Plasmon-assisted chemistry is the result of a complex interplay between electromagnetic near fields, heat and charge transfer on the nanoscale. The disentanglement of their roles is non-trivial. Therefore, a thorough knowledge of the chemical, structural and spectral properties of the plasmonic/molecular system being used is required. Specific techniques are needed to fully characterize optical near fields, temperature and hot carriers with spatial, energetic and/or temporal resolution. The timescales for all relevant physical and chemical processes can range from a few femtoseconds to milliseconds, which necessitates the use of time-resolved techniques for monitoring the underlying dynamics. In this Review, we focus on experimental techniques to tackle these challenges. We further outline the difficulties when going from the ensemble level to single-particle measurements. Finally, a thorough understanding of plasmon-assisted chemistry also requires a substantial joint experimental and theoretical effort. [Figure not available: see fulltext.] © 2022, Springer Nature Limited.

Gold nanorods (AuNRs) are promising agents for diverse biomedical applications such as drug and gene delivery, bioimaging, and cancer treatment. Upon in vivo application, AuNRs quickly interact with cells of the immune system. On the basis of their strong intrinsic phagocytic activity, polymorphonuclear neutrophils (PMNs) are specifically equipped for the uptake of particulate materials such as AuNRs. Therefore, understanding the interaction of AuNRs with PMNs is key for the development of safe and efficient therapeutic applications. In this study, we investigated the uptake, intracellular processing, and cell biological response induced by AuNRs in PMNs. We show that uptake of AuNRs mainly occurs via phagocytosis and macropinocytosis with rapid deposition of AuNRs in endosomes within 5 min. Within 60 min, AuNR uptake induced an unfolded protein response (UPR) along with induction of inositol-requiring enzyme 1 α (IREα) and features of endoplasmic reticulum (ER) stress. This early response was followed by a pro-inflammatory autocrine activation loop that involves LOX1 upregulation on the cell surface and increased secretion of IL8 and MMP9. Our study provides comprehensive mechanistic insight into the interaction of AuNRs with immune cells and suggests potential targets to limit the unwanted immunopathological activation of PMNs during application of AuNRs. © 2022 American Chemical Society.

The central dilemma in label-free in situ surface-enhanced Raman scattering (SERS) for monitoring of heterogeneously catalyzed reactions is the need of plasmonically active nanostructures for signal enhancement. Here, we show that the assembly of catalytically active transition-metal nanoparticles into dimers boosts their intrinsically insufficient plasmonic activity at the monomer level by several orders of magnitude, thereby enabling the in situ SERS monitoring of various important heterogeneously catalyzed reactions at the single-dimer level. Specifically, we demonstrate that Pd nanocubes (NCs), which alone are not sufficiently plasmonically active as monomers, can act as a monometallic yet bifunctional platform with both catalytic and satisfactory plasmonic activity via controlled assembly into single dimers with an ∼1 nm gap. Computer simulations reveal that the highest enhancement factors (EFs) occur at the corners of the gap, which has important implications for the SERS-based detection of catalytic conversions: it is sufficient for molecules to come in contact with the "hot spot corners", and it is not required that they diffuse deeply into the gap. For the widely employed Pd-catalyzed Suzuki-Miyaura cross-coupling reaction, we demonstrate that such Pd NC dimers can be employed for in situ kinetic SERS monitoring, using a whole series of aryl halides as educts. Our generic approach based on the controlled assembly into dimers can easily be extended to other transition-metal nanostructures. © 2022 American Chemical Society.

DNA-scaffolded enzymes typically show altered kinetic properties; however, the mechanism behind this phenomenon is still poorly understood. We address this question using thrombin, a model of allosterically regulated serine proteases, encaged into DNA origami cavities with distinct structural and electrostatic features. We compare the hydrolysis of substrates that differ only in their net charge due to a terminal residue far from the cleavage site and presumably involved in the allosteric activation of thrombin. Our data show that the reaction rate is affected by DNA/substrate electrostatic interactions, proportionally to the degree of DNA/enzyme tethering. For substrates of opposite net charge, this leads to an inversion of the catalytic response of the DNA-scaffolded thrombin when compared to its freely diffusing counterpart. Hence, by altering the electrostatic environment nearby the encaged enzyme, DNA nanostructures interfere with charge-dependent mechanisms of enzyme-substrate recognition and may offer an alternative tool to regulate allosteric processes through spatial confinement. © 2022 The Authors.

A detailed study of the adsorption structure of self-assembled monolayers of 4-nitrothiophenol on the Au(111) surface was performed from a theoretical perspective via first-principles density functional theory calculations and experimentally by Raman and vibrational sum frequency spectroscopy (vSFS) with an emphasis on the molecular orientation. Simulations—including an explicit van der Waals (vdW) description—for different adsorbate structures, namely, for (3×3), (2 × 2), and (3 × 3) surface unit cells, reveal a significant tilting of the molecules toward the surface with decreasing coverage from 75° down to 32° tilt angle. vSFS suggests a tilt angle of 50°, which agrees well with the one calculated for a structure with a coverage of 0.25. Furthermore, calculated vibrational eigenvectors and spectra allowed us to identify characteristic in-plane (NO2 scissoring) and out-of-plane (C-H wagging) modes and to predict their strength in the spectrum in dependence of the adsorption geometry. We additionally performed calculations for biphenylthiol and terphenylthiol to assess the impact of multiple aromatic rings and found that vdW interactions are significantly increasing with this number, as evidenced by the absorption energy and the molecule adopting a more upright-standing geometry. © 2021 Author(s).

Several fields of applications require a reliable characterization of the photothermal response and heat dissipation of nanoscopic systems, which remains a challenging task for both modeling and experimental measurements. Here, we present an implementation of anti-Stokes thermometry that enables the in situ photothermal characterization of individual nanoparticles (NPs) from a single hyperspectral photoluminescence confocal image. The method is label-free, potentially applicable to any NP with detectable anti-Stokes emission, and does not require any prior information about the NP itself or the surrounding media. With it, we first studied the photothermal response of spherical gold NPs of different sizes on glass substrates, immersed in water, and found that heat dissipation is mainly dominated by the water for NPs larger than 50 nm. Then, the role of the substrate was studied by comparing the photothermal response of 80 nm gold NPs on glass with sapphire and graphene, two materials with high thermal conductivity. For a given irradiance level, the NPs reach temperatures 18% lower on sapphire and 24% higher on graphene than on bare glass. The fact that the presence of a highly conductive material such as graphene leads to a poorer thermal dissipation demonstrates that interfacial thermal resistances play a very significant role in nanoscopic systems and emphasize the need for in situ experimental thermometry techniques. The developed method will allow addressing several open questions about the role of temperature in plasmon-assisted applications, especially ones where NPs of arbitrary shapes are present in complex matrixes and environments. © 2021 American Chemical Society. All rights reserved.

