High-Throughput All-Optical Determination of Nanorod Size and Orientation.

As a single-particle characterization technique, optical microscopy has transformed our understanding of structure-function relationships of plasmonic nanoparticles, but the need for ex-situ-correlated electron microscopy to obtain structural information handicaps an otherwise exceptional high-throughput technique. Here, we present an all-optical alternative to electron microscopy to accurately and quickly extract structural information about single gold nanorods (Au NRs) using calcite-assisted localization and kinetics (CLocK) microscopy. Color CLocK images of single Au NRs allow scattering from the longitudinal and transverse plasmon modes to be imaged simultaneously, encoding spectral data in CLocK images that can then be extracted to obtain Au NR size and orientation. Moreover, through the use of convolutional neural networks, Au NR length, width, and aspect ratio can be predicted directly from color CLocK images within ∼10% of the true value measured by electron microscopy.

Nonlinear effects in single-particle photothermal imaging.

Although photothermal imaging was originally designed to detect individual molecules that do not emit or small nanoparticles that do not scatter, the technique is now being applied to image and spectroscopically characterize larger and more sophisticated nanoparticle structures that scatter light strongly. Extending photothermal measurements into this regime, however, requires revisiting fundamental assumptions made in the interpretation of the signal. Herein, we present a theoretical analysis of the wavelength-resolved photothermal image and its extension to the large particle scattering regime, where we find the photothermal signal to inherit a nonlinear dependence upon pump intensity, together with a contraction of the full-width-at-half-maximum of its point spread function. We further analyze theoretically the extent to which photothermal spectra can be interpreted as an absorption spectrum measure, with deviations between the two becoming more prominent with increasing pump intensities. Companion experiments on individual 10, 20, and 100 nm radius gold nanoparticles evidence the predicted nonlinear pump power dependence and image contraction, verifying the theory and demonstrating new aspects of photothermal imaging relevant to a broader class of targets.

Wavelength-Dependent Photothermal Imaging Probes Nanoscale Temperature Differences among Subdiffraction Coupled Plasmonic Nanorods.

Plasmonic structures confine electromagnetic energy at the nanoscale, resulting in local, inhomogeneous, controllable heating, but reading out the temperature using optical techniques poses a difficult challenge. Here, we report on the optical thermometry of individual gold nanorod trimers that exhibit multiple wavelength-dependent plasmon modes resulting in measurably different local temperature distributions. Specifically, we demonstrate how photothermal microscopy encodes different wavelength-dependent temperature profiles in the asymmetry of the photothermal image point spread function. These asymmetries are interpreted through companion numerical simulations to reveal how thermal gradients within the trimer can be controlled by exciting its hybridized plasmon modes. We also find that plasmon modes that are optically dark can be excited by focused laser beam illumination, providing another route to modify thermal profiles beyond wide-field illumination. Taken together these findings demonstrate an all-optical thermometry technique to actively create and measure nanoscale thermal gradients below the diffraction limit.

Special topic on emerging directions in plasmonics.

Plasmonics enables a wealth of applications, including photocatalysis, photoelectrochemistry, photothermal heating, optoelectronic devices, and biological and chemical sensing, while encompassing a broad range of materials, including coinage metals, doped semiconductors, metamaterials, 2D materials, bioconjugates, and chiral assemblies. Applications in plasmonics benefit from the large local electromagnetic field enhancements generated by plasmon excitation, as well as the products of plasmon decay, including photons, hot charge carriers, and heat. This special topic highlights recent work in both theory and experiment that advance our fundamental understanding of plasmon excitation and decay mechanisms, showcase new applications enabled by plasmon excitation, and highlight emerging classes of materials that support plasmon excitation.

Super-Resolution Imaging and Plasmonics.

