchy particles into open percolating gel networks by exploiting solvophobic attractions between the protrusions.

Flickering nanometre-scale disorder in a crystal lattice tracked by plasmonic flare light emission.

The dynamic restructuring of metal nanoparticle surfaces is known to greatly influence their catalytic, electronic transport, and chemical binding functionalities. Here we show for the first time that non-equilibrium atomic-scale lattice defects can be detected in nanoparticles by purely optical means. These fluctuating states determine interface electronic transport for molecular electronics but because such rearrangements are low energy, measuring their rapid dynamics on single nanostructures by X-rays, electron beams, or tunnelling microscopies, is invasive and damaging. We utilise nano-optics at the sub-5nm scale to reveal rapid (on the millisecond timescale) evolution of defect morphologies on facets of gold nanoparticles on a mirror. Besides dynamic structural information, this highlights fundamental questions about defining bulk plasma frequencies for metals probed at the nanoscale.

Inhibiting Analyte Theft in Surface-Enhanced Raman Spectroscopy Substrates: Subnanomolar Quantitative Drug Detection.

Quantitative applications of surface-enhanced Raman spectroscopy (SERS) often rely on surface partition layers grafted to SERS substrates to collect and trap-solvated analytes that would not otherwise adsorb onto metals. Such binding layers drastically broaden the scope of analytes that can be probed. However, excess binding sites introduced by this partition layer also trap analytes outside the plasmonic "hotspots". We show that by eliminating these binding sites, limits of detection (LODs) can effectively be lowered by more than an order of magnitude. We highlight the effectiveness of this approach by demonstrating quantitative detection of controlled drugs down to subnanomolar concentrations in aqueous media. Such LODs are low enough to screen, for example, urine at clinically relevant levels. These findings provide unique insights into the binding behavior of analytes, which are essential when designing high-performance SERS substrates.

Unexpected stability of aqueous dispersions of raspberry-like colloids.

Aqueous colloidal suspensions, both man-made and natural, are part of our everyday life. The applicability of colloidal suspensions, however, is limited by the range of conditions over which they are stable. Here we report a novel type of highly monodisperse raspberry-like colloids, which are prepared in a single-step synthesis that relies on simultaneous dispersion and emulsion polymerisation. The resulting raspberry colloids behave almost like hard spheres. In aqueous solutions, such prepared raspberries show unexpected stability against aggregation over large variations of added salt concentrations without addition of stabilisers. We present simple Derjaguin-Landau-Verwey-Overbeek (DLVO) calculations performed on raspberry-like and smooth colloids showing that this stability results from our raspberries' unique morphology, which extends our understanding of colloidal stability against salting. Further, the raspberries' stability facilitates the formation of superspheres and thin films in which the raspberry colloids self-assemble into hexagonally close-packed photonic crystals with exquisite reproducibility.

Protein-mediated gelation and nano-scale assembly of unfunctionalized hyaluronic acid and chondroitin sulfate.

Background: Hyaluronic acid (HA) is a major component of the extracellular matrix (ECM) in the central nervous system and the only purely supramolecular glycosaminoglycan. Much focus has been given to using this high molecular weight polysaccharide for tissue engineering applications. In most studies, the backbone of HA is functionalized with moieties that can facilitate network formation through physical self-assembly, or covalent crosslinking (e.g. photo-catalyzed) at concentrations where the polysaccharide does not gel on its own. However, these crosslinks often utilize functional groups not found in biological tissues. Methods: Oscillatory rheology, dynamic light scattering, and scanning electron microscopy were used to study albumin/HA structures. Dynamic light scattering and transmission electron microscopy were used to study albumin/chondroitin sulfate (CS) structures. UV-vis spectroscopy was used to demonstrate the potential for using protein-polymer blends as an ECM-mimetic model to study transport of small molecules. Results: We examine the intermolecular interactions of two major glycosaminoglycans found in the human brain, HA and the lower molecular weight CS, with the model protein albumin. We report the properties of the resulting micro- and nano materials. Our albumin/HA systems formed gels, and albumin/CS systems formed micro- and nanoparticles. These systems are formed from unfunctionalized polysaccharides, which is an attractive and simple method of forming HA hydrogels and CS nanoparticles. We also summarize the concentrations of HA and CS found in various mammalian brains, which could potentially be useful for biomimetic scaffold development. Conclusions: Simple preparation of commercially available charged biomacromolecules results in interesting materials with structures at the micron and nanometer length-scales. Such materials may have utility in serving as cost-effective models of nervous system electrostatic interactions and as in vitro drug release and model system for ECM transport studies.