Accurate diagnosis relies on precise detection of trace marker molecules in bio-fluids and/or analysis of multiple disease-related molecules in cell/tissue specimens. While SERS provides ultra-sensitivity and intensive multiplexing capacity, conjugation of SERS tags with targeting molecules such as antibodies and aptamers enables specific recognition of these disease-related marker molecules. In this chapter, we will introduce recent progress of iSERS, the new technique that combines SERS-labeled targeting molecules with Raman microspectroscopy, and its applications in the field of lateral flow assay for point-of care diagnosis as well as in cell/tissue imaging for multiplexed tumor marker characterization. © 2022 Elsevier Inc. All rights reserved.

Despite extensive scientific efforts to understand the exact nature of hydrogen bonding in Pyridine (Py) aqueous solutions, the exact mechanism causing the blue-shift in Py ring breathing mode has remained unclear so far. The present study is an attempt to investigate the possible causes of the experimentally observed blue-shifts using density functional theory (DFT) by computing the complexation-induced electronic charge redistribution due to hydrogen bonding in distinct Py–water complexes. Electron density difference map (EDDM) calculations, natural population analysis (NPA), and natural bond orbital (NBO) analysis were used to explore the blue-shift in Py ring breathing mode upon complexation with water. The combined NPA and NBO analysis yielded that interaction between the nitrogen lone pair in Py and hydrogen atom of W causes a net charge transfer from Py to W, which—through hyperconjugation—depopulates the σ* (C–C) orbitals. This finally leads to the strengthening of the local C–C force constants. The relative contributions of normal coordinates were obtained by potential energy distribution (PED) calculations. Our computation reveals that increasing contributions of C–C stretching motions to the Py ring breathing mode (ν1) are de facto causing a blue-shift in the ν1 vibration upon water addition. Overall, this analysis suggests that complementary information from both (a) electronic charge redistribution and (b) potential energy distribution of normal coordinates is required to explain the blue-shift in ν1 (Py) mode in aqueous solutions. © 2021 John Wiley & Sons, Ltd.

Ultraviolet resonance Raman scattering (UVRR) has been frequently used for studying peptide and protein structure and dynamics, while applications in supramolecular chemistry are quite rare. Since UVRR offers the additional advantages of chromophore selectivity and high sensitivity compared with conventional non-resonant Raman scattering, it is ideally suited for label-free probing of relatively small artificial/supramolecular ligands exhibiting electronic resonances in the UV. In this perspective article, we first summarize results of UVRR spectroscopy in supramolecular chemistry in the context of peptide/protein recognition. We focus on selected artificial ligands which were rationally designed as selective carboxylate binders (guanidiniocarbonyl pyrrole, GCP, and guanidiniocarbonyl indole, GCI) and selective lysine binder (molecular tweezer, CLR01), respectively, via a combination of non-covalent interactions involving electrostatics, hydrogen bonding, and hydrophobic effects/van der Waals forces. Current limitations of applying UVRR as a universally applicable method for label-free and site-specific probing of molecular recognition between supramolecular ligands and proteins are highlighted. We then propose solutions to overcome these limitations for transforming UVRR spectroscopy into a generic tool in supramolecular chemistry on proteins, with an emphasis on mono- and multivalent GCP- and GCI-based ligands. Finally, we outline specific cases of supramolecular ligands such as molecular tweezers where alternative approaches such as laser-based mid-IR spectroscopy are required since UVRR can intrinsically not provide the required molecular information. © 2021 The Authors

As an immune response to COVID-19 infection, patients develop SARS-CoV-2-specific IgM/IgG antibodies. Here, we compare the performance of a conventional lateral flow assay (LFA) with a surface-enhanced Raman scattering (SERS)-based LFA test for the detection of SARS-CoV-2-specific IgM/IgG in sera of COVID-19 patients. Sensitive detection of IgM might enable early serological diagnosis of acute infections. Rapid detection in serum using a custom-built SERS reader is at least an order of magnitude more sensitive than the conventional LFAs with naked-eye detection. For absolute quantification and the determination of the limit of detection (LOD), a set of reference measurements using purified (total) IgM in buffer was performed. In this purified system, the sensitivity of SERS detection is even 7 orders of magnitude higher: the LOD for SERS was ca. 100 fg/mL compared to ca. 1 μg/mL for the naked-eye detection. This outlines the high potential of SERS-based LFAs in point-of-care testing once the interference of serum components with the gold conjugates and the nitrocellulose membrane is minimized. © 2021 American Chemical Society

Bare gold nanocubes and nanospheres with different sizes are incorporated into a rationally designed 3D DNA origami box. The encaged particles expose a gold surface accessible for subsequent site-specific functionalization, for example, for applications in molecular plasmonics such as SERS or SEF. © The Royal Society of Chemistry 2021.

Ultraviolet resonance Raman (UVRR) scattering is a highly sensitive and selective vibrational spectroscopic technique with a broad range of applications from polyaromatic hydrocarbons (PAHs) to biomolecular systems (peptides/proteins and nucleic acids) and catalysts. The interpretation of experimental UVRR spectra is not as straightforward as in purely vibrational Raman scattering (Placzek approximation) due to the involvement of higher lying electronic states and vibronic coupling. This necessitates the comparison with theoretical UVRR spectra computed by electronic structure calculations. Anthracene is an ideal model system for such a comparison between experiment and theory because it is rigid, symmetric, and of moderate size. By taking into account Herzberg–Teller contributions including Duschinsky effects, bulk solvent effects, and anharmonic contributions, a good qualitative agreement close to the resonance condition is achieved. The present study shows that within the framework of time-dependent density functional theory (TD-DFT), a general and robust approach for the analysis and interpretation of resonance Raman spectra of medium- to large-size molecules is available. © 2021 The Authors. Journal of Raman Spectroscopy published by John Wiley & Sons Ltd.