This review describes the growing partnership between super-resolution imaging and plasmonics, by describing the various ways in which the two topics mutually benefit one another to enhance our understanding of the nanoscale world. First, localization-based super-resolution imaging strategies, where molecules are modulated between emissive and nonemissive states and their emission localized, are applied to plasmonic nanoparticle substrates, revealing the hidden shape of the nanoparticles while also mapping local electromagnetic field enhancements and reactivity patterns on their surface. However, these results must be interpreted carefully due to localization errors induced by the interaction between metallic substrates and single fluorophores. Second, plasmonic nanoparticles are explored as image contrast agents for both superlocalization and super-resolution imaging, offering benefits such as high photostability, large signal-to-noise, and distance-dependent spectral features but presenting challenges for localizing individual nanoparticles within a diffraction-limited spot. Finally, the use of plasmon-tailored excitation fields to achieve subdiffraction-limited spatial resolution is discussed, using localized surface plasmons and surface plasmon polaritons to create confined excitation volumes or image magnification to enhance spatial resolution.

Quantifying photothermal heating at plasmonic nanoparticles by scanning electrochemical microscopy.

Photothermal heating at metal nanoparticles results from the non-radiative decay of localized surface plasmons. The local heat generation enhances the mass transport rate of redox molecules and causes a shift in their formal potential, both of which can impact an electrochemical process at the nanoparticle interface. Here we present a methodology for probing the surface temperature at a plasmonic nanoparticle substrate using scanning electrochemical microscopy (SECM). Light is used to excite a plasmonic substrate electrode, while an ultramicroelectrode tip is positioned close to the substrate to read out both the mass transfer rate and concentration profile of the redox molecules. The measured mass transfer rate and the shift in the equilibrium potential provide a quantitative value of the temperature increase at the substrate surface, which is verified by simulations using a mass transfer model coupled with heat dissipation. The developed SECM approach is suitable for probing heat generation at a variety of both plasmonic and non-plasmonic nanostructures.

Visualizing and Calculating Tip-Substrate Distance in Nanoscale Scanning Electrochemical Microscopy Using 3-Dimensional Super-Resolution Optical Imaging.

We report a strategy for the optical determination of tip-substrate distance in nanoscale scanning electrochemical microscopy (SECM) using three-dimensional super-resolution fluorescence imaging. A phase mask is placed in the emission path of our dual SECM/optical microscope, generating a double helix point spread function at the image plane, which allows us to measure the height of emitting objects relative to the focus of the microscope. By exciting both a fluorogenic reaction at the nanoscale electrode tip as well as fluorescent nanoparticles at the substrate, we are able to calculate the tip-substrate distance as the tip approaches the surface with precision better than 25 nm. Attachment of a fluorescent particle to the insulating sheath of the SECM tip extends this technique to nonfluorogenic electrochemical reactions. Correlated electrochemical and optical determination of tip-substrate distance yielded excellent agreement between the two techniques. Not only does super-resolution imaging offer a secondary feedback mechanism for measuring the tip-sample gap during SECM experiments, it also enables facile tip alignment and a strategy for accounting for electrode tilt relative to the substrate.

Investigating Nanoscale Electrochemistry with Surface- and Tip-Enhanced Raman Spectroscopy.