Regiospecific Nucleation and Growth of Silane Coupling Agent Droplets onto Colloidal Particles.

Nucleation-and-growth processes are used extensively in the synthesis of spherical colloids, and more recently regiospecific nucleation-and-growth processes have been exploited to prepare more complex colloids such as patchy particles. We demonstrate that surface geometry alone can be made to play the dominant role in determining the final particle geometry in such syntheses, meaning that intricate chemical surface patternings are not required. We present a synthesis method for "lollipop"-shaped colloidal heterodimers (patchy particles), combining a recently published nucleation-and-growth technique with our recent findings that particle geometry influences the locus of droplet adsorption onto anisotropic template particles. Specifically, 3-methacryloxypropyl trimethoxysilane (MPTMS) is nucleated and grown onto bullet-shaped and nail-shaped colloids. The shape of the template particle can be chosen such that the MPTMS adsorbs regiospecifically onto the flat ends. In particular, we find that particles with a wider base increase the range of droplet volumes for which the minimum in the free energy of adsorption is located at the flat end of the particle compared with bullet-shaped particles of the same aspect ratio. We put forward an extensive analysis of the synthesis mechanism and experimentally determine the physical properties of the heterodimers, supported by theoretical simulations. Here we numerically optimize, for the first time, the shape of finite-sized droplets as a function of their position on the rod-like silica particle surface. We expect that our findings will give an impulse to complex particle creation by regiospecific nucleation and growth.

Plasmonic tunnel junctions for single-molecule redox chemistry.

Nanoparticles attached just above a flat metallic surface can trap optical fields in the nanoscale gap. This enables local spectroscopy of a few molecules within each coupled plasmonic hotspot, with near thousand-fold enhancement of the incident fields. As a result of non-radiative relaxation pathways, the plasmons in such sub-nanometre cavities generate hot charge carriers, which can catalyse chemical reactions or induce redox processes in molecules located within the plasmonic hotspots. Here, surface-enhanced Raman spectroscopy allows us to track these hot-electron-induced chemical reduction processes in a series of different aromatic molecules. We demonstrate that by increasing the tunnelling barrier height and the dephasing strength, a transition from coherent to hopping electron transport occurs, enabling observation of redox processes in real time at the single-molecule level.

Smart supramolecular sensing with cucurbit[n]urils: probing hydrogen bonding with SERS.

Rigid gap nano-aggregates of Au nanoparticles formed using cucurbit[n]uril (CB[n]) molecules are used to investigate the competitive binding of ethanol and methanol in an aqueous environment. We show it is possible to detect as little as 0.1% methanol in water and a ten times higher affinity to methanol over ethanol, making this a useful technology for quality control in alcohol production. We demonstrate strong interaction effects in the SERS peaks, which we demonstrate are likely from the hydrogen bonding of water complexes in the vicinity of the CB[n]s.

Selective Depletion Interactions in Mixtures of Rough and Smooth Silica Spheres.

The depletion interaction as induced between colloids by the addition of a polymer depletant is one of the few ways in which short-ranged attractions between particles can be controlled. Due to these tunable interactions, colloid-polymer mixtures have contributed to a better understanding of the role of attractions both in equilibrium phenomena such as phase transitions and liquid surfaces as well as in systems out of equilibrium such as gelation and the glass transition. It is known that, by simple geometric effects, surface roughness decreases the strength of the depletion interaction. In this study, we demonstrate both by Monte Carlo simulations and experiments that it is possible to generate enough difference in attraction strength to induce phase separation in smooth particles but not in rough particles. Roughness was induced by coating smooth particles with smaller spherical colloids. We indicate how effective potentials can be obtained through simulations and how the interplay between gravity and the depletion interaction with a flat container wall can be used to obtain a simple measure of the interaction strengths as a function of roughness.

Site-specific growth of polymers on silica rods.

Colloids specifically developed for self-assembly (SA) into advanced functional materials have rapidly become more complex, as this complexity allows for more ways to optimize both the SA process and the properties of the resulting materials. For instance, by creating 'patchy' particles more open structures can be achieved through directional interactions. However, the number of ways in which site-specific chemistry can be achieved on particle surfaces is still limited. Here, we show how polymer patches can be specifically grown onto only the flat end of bullet-shaped silica rods by utilizing a subtle anisotropy in surface tension and shape caused by the growth mechanism of the rods. Conversely, if the bullet-shaped silica rods are used as 'Pickering-emulsion' stabilizers the same surface tension effects exclusively direct the orientation of the rods into a 'hedgehog-morphology'. Finally, we demonstrate how an external electric field can direct the particles in a 'vectorial' way.