We present a UVRR spectroscopy setup which is equipped with a picosecond pulsed laser excitation source continuously tunable in the 210–2600 nm wavelength range. This laser source is based on a three-stage optical parametric amplifier (OPA) pumped by a bandwidth-compressed second harmonic output of an amplified Yb:KGW laser. It provides <15 cm−1 linewidth pulses below 270 nm, which is sufficient for resolving Raman lines of samples in condensed phase studies. For demonstrating the capability of this tunable setup for UVRR spectroscopy we present its application to the artificial ligand guanidiniocarbonyl pyrrole (GCP), a carboxylate binder used in peptide and protein recognition. A UVRR excitation study in the range 244–310 nm was performed for identifying the optimum laser excitation wavelength for UVRR spectroscopy of this ligand (λmax = 298 nm) at submillimolar concentrations (400 µM) in aqueous solution. The optimum UVRR spectrum is observed for laser excitation with λexc = 266 nm. Only in the relatively narrow range of λexc = 266–275 nm UVRR spectra with a sufficiently high signal-to-noise ratio and without severe interference from autofluorescence (AF) were detectable. At longer excitation wavelengths the UVRR signal is masked by AF. At shorter excitation wavelengths the UVRR spectrum is sufficiently separated from the AF, but the resonance enhancement is not sufficient. The presented tunable UVRR setup provides the flexibility to also identify optimum conditions for other supramolecular ligands for peptide/protein recognition. © 2020

There is an urgent clinical need for multicolor imaging of single cancer cells (no ensemble averaging) for identifying heterogenous expression of predictive biomarkers. Specifically, the comprehensive characterization of single disseminated tumor cells (sDTCs) responsible for metastatic relapse is the key to personalized therapy for patients. Current bioimaging methods lack the necessary multicolor capacity and suffer from background/autofluorescence. Both these central limitations can be overcome by immuno-SERS microscopy using SERS nanotags conjugated to antibodies. Here, we demonstrate the proof of concept for 6-color iSERS microscopy on the same single cancer cell. Human epidermal growth factor receptor 2 (HER2), the most prominent breast cancer marker, is localized on the membrane of single SkBr-3 cells, which overexpress HER2 and are an accepted model for sDTCs in breast cancer. This work paves the way for future multicolor/multitarget imaging for characterizing heterogeneous protein expression at the single-cell level. Copyright © 2020 American Chemical Society.

The variable configuration of Raman spectroscopic platforms is one of the major obstacles in establishing Raman spectroscopy as a valuable physicochemical method within real-world scenarios such as clinical diagnostics. For such real world applications like diagnostic classification, the models should ideally be usable to predict data from different setups. Whether it is done by training a rugged model with data from many setups or by a primary-replica strategy where models are developed on a 'primary' setup and the test data are generated on 'replicate' setups, this is only possible if the Raman spectra from different setups are consistent, reproducible, and comparable. However, Raman spectra can be highly sensitive to the measurement conditions, and they change from setup to setup even if the same samples are measured. Although increasingly recognized as an issue, the dependence of the Raman spectra on the instrumental configuration is far from being fully understood and great effort is needed to address the resulting spectral variations and to correct for them. To make the severity of the situation clear, we present a round robin experiment investigating the comparability of 35 Raman spectroscopic devices with different configurations in 15 institutes within seven European countries from the COST (European Cooperation in Science and Technology) action Raman4clinics. The experiment was developed in a fashion that allows various instrumental configurations ranging from highly confocal setups to fibre-optic based systems with different excitation wavelengths. We illustrate the spectral variations caused by the instrumental configurations from the perspectives of peak shifts, intensity variations, peak widths, and noise levels. We conclude this contribution with recommendations that may help to improve the inter-laboratory studies. © 2020 American Chemical Society.

Programmed cell death-ligand 1 (PD-L1) is an important predictive biomarker. The detection of PD-L1 can be crucial for patients with advanced cancer where the use of immunotherapy is considered. Here, we demonstrate the use of immuno-SERS microscopy (iSERS) for localizing PD-L1 on single cancer SkBr-3 cells. A central advantage of iSERS is that the disturbing autofluorescence from cells and tissues can be efficiently minimized by red to near-infrared laser excitation. In this study we employed Au/Au core/satellite nanoparticles as SERS nanotags because of their remarkable signal brightness and colloidal stability upon red laser excitation. False-color iSERS images of the positive and negative controls clearly reveal the specific localization of PD-L1 with SERS nanotag-labeled antibodies. © 2019 The Authors. Journal of Biophotonics published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

The discovery of the enhancement of Raman scattering by molecules adsorbed on nanostructured metal surfaces is a landmark in the history of spectroscopic and analytical techniques. Significant experimental and theoretical effort has been directed toward understanding the surface-enhanced Raman scattering (SERS) effect and demonstrating its potential in various types of ultrasensitive sensing applications in a wide variety of fields. In the 45 years since its discovery, SERS has blossomed into a rich area of research and technology, but additional efforts are still needed before it can be routinely used analytically and in commercial products. In this Review, prominent authors from around the world joined together to summarize the state of the art in understanding and using SERS and to predict what can be expected in the near future in terms of research, applications, and technological development. This Review is dedicated to SERS pioneer and our coauthor, the late Prof. Richard Van Duyne, whom we lost during the preparation of this article. © 2020 American Chemical Society. All rights reserved.

The characteristic vibrational spectroscopic fingerprint of Raman reporter molecules adsorbed on noble metal nanoparticles is employed for the identification of target proteins by the corresponding surface-enhanced Raman scattering (SERS) nanotag-labeled antibodies. Here, we present the modular synthesis of thiolated polyenes with two to five C═C double bonds introduced via stepwise Wittig reactions. The experimental characterization of their electronic and vibrational properties is complemented by density functional theory calculations. Highly SERS-active nanotags are generated by using the thiolated polyenes as Raman reporter molecules in Au/Au core/satellite supraparticles with multiple hot spots. The cytokines IL-1β and IFN-γ are detected in a duplex SERS-based lateral flow assay on a nitrocellulose test strip by Raman microscopy. The thiolated polyenes are suitable for use in immuno-SERS applications such as point-of-care testing as well as cellular and tissue imaging. © 2020 The Authors. Journal of Biophotonics published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Surfactant-free silicon nanoparticles of a predefined and narrow (σ < 10 nm) size range can be selectively immobilized on a substrate by optical printing from a polydisperse colloidal suspension by tuning the light wavelength to their size-dependent magnetic dipolar resonance. © 2020 SPIE.

Experimental results obtained in different laboratories world-wide by researchers using surface-enhanced Raman scattering (SERS) can differ significantly. We, an international team of scientists with long-standing expertise in SERS, address this issue from our perspective by presenting considerations on reliable and quantitative SERS. The central idea of this joint effort is to highlight key parameters and pitfalls that are often encountered in the literature. To that end, we provide here a series of recommendations on: a) the characterization of solid and colloidal SERS substrates by correlative electron and optical microscopy and spectroscopy, b) on the determination of the SERS enhancement factor (EF), including suitable Raman reporter/probe molecules, and finally on c) good analytical practice. We hope that both newcomers and specialists will benefit from these recommendations to increase the inter-laboratory comparability of experimental SERS results and further establish SERS as an analytical tool. © 2019 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.