The chemical sensitivity of surface-enhanced Raman spectroscopy (SERS) methodologies allows for the investigation of heterogeneous chemical reactions with high sensitivity. Specifically, SERS methodologies are well-suited to study electron transfer (ET) reactions, which lie at the heart of numerous fundamental processes: electrocatalysis, solar energy conversion, energy storage in batteries, and biological events such as photosynthesis. Heterogeneous ET reactions are commonly monitored by electrochemical methods such as cyclic voltammetry, observing billions of electrochemical events per second. Since the first proof of detecting single molecules by redox cycling, there has been growing interest in examining electrochemistry at the nanoscale and single-molecule levels. Doing so unravels details that would otherwise be obscured by an ensemble experiment. The use of optical spectroscopies, such as SERS, to elucidate nanoscale electrochemical behavior is an attractive alternative to traditional approaches such as scanning electrochemical microscopy (SECM). While techniques such as single-molecule fluorescence or electrogenerated chemiluminescence have been used to optically monitor electrochemical events, SERS methodologies, in particular, have shown great promise for exploring electrochemistry at the nanoscale. SERS is ideally suited to study nanoscale electrochemistry because the Raman-enhancing metallic, nanoscale substrate duly serves as the working electrode material. Moreover, SERS has the ability to directly probe single molecules without redox cycling and can achieve nanoscale spatial resolution in combination with super-resolution or scanning probe microscopies. This Account summarizes the latest progress from the Van Duyne and Willets groups toward understanding nanoelectrochemistry using Raman spectroscopic methodologies. The first half of this Account highlights three techniques that have been recently used to probe few- or single-molecule electrochemical events: single-molecule SERS (SMSERS), superlocalization SERS imaging, and tip-enhanced Raman spectroscopy (TERS). While all of the studies we discuss probe model redox dye systems, the experiments described herein push the study of nanoscale electrochemistry toward the fundamental limit, in terms of both chemical sensitivity and spatial resolution. The second half of this Account discusses current experimental strategies for studying nanoelectrochemistry with SERS techniques, which includes relevant electrochemically and optically active molecules, substrates, and substrate functionalization methods. In particular, we highlight the wide variety of SERS-active substrates and optically active molecules that can be implemented for EC-SERS, as well as the need to carefully characterize both the electrochemistry and resultant EC-SERS response of each new redox-active molecule studied. Finally, we conclude this Account with our perspective on the future directions of studying nanoscale electrochemistry with SERS/TERS, which includes the integration of SECM with TERS and the use of theoretical methods to further describe the fundamental intricacies of single-molecule, single-site electrochemistry at the nanoscale.

Chemically Driven Interfacial Coupling in Charge-Transfer Mediated Functional Superstructures.

Organic charge-transfer superstructures are enabling new interfacial electronics, such as organic thermoelectrics, spin-charge converters, and solar cells. These carbon-based materials could also play an important role in spin-based electronics due to their exceptionally long spin lifetime. However, to explore these potentials a coherent design strategy to control interfacial charge-transfer interaction is indispensable. Here we report that the control of organic crystallization and interfacial electron coupling are keys to dictate external stimuli responsive behaviors in organic charge-transfer superstructures. The integrated experimental and computational study reveals the importance of chemically driven interfacial coupling in organic charge-transfer superstructures. Such degree of engineering opens up a new route to develop a new generation of functional charge-transfer materials, enabling important advance in all organic interfacial electronics.

Unforeseen distance-dependent SERS spectroelectrochemistry from surface-tethered Nile Blue: the role of molecular orientation.

Covalent immobilization of redox-active dyes is an important strategy to evaluate structure-activity relationships in nanoscale electrochemistry by using optical readouts such as surface-enhanced Raman scattering (SERS). Here we investigate the role of the tether length in the SERS spectroelectrochemistry of surface-attached Nile Blue. Differential pulse voltammetry and a potential-dependent SERS derivative analysis reveal that the Nile Blue molecules adopt a different orientation with respect to the electrode surface as the number of carbons in a carboxylic acid-terminated alkanethiol monolayer is varied, which leads to unique SERS spectroelectrochemical behaviors. We use the relative molecular orientations and spectral characteristics to propose a model in which tethers shorter than the length of the molecule limit molecular motion under electrochemical perturbation, but tethers longer than the length of the molecule allow dye intercalation into the hydrophobic self-assembled monolayer, producing an unexpected decrease in the SERS intensity when the molecule is in the oxidized form.

Imaging Electrogenerated Chemiluminescence at Single Gold Nanowire Electrodes.

We report electrogenerated chemiluminescence (ECL) generated at single gold nanowire electrodes supported on tin-doped indium oxide. Unlike other single nanoparticle electrochemical characterization techniques, ECL provides a massively parallel direct readout of electrochemical activity on individual nanoparticle electrodes without the need for extrinsic illumination or a scanning electrochemical probe. While ECL is not observed from as-purchased nanowires due to the surfactant layer, by removing the layer and coating the nanowires with a polymer blend, ECL from single nanowire electrodes is readily measured. With an increase in polymer thickness, an increase in ECL image quality and reproducibility over multiple redox cycles is observed. The polymer coating also provides a strategy for stabilizing gold nanoparticle electrodes against complete surface oxidation in aqueous environments.