Ultraviolet resonance Raman (UVRR) spectroscopy is a powerful vibrational spectroscopic technique for the label-free monitoring of molecular recognition of peptides or proteins with supramolecular ligands such as guanidiniocarbonyl pyrroles (GCPs). The use of UV laser excitation enables Raman binding studies of this class of supramolecular ligands at submillimolar concentrations in aqueous solution and provides a selective signal enhancement of the carboxylate binding site (CBS). A current limitation for the extension of this promising UVRR approach from peptides to proteins as binding partners for GCPs is the UV-excited autofluorescence from aromatic amino acids observed for laser excitation wavelengths >260 nm. These excitation wavelengths are in the electronic resonance with the GCP for achieving both a signal enhancement and the selectivity for monitoring the CBS, but the resulting UVRR spectrum overlaps with the UV-excited autofluorescence from the aromatic binding partners. This necessitates the use of a laser excitation <260 nm for spectrally separating the UVRR spectrum of the supramolecular ligand from the UV-excited autofluorescence of the peptide or protein. Here, we demonstrate the use of UVRR spectroscopy with 244 nm laser excitation for the characterization of GCP as well as guanidiniocarbonyl indole (GCI), a next generation supramolecular ligand for the recognition of carboxylates. For demonstrating the feasibility of the UVRR binding studies without an interference from the disturbing UV-excited autofluorescence, benzoic acid (BA) was chosen as an aromatic binding partner for GCI. We also present the UVRR results from the binding of GCI to the ubiquitous RGD sequence (arginylglycylaspartic acid) as a biologically relevant peptide. In the case of RGD, the more pronounced differences between the UVRR spectra of the free and complexed GCI (1:1 mixture) clearly indicate a stronger binding of GCI to RGD compared with BA. A tentative assignment of the experimentally observed changes upon molecular recognition is based on the results from density functional theory (DFT) calculations. © 2020 Beilstein-Institut Zur Forderung der Chemischen Wissenschaften. All rights reserved.

We investigated the suitability of immuno-SERS (iSERS) microscopy for imaging of smooth muscle cells (SMCs) in atherosclerotic plaques. Localization of SMCs is achieved by using SERS-labelled antibodies direct against alpha-smooth muscle actin (SMA). The staining quality of the false-colour iSERS images obtained by confocal Raman microscopy with point mapping is compared with wide-field immunofluorescence images. Both direct (labelled primary antibody) and indirect iSERS staining (unlabelled primary and labelled secondary antibody) techniques were employed. Direct iSERS staining yields results comparable to indirect IF staining, demonstrating the suitability of iSERS in research on atherosclerosis and paving the way for future multiplexed imaging experiments. © 2019 Elsevier B.V.

Vibrational sum frequency spectra (vSFS) of 4-nitrothiophenol (4-NTP) have been utilized to study plasmonic effects arising from its interaction with gold nanoparticles (Au NPs). To this end three systems have been studied: 4-NTP adsorbed on deposited Au nanoparticles, a self-assembled monolayer of 4-NTP on flat Au with dispersed Au NPs atop and, as reference, a self-assembled monolayer of 4-NTP. For 4-NTP on 50 and 80 nm Au NPs an enhancement of the SF intensity due to plasmonic effects by less than 1 order of magnitude is inferred in comparison to the monolayer system after taking the particle density into account. The addition of Au NPs to the monolayer system on flat Au also selectively enhances the response to an in-plane field component of the 532 nm upconversion light. © 2019 American Chemical Society.

Monomers and dimers of spherical gold nanoparticles (NPs) exhibit highly uniform plasmonic properties at the single-particle level due to their high structural homogeneity (precision plasmonics). Recent investigations in precision plasmonics have largely focused on static properties using conventional techniques such as transmission electron microscopy and optical dark-field microscopy. In this Feature Article, we first highlight the application of femtosecond time-resolved electron diffraction for monitoring the nonequilibrium dynamics of spherical gold NPs after ultrafast optical excitation. The analysis of the transient diffraction patterns allows us to directly obtain quantitative information on the incoherent excitation of the lattice, that is, heating upon electron-lattice equilibration, as well as on the development of strain due to lattice expansion on picosecond time scales. The controlled assembly of two spherical gold NPs into a dimer with a few nanometers gap leads to unique optical properties. Specifically, extremely high electric fields (hot spot) in the gap are generated upon resonant optical excitation. Conventional optical microscopy cannot spatially resolve this unique hot spot due to the optical diffraction limit. We therefore employed nonlinear photoemission electron microscopy to visualize hot spots in single dimers of spherical gold NPs. A quantitative comparison of different single dimers confirms the homogeneity of the hot spots on the single-particle level. Overall, these initial results are highly encouraging because they pave the way to investigate nonequilibrium dynamics in highly uniform plasmonic nanostructures at the time and space limits. © 2019 American Chemical Society.

The design of a portable Raman/SERS-LFA reader with line illumination using a custom-made fiber optic probe for rapid, quantitative, and ultrasensitive point-of-care testing (POCT) is presented. The pregnancy hormone human chorionic gonadotropin (hCG) is detectable in clinical samples within only 2–5 s down to approximately 1.6 mIU mL−1. This acquisition time is several orders of magnitude shorter than those of existing approaches requiring expensive Raman instrumentation, and the method is 15-times more sensitive than a commercially available lateral flow assay (LFA) as the gold standard. The SERS-LFA technology paves the way for affordable, quantitative, and ultrasensitive POCT with multiplexing potential in real-world applications, ranging from clinical chemistry to food and environmental analysis as well as drug and biowarfare agent testing. © 2018 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.

Silicon nanoparticles possess unique size-dependent optical properties due to their strong electric and magnetic resonances in the visible range. However, their widespread application has been limited, in comparison with other (e.g., metallic) nanoparticles, because their preparation on monodisperse colloids remains challenging. Exploiting the unique properties of Si nanoparticles in nano- and microdevices calls for methods able to sort and organize them from a colloidal suspension onto specific positions of solid substrates with nanometric precision. We demonstrate that surfactant-free silicon nanoparticles of a predefined and narrow (σ < 10 nm) size range can be selectively immobilized on a substrate by optical printing from a polydisperse colloidal suspension. The size selectivity is based on differential optical forces that can be applied on nanoparticles of different sizes by tuning the light wavelength to the size-dependent magnetic dipolar resonance of the nanoparticles. © 2019 American Chemical Society.

Plasmonic hot carriers have been recently identified as key elements for photocatalysis at visible wavelengths. The possibility to transfer energy between metal plasmonic nanoparticles and nearby molecules depends not only on carrier generation and collection efficiencies but also on their energy at the metal-molecule interface. Here an energy screening study was performed by monitoring the aniline electro-polymerization reaction via an illuminated 80 nm gold nanoparticle. Our results show that plasmon excitation reduces the energy required to start the polymerization reaction as much as 0.24 eV. Three possible photocatalytic mechanisms were explored: the enhanced near field of the illuminated particle, the temperature increase at the metal-liquid interface, and the excited electron-hole pairs. This last phenomenon is found to be the one contributing most prominently to the observed energy reduction. Copyright © 2019 American Chemical Society.

The surface plasmon (SP) coupling in dimers of spherical and faceted gold nanoparticles was investigated experimentally and computationally at the single-particle level. Individual ideal dimers of two spherical gold nanoparticles with a constant gap, filled by a self-assembled monolayer of 1,8-octanedithiol linker molecules, exhibit highly uniform dark-field (DF) scattering spectra. In contrast, single nonideal dimers of two faceted gold nanoparticles with the same constant gap exhibit a high degree of spectral nonuniformity. We attribute this significant spectral heterogeneity to the many possible gap morphologies, that is, the many possible configurations of the crystal facet orientations at the junction of the two faceted particles in nonideal dimers. This configurational complexity is reduced in so-called hybrid dimers, which comprise an ideal spherical particle and a nonideal faceted particle. Hybrid dimers were therefore investigated theoretically and experimentally for revealing the influence of crystal facet orientation in the gap. Computed scattering spectra of ideal dimers (two spheres) and three different configurations of hybrid dimers (sphere and icosahedron) with face, edge, or point contact, respectively, were obtained using the finite-difference time-domain method. Theory predicts a blue shift of the longitudinal bonding dipolar plasmon coupling mode for hybrid dimers with point and edge contacts, but a red shift for face contact due to the stronger SP coupling. Experimental DF scattering spectra of individual hybrid dimers were measured for comparison with the predictions from theory. All single-particle DF scattering spectra of hybrid dimers exhibit either a blue shift (two distinct classes) or a red shift relative to the corresponding ideal dimers, in agreement with the predictions from computer simulations. Copyright © 2019 American Chemical Society.

The photochromic molecule 2-(2,4-dinitro-benzyl)-pyridine (α-DNBP) is characterized in solution by a combination of density functional theory employing a polarizable continuum model and polarization-resolved spontaneous and nonlinear Raman spectroscopies. By the comparison of theoretically predicted wavenumber positions and depolarization ratios with the experimental spectra acquired under electronically nonresonant conditions, polarized and depolarized Raman bands are assigned. Specifically, the symmetric stretching vibrations of the two nitro groups in ortho and para positions to the pyridine ring can be experimentally differentiated, mainly because of their different Raman depolarization ratios, which supports our prediction from theory. Compared to the polarization-resolved spontaneous Raman experiments, the vibrational spectroscopic differentiation of the two nitro groups is more pronounced in time-delayed polarization-resolved coherent anti-Stokes Raman scattering experiments. Overall, this linear and nonlinear vibrational spectroscopic characterization of the CH form paves the way for the interpretation of future time-resolved pump/nonlinear Raman probe studies on the ultrafast photoinduced intramolecular proton transfer in α-DNBP involving a nitro group as an intramolecular proton acceptor. © 2019 American Chemical Society.

Immunohistochemical analysis of formalin-fixed paraffin-embedded (FFPE) tissues provides important diagnostic and prognostic information in pathology. Metal nanoparticles (NPs) and, in particular, surface-enhanced Raman scattering (SERS) nanotags as a new class of labeling reagents are promising to be used for multiplexed protein profiling on tissue sections. However, nonspecific binding of NPs onto the tissue specimens greatly hampers their clinical applications. In this study, we found that the antigen retrieval method strongly influences the extent of nonspecific binding of the antibody-SERS NP conjugates to the tissue. Our SERS labels comprised ca. 70 nm Au nanostars coated with ethylene glycol-modified Raman reporter molecules for hydrophilic stabilization and subsequent covalent bioconjugation to antibodies. We systematically investigated the influence of heat- and protease-induced epitope retrieval (HIER and PIER, respectively) on the immunostaining quality of prostate-specific antigen (PSA) on human prostate tissue sections. The best staining results were obtained with PIER. Pretreatment of the tissue sections by HIER led to selective but nonspecific adsorption of the antibody-Au nanostar conjugates onto epithelial cells, while enzymatic treatment within PIER did not. In addition to gold nanostars, also other types of metal NPs with different shapes and sizes (including ca. 20 nm quasi-spherical Au NPs and ca. 60 nm quasi-spherical Au/Ag nanoshells) as well as tissue sections from different organs (including prostate and breast) were tested; in each case the same tendency was observed, i.e., PIER yielded better results than HIER. Therefore, we recommend PIER for future NP-based tissue immunostaining such as immuno-SERS microscopy. Alternatively, for antigens that can only be unmasked by heating, PEGylation of the NPs is recommended to avoid nonspecific binding. © 2017 American Chemical Society.

Ideal dimers comprising gold nanoparticles with a smooth surface and high sphericity are synthesized by a substrate-based assembly strategy with efficient cetyltrimethylammonium bromide removal. An unprecedented structural and plasmonic uniformity at the single-particle level is observed since inhomogeneities resulting from variations in gap morphology are eliminated. Single ideal dimers are analyzed by polarization-resolved dark-field scattering spectroscopy. Contributions from transverse as well as quadrupolar and octupolar longitudinal plasmon coupling modes can be discriminated because of their orthogonal polarization behavior. The assignment of these higher order coupling modes is supported by computer simulations. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Immuno-SERS (iSERS) microscopy is an emerging imaging technique in tissue-based cancer diagnostics, which is based on antibodies labelled with SERS-active noble metal nanoparticles (NPs) in conjunction with Raman microspectroscopy for localizing target proteins. Advantages of SERS over existing labeling approaches include its high capacity for spectral multiplexing (parallel detection of target molecules), quantification (based on the characteristic SERS signatures), high photostability (no or minimal photobleaching), minimization of autofluorescence from biological specimens (via red to near-infrared excitation), and the technical advantage of using only a single laser excitation line. In this book chapter, we will first give a very brief tutorial on the fundamentals of SERS, followed by an introduction into the concept and current designs of target-specific SERS probes based on noble metal NPs. Next, the fast developing applications of iSERS microscopy for tissue-based cancer diagnostics are highlighted, and finally the challenges and future perspectives of this emerging field are presented. © Springer International Publishing AG 2018.