Objective-Induced Point Spread Function Aberrations and Their Impact on Super-Resolution Microscopy.

This study demonstrates how different microscope objectives can lead to asymmetric imaging aberrations in the point spread function of dipolar emitters, which can adversely affect the quality of fit in super-resolution imaging. Luminescence from gold nanorods was imaged with four different objectives to measure the diffraction-limited emission and characterize deviations from the expected dipolar emission patterns. Each luminescence image was fit to a three-dipole emission model to generate fit residuals that visually relay aberrations in the point spread function caused by the different microscope objectives. Output parameters from the fit model were compared to experimentally measured values, and we find that while some objectives provide high quality fits across all nanorods studied, others show significant aberrations and are inappropriate for super-resolution imaging. This work presents a simple and robust strategy for quickly assessing the quality of point spread functions produced by different microscope objectives.

Observation of nanometer-sized electro-active defects in insulating layers by fluorescence microscopy and electrochemistry.

We report a method to study electro-active defects in passivated electrodes. This method couples fluorescence microscopy and electrochemistry to localize and size electro-active defects. The method was validated by comparison with a scanning probe technique, scanning electrochemical microscopy. We used our method for studying electro-active defects in thin TiO2 layers electrodeposited on 25 µm diameter Pt ultramicroelectrodes (UMEs). The permeability of the TiO2 layer was estimated by measuring the oxidation of ferrocenemethanol at the UME. Blocking of current ranging from 91.4 to 99.8% was achieved. Electro-active defects with an average radius ranging between 9 and 90 nm were observed in these TiO2 blocking layers. The distribution of electro-active defects over the TiO2 layer is highly inhomogeneous and the number of electro-active defect increases for lower degree of current blocking. The interest of the proposed technique is the possibility to quickly (less than 15 min) image samples as large as several hundreds of µm(2) while being able to detect electro-active defects of only a few tens of nm in radius.

Super-resolution imaging of SERS hot spots.

Surface-enhanced Raman scattering (SERS) hot spots occur when molecules are positioned near regions of strongly enhanced electromagnetic fields on the surface of nano-featured plasmonic substrates. The emission from the molecule is coupled out into the far field by the plasmon modes of the substrate, but due to the diffraction-limit of light, the properties of this coupled molecule-plasmon emitter cannot be resolved using typical far-field optical microscopy techniques. However, by fitting the emission to a model function such as 2-dimensional Gaussian, the relative position of the emitter can be determined with precision better than 5 nm in a process known as super-resolution imaging. This tutorial review describes the basic principles of super-resolution imaging of SERS hot spots using single molecules to probe local electromagnetic field enhancements. New advances using dipole-based fitting functions and spectrally- and spatially-resolved measurements are described, providing new insight into SERS hot spots and the important roles of both the molecule and the substrate in defining their properties.

Visualizing site-specific redox potentials on the surface of plasmonic nanoparticle aggregates with superlocalization SERS microscopy.

In this Letter, we demonstrate site-specific redox potentials for Nile Blue adsorbed to Ag nanoparticle electrodes using surface-enhanced Raman scattering (SERS) superlocalization microscopy. Nile Blue is electrochemically modulated between its oxidized and reduced form, which can be optically read out through a corresponding gain or loss in SERS intensity. SERS emission centroids are calculated by fitting the diffraction-limited SERS emission to a two-dimensional Gaussian to determine the approximate location of the emitter with 5-10 nm precision. With molecular coverage above the single molecule level, the SERS centroid trajectories shift reversibly with applied potential over multiple reduction and oxidation cycles. A mechanism is proposed to explain the centroid trajectories based on site-specific redox potentials on the nanoparticle electrode surface, where the first molecule reduced is the last to be oxidized, consistent with reversible electrochemical behavior of redox probes adsorbed to electrode surfaces.

Triplet-state-mediated super-resolution imaging of fluorophore-labeled gold nanorods.