Ultraviolet resonance Raman (UVRR) spectroscopy is a selective, sensitive and label-free vibrational spectroscopic technique. Here, we demonstrate as proof of concept that UVRR can be used for probing the recognition between a multivalent supramolecular ligand and acidic residues in leucine zipper, an α-helical structural motif of many proteins. © 2017 Owner Societies.

Plasmonic metal nanostructures are widely used as subwavelength light concentrators to enhance light harvesting of organic solar cells through two photophysical effects, including enhanced local electric field (ELEF) and antenna-amplified light scattering (AALS), while their adverse quenching effect from surface energy transfer (SET) should be suppressed. In this work, a comprehensive study to unambiguously distinguish and quantitatively determine the specific influence and contribution of each effect on the overall performance of organic solar cells incorporated with Ag@SiO2 core–shell nanoparticles (NPs) is presented. By investigating the photon conversion efficiency (PCE) as a function of the SiO2 shell thickness, a strong competition between the ELEF and SET effects in the performance of the devices with the NPs embedded in the active layers is found, leading to a maximum PCE enhancement of 12.4% at the shell thickness of 5 nm. The results give new insights into the fundamental understanding of the photophysical mechanisms responsible for the performance enhancement of plasmonic organic solar cells and provide important guidelines for designing more-efficient plasmonic solar cells in general. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Many applications of nano- and microparticles require molecular functionalization. Assessing the heterogeneity of a colloidal sample in terms of its molecular functionalization is highly desirable but not accessible by conventional ensemble experiments. Retrieving this information necessitates single-particle experiments which simultaneously detect both functionalized and nonfunctionalized particles via two separate imaging channels. In this contribution, we present an optical setup for performing correlative single-particle imaging using laser light-sheet illumination: the first detection channel records elastic light scattering (Rayleigh/Mie), while the second channel detects inelastic light scattering (Raman) or fluorescence. The instrument is tested with Raman reporter-functionalized SERS-active metal nanoparticles (core/satellite silver nanoparticles, dimers and monomers of gold nanoparticles) and fluorophore-functionalized colloids (fluorescent polymer microparticles, dye-labeled protein on gold nanoparticles). © 2017 American Chemical Society.

The in situ detection of reactions catalyzed by metal NPs is challenging because the underlying chemical transformations occur at interfaces. Surface-enhanced Raman scattering (SERS), a surface-selective, sensitive and label-free vibrational spectroscopic technique, is ideally suited for monitoring of heterogeneous catalysis with high chemical specificity. A major limitation in the past, however, was that small, catalytically active metal NPs do not exhibit the high plasmonic activity required for SERS. This feature article focuses on the design, synthesis and use of bifunctional NPs with both catalytic and plasmonic activity for in situ SERS detection of reactions catalyzed by metal NPs. We focus on model reactions induced by chemical reducing agents such as hydride or molecular hydrogen as well as on plasmon-induced photo-catalysis including both photo-oxidation and photo-reduction. Finally, we highlight the concept of photo-recycling on halide-containing silver surfaces for unprecedented multi-electron reduction chemistry. © 2018 The Royal Society of Chemistry.

A reduction in the number of loss decay channels present in optical nanoantennas could help enhance an emitter's radiation efficiency. These losses get amplified for emitters in close proximity to metallic surfaces, such as for self-assembled monolayers, reducing the fluorescence rate. However, such a proximity strongly enhances Raman scattering. A dual-sensing scheme should bypass this shortcoming, and switching from metals to high refractive index dielectrics could aid in that direction. In order to show this, we fabricated silicon nanodimers and coated them with a β-carotenal monolayer for detecting surface-enhanced Raman scattering and fluorescence emission of the same probe. We obtained a surface-enhanced Raman scattering (SERS) factor of 1720 ± 300 for the C-C bond stretching of the polyene chain and a surface fluorescence enhancement (SEF) factor of 470 ± 90. Furthermore, our theoretical studies of different materials and emitters located on the surface of nanostructures demonstrate that low-loss dielectric materials provide a robust architecture for enhancing the response of efficient emitters. These results could have a direct impact on the development of deterministic high-rate single-photon sources. © 2018 American Chemical Society.

A preconcentrating surface-enhanced Raman scattering (SERS) sensor for the analysis of liquid-soaked tissue, tiny liquid droplets and thin liquid films without the necessity to collect the analyte is reported. The SERS sensor is based on a block-copolymer membrane containing a spongy-continuous pore system. The sensor's upper side is an array of porous nanorods having tips functionalized with Au nanoparticles. Capillarity in combination with directional evaporation drives the analyte solution in contact with the flat yet nanoporous underside of the SERS sensor through the continuous nanopore system toward the nanorod tips where non-volatile components of the analyte solution precipitate at the Au nanoparticles. The nanorod architecture increases the sensor surface in the detection volume and facilitates analyte preconcentration driven by directional solvent evaporation. The model analyte 5,5'-dithiobis(2-nitrobenzoic acid) can be detected in a 1 × 10-3m solution ≈300 ms after the sensor is brought into contact with the solution. Moreover, a sensitivity of 0.1 ppm for the detection of the dissolved model analyte is achieved. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Speed is often a bottleneck in conventional Raman microscopy on biological specimens. In immuno-Raman microspectroscopy, or for short iSERS microscopy, the acquisition times per pixel have been reduced by more than one order of magnitude over the past decade since its proof of concept. Typically rather high laser power densities are employed with the intention of compensating for the shorter acquisition times, without checking the reproducibility of the results in repeated experiments on the same sample. Here, we systematically analyze this aspect at the single-cell level since it forms the basis of quantification and is very important for reinspection of the same specimen. Specifically, we investigate experimentally the role of the laser power density in conjunction with the acquisition times per pixel in a series of repeated iSERS experiments on the same single cell overexpressing the breast cancer tumor marker human epidermal growth factor receptor 2 (HER2). Confocal iSERS mapping experiments were guided by wide-field fluorescence microscopy for selecting the regions of interest. We demonstrate that the combination of ca. a 1-2 mW laser power (40× objective, NA 0.6), 50 ms acquisition time per pixel and a high EM-CCD signal gain yields highly reproducible iSERS images in a series of four repeated experiments on the same single cell. In contrast, longer acquisition times (0.8 s, no EM gain) and in particular higher laser power (4 mW up to 18 mW) densities lead to non-reproducible iSERS results due to signal degradation. © 2017 The Royal Society of Chemistry.

Nanoscale localization of electromagnetic fields near metallic nanostructures underpins the fundamentals and applications of plasmonics. The unavoidable energy loss from plasmon decay, initially seen as a detriment, has now expanded the scope of plasmonic applications to exploit the generated hot carriers. However, quantitative understanding of the spatial localization of these hot carriers, akin to electromagnetic near-field maps, has been elusive. Here we spatially map hot-electron-driven reduction chemistry with 15 nm resolution as a function of time and electromagnetic field polarization for different plasmonic nanostructures. We combine experiments employing a six-electron photo-recycling process that modify the terminal group of a self-assembled monolayer on plasmonic silver nanoantennas, with theoretical predictions from first-principles calculations of non-equilibrium hot-carrier transport in these systems. The resulting localization of reactive regions, determined by hot-carrier transport from high-field regions, paves the way for improving efficiency in hot-carrier extraction science and nanoscale regio-selective surface chemistry. © The Author(s) 2017.

An assay for Survivin, a small dimeric protein which functions as modulator of apoptosis and cell division and serves as a promising diagnostic biomarker for different types of cancer, is presented. The assay is based on switching on surface-enhanced Raman scattering (SERS) upon incubation of the Survivin protein dimer with Raman reporter-labeled gold nanoparticles (AuNP). Site-specificity is achieved by complexation of nickel-chelated N-nitrilo-triacetic acid (Ni-NTA) anchors on the particle surface by multiple histidines (His6-tag) attached to each C-terminus of the centrosymmetric protein dimer. Correlative single-particle analysis using light sheet laser microscopy enables the simultaneous observation of both elastic and inelastic light scattering from the same sample volume. Thereby, the SERS-inactive AuNP-protein monomers can be directly discriminated from the SERS-active AuNP-protein dimers/oligomers. This information, i.e. the percentage of SERS-active AuNP in colloidal suspension, is not accessible from conventional SERS experiments due to ensemble averaging. The presented correlative single-particle approach paves the way for quantitative site-specific SERS assays in which site-specific protein recognition by small chemical and in particular supramolecular ligands can be tested. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Rhodamines are widely used dyes in fluorescence and surface-enhanced Raman spectroscopy (SERS). The latter requires adsorption of the dye onto the surface of plasmonic nanostructures, a process which requires attractive molecule-surface interactions. Here, we report an experimental SERS and computational density functional theory (DFT) study investigating the role of thiol functionalization at the xanthene ring of the rhodamine in the adsorption onto gold nanoparticles. For this purpose, a new bisthiolated rhodamine derivative was rationally designed and synthesized via a PPh3/I2 reduction route. The introduction of two thiol moieties directly at the xanthene ring provides the shortest possible distance between the molecular π-system and the metal surface for maximum SERS enhancement combined with the strong Au-S interaction for chemisorption. The comparison of experimental SERS spectra obtained from gold nanostars and a film of gold nanoparticles with results from DFT calculations (molecular electrostatic potential, normal modes) suggests adsorption via the thiol groups at the xanthene moiety. © 2017 American Chemical Society.

Surface-enhanced Raman scattering (SERS) microscopy is an emerging imaging technique for tissue-based cancer diagnostics. Specifically, immuno-SERS (iSERS) microscopy employs antibodies labelled by molecularly functionalized noble metal colloids for antigen localization on tissue specimen. Spectrally resolved iSERS acquisition schemes are typically rather time-consuming when large tissue areas must be scanned. Here, we demonstrate the application of iSERS imaging guided by wide field immunofluorescence (IF) for localization of the human epidermal growth factor receptor 2 (HER2) on breast tissue sections. The addition of unlabelled anti-HER2 primary antibodies to the tissue is followed by the incubation with secondary antibodies labelled with both Alexa-647 (for IF) and hydrophilically stabilized gold nanostars coated with aromatic thiols (for iSERS). False-color iSERS images clearly reveal the different HER2 expression levels on normal and breast cancer tissue, respectively. A series of negative controls confirms that the binding specificity of the secondary antibody is maintained after conjugation to the SERS nanoparticles. © 2016 The Royal Society of Chemistry.

Surface-enhanced Raman scattering (SERS) is a promising molecular spectroscopic technique to study chemical reactions catalyzed by nobel metal nanoparticles. Herein, the Au nanostar (NS)-catalyzed hydride reduction of 4-nitrothiophenol to 4-aminothiophenol chemisorbed at the metal/solution interface was followed by SERS. By developing a simple two-step route, we demonstrated shape-controlled synthesis and morphology-dependent catalytic activity of AuNSs. The kinetic SERS monitoring of the reaction on the Au surface indicates that the catalytic activity of the AuNSs correlates with the sharpness of their tips. This is consistent with the known size-dependent catalytic activity of small AuNPs. In contrast to the complex bifunctional nanostructures reported previously, this work shows that AuNSs are bifunctional nanostructures suitable for investigating Au-catalyzed reactions. A proposed electron transfer mechanism explains the reduction of 4-nitrothiophenol to 4-aminothiophenol on AuNS tips where –NO2 groups have no direct contact with the catalyst. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Hydrides are widely used in reduction reactions. In protic solvents, their hydrolysis generates molecular hydrogen as a second reducing agent. The competition between these two parallel reduction pathways has been overlooked so far since both typically yield the same product. We investigated the platinum-catalyzed reduction of 4-nitrothiophenol to 4-aminothiophenol in aqueous sodium borohydride solution as a prominent model reaction, by using label-free SERS monitoring in a microfluidic reactor. Kinetic analysis revealed a strong pH dependence. Surprisingly, only at pH>13 the reduction is driven exclusively by sodium borohydride. This study demonstrates the potential of microfluidics-based kinetic SERS monitoring of heterogeneous catalysis in colloidal suspension. © 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

This roadmap, through the contributions of ten groups worldwide, contains different techniques, methods and materials devoted to sensing in nanomedicine. Optics is used in different ways in the detection schemes. Raman, fluorescence and infrared spectroscopies, plasmonics, second harmonic generation and optical tweezers are all used in applications from single molecule detection (both in highly diluted and in highly concentrated solutions) to single cell manipulation. In general, each optical scheme, through device miniaturization and electromagnetic field localization, exploits an intrinsic optical enhancement mechanism in order to increase the sensitivity and selectivity of the device with respect to the complex molecular construct. The materials used for detection include nanoparticles and nanostructures fabricated with different 2D and 3D lithographic methods. It is shown that sensitivity to a single molecule is already accessible whether the system under study is a single cell or a multitude of cells in a molecular mixture. Throughout the roadmap there is an attempt to foresee and to suggest future directions in this interdisciplinary field. © 2016 IOP Publishing Ltd.