Triplet-state-mediated super-resolution imaging was used to map the positions of fluorescently labeled double-stranded DNA bound to the surface of gold nanorods. In order to isolate individual fluorophores bound to the nanorod surface, imaging conditions were optimized such that the majority of the fluorophores were forced into a triplet dark state, and fluorescence from approximately one molecule at a time was detected. The fluorescence from the emitting single molecule was then fit to a two-dimensional (2D) Gaussian to localize its position relative to the nanorod substrate. The reconstructed super-resolution images showed excellent agreement with the shape and orientation of the nanorods, although, based on correlated atomic force microscopy, they consistently under-estimated nanorod size. The apparent DNA ligand binding on the gold nanorod surface showed significant heterogeneity, with examples of preferential binding to nanorod ends, uniform binding across the nanorod surface, and site-specific binding to a single end of the nanorod. This heterogeneity would be hidden in a typical ensemble or diffraction-limited measurement, highlighting the need for single nanoparticle super-resolution imaging studies.

A first-principles polarized Raman method for determining whether a uniform region of a sample is crystalline or isotropic.

Previous methods for determining whether a uniform region of a sample is crystalline or isotropic-what we call the "state of internal orientation" S-require a priori knowledge of properties of the purely crystalline and purely isotropic states. In addition, these methods can be ambiguous in their determination of state S for particular materials and, for a given material, the spectral methods can be ambiguous when using particular peaks. Using first-principles Raman theory, we have discovered a simple, non-resonance, polarized Raman method for determining the state S that requires no information a priori and will work unambiguously for any material using any vibrational mode. Similar to the concept behind "magic angle spinning" in NMR, we have found that for a special set of incident/analyzed polarizations and scattering angle, the dependence of the Raman modulation depth M on the sample composition-and, for crystalline regions, the unit cell orientation-falls out completely, leaving dependence on only whether the region is crystalline (M = 1) or isotropic (M = 0). Further, upon scanning between homogeneous regions or domains within a heterogeneous sample, our signal M is a clear detector of the region boundaries, so that when combined with methods for determining the orientations of the crystalline domains, our method can be used to completely characterize the molecular structure of an entire heterogeneous sample to a very high certainty. Interestingly, our method can also be used to determine when a given mode is vibrationally degenerate. While simulations on realistic terthiophene systems are included to illustrate our findings, our method should apply to any type of material, including thin films, molecular crystals, and semiconductors. Finally, our discovery of these relationships required derivations of Raman intensity formulas that are at least as general as any we have found, and herein we present our comprehensive formulas for both the crystalline and isotropic states.

New tools for investigating electromagnetic hot spots in single-molecule surface-enhanced Raman scattering.

Surface-enhanced Raman scattering (SERS) is quickly growing as an analytical technique, because it offers both molecular specificity and excellent sensitivity. For select substrates, SERS can even be observed from single molecules, which is the ultimate limit of detection. This review describes recent developments in the field of single-molecule SERS (SM-SERS) with a focus on new tools for characterizing SM-SERS-active substrates and how they interact with single molecules on their surface. In particular, techniques that combine optical spectroscopy and microscopy with electron microscopy are described, including correlated optical and transmission electron microscopy, correlated super-resolution imaging and scanning electron microscopy, and correlated optical microscopy and electron energy loss spectroscopy.

Super-resolution imaging of interactions between molecules and plasmonic nanostructures.

Super-resolution far-field imaging has recently emerged as a novel strategy for imaging interactions between plasmonic nanostructures and single molecules with spatial resolution <5 nm. In these experiments, the emission centroid of a diffraction-limited spot is modeled as a two-dimensional Gaussian, allowing the position of an emitter to be determined with nanoscale precision. In this perspective, we describe the principles of super-resolution far-field imaging and then highlight its application to several different problems in plasmonics, including surface-enhanced fluorescence of ligands bound to nanoparticle surfaces, nanoparticle-mediated catalysis, and mapping electromagnetic hot spots. In all cases, the complex coupling between molecular emission and plasmon modes of the underlying nanostructure must be considered. While this complicates the interpretation of super-resolution images of plasmonic systems, the coupling also opens new doors for understanding the fundamental interactions between molecules and plasmonic nanostructures.