Light harvesting strategy using plasmonic metal nanostructures as subwavelength light concentrators provides a highly attractive solution to enhancing the performance of dye-sensitized solar cells (DSSCs). Through comprehensive optical spectroscopy and electrical characterizations together with a theoretical analysis, we demonstrate a strong competition between the surface energy transfer induced non-radiative quenching and the plasmonic electromagnetic enhancement effect in metal-dielectric-semiconductor core-shell-shell nanoparticle doped DSSCs, a generic yet unavoidable phenomenon in all types of plasmonic solar cells. The competition of the two effects results in a non-monotonic relationship between the device efficiency and the thickness of the dielectric shell covering the metal nanoparticles, and leads to an optimal thickness for the highest power conversion efficiency. This observation is further corroborated by photoluminescence spectroscopic measurements. Our experimental results are in good agreement with the Persson model that predicts a strong energy quenching effect when the distance between the photogenerated charge carrier and the metal core is short enough. Both experiment and theory show that the localized surface plasmon resonance enhanced light harvesting efficiency is suppressed by the surface energy transfer to the metal cores for the dielectric shell thickness shorter than a characteristic value (~7 nm in our study). Our work sheds new insights into the fundamental understanding of the photophysics mechanisms of plasmonic DSSCs and could push forward the study of plasmonic solar cells in terms of device design and fabrication. © 2016 Elsevier Ltd.

Quantitative multiplexing experiments using mixtures of up to six spectrally distinct surface-enhanced Raman scattering (SERS) nanoparticle labels are demonstrated. The SERS labels comprise gold nanoparticles, Raman reporter molecules chemisorbed onto the metal surface, and a silica shell providing chemical and mechanical stability as well as spectral reproducibility. Spectral unmixing is performed with a simple least-square algorithm because the overall SERS signal of the mixtures can be represented as a linear combination of the known spectral signatures from the individual silica-protected labels. In contrast, simple spectral unmixing using unprotected SERS particles, i.e. without a silica shell, was not successful because of band shifts and alternating peak heights in the mixtures, which is indicative of perturbing interactions between the SERS labels. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Noble metals are important photocatalysts due to their ability to convert light into chemical energy. Hot electrons, generated via the non-radiative decay of localized surface plasmons, can be transferred to reactants on the metal surface. Unfortunately, the number of hot electrons per molecule is limited due to charge-carrier recombination. In addition to the reduction half-reaction with hot electrons, also the corresponding oxidation counter-half-reaction must take place since otherwise the overall redox reaction cannot proceed. Here we report on the conceptual importance of promoting the oxidation counter-half-reaction in plasmon-mediated catalysis by photorecycling in order to overcome this general limitation. A six-electron photocatalytic reaction occurs even in the absence of conventional chemical reducing agents due to the photoinduced recycling of Ag atoms from hot holes in the oxidation half-reaction. This concept of multi-electron, counter-half-reaction-promoted photocatalysis provides exciting new opportunities for driving efficient light-to-energy conversion processes. © 2015 Macmillan Publishers Limited. All rights reserved.

A fast, generic, and suspension-based route to highly SERS-active assemblies of noble metal nanoparticles (Au, Ag) with small core-satellite gaps and single-particle Raman sensitivity is presented. Rationally designed, heterobifunctional Raman reporters serve as molecular linkers for electrostatic conjugation of the small satellites to the large core. © 2015 the Owner Societies.

Simultaneous detection of multiple molecular targets can greatly facilitate early diagnosis and drug discovery. Encoding micron-sized beads with optically active tags is one of the most popular methods to achieve multiplexing. Noble metal nanoparticle labels for optical detection by surface-enhanced Raman spectroscopy (SERS) exhibit narrow bandwidths, high photostability and intense Raman signals. In this study, we demonstrate the feasibility of spectral multiplexing by SERS using micron-sized polystyrene (PS) beads loaded with SERS-active nanoparticles. The silica-encapsulated SERS nanotags comprise gold nanocrystals with a self-assembled monolayer (SAM) of aromatic thiols as Raman reporter molecules for spectral identification. SERS microspectroscopic images of single Raman-encoded PS microbeads indicate the homogeneous spatial distribution of the SERS-active nanoparticles on the surface of the beads. By using up to five different Raman reporters, 31 spectrally distinct micron-sized beads were encoded and characterized spectroscopically at the single-bead level. This journal is © The Royal Society of Chemistry 2015.

The redox reaction of superoxide (KO<inf>2</inf>) with highly charged iron porphyrins (Fe(P4+), Fe(P8+), and Fe(P8-)) has been investigated in the ionic liquids (IL) [EMIM][Tf<inf>2</inf>N] (1-ethyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide) and [EMIM][B(CN)<inf>4</inf>] (1-ethyl-3-methylimidazolium tetracyanoborate) by using time-resolved UV/vis stopped-flow, electrochemistry, cryospray mass spectrometry, EPR, and XPS measurements. Stable KO<inf>2</inf> solutions in [EMIM][Tf<inf>2</inf>N] can be prepared up to a 15 mM concentration and are characterized by a signal in EPR spectrum at g = 2.0039 and by the 1215 cm-1 stretching vibration in the resonance Raman spectrum. While the negatively charged iron porphyrin Fe(P8-) does not react with superoxide in IL, Fe(P4+) and Fe(P8+) do react in a two-step process (first a reduction of the Fe(III) to the Fe(II) form, followed by the binding of superoxide to Fe(II)). In the reaction with KO<inf>2</inf>, Fe(P4+) and Fe(P8+) show similar rate constants (e.g., in the case of Fe(P4+): k<inf>1</inf> = 18.6 ± 0.5 M-1 s-1 for the first reaction step, and k<inf>2</inf> = 2.8 ± 0.1 M-1 s-1 for the second reaction step). Notably, these rate constants are four to five orders of magnitude lower in [EMIM][Tf<inf>2</inf>N] than in conventional solvents such as DMSO. The influence of the ionic liquid is also apparent during electrochemical experiments, where the redox potentials for the corresponding Fe(III)/Fe(II) couples are much more negative in [EMIM][Tf<inf>2</inf>N] than in DMSO. This modified redox and kinetic behavior of the positively charged iron porphyrins results from their interactions with the anions of the ionic liquid, while the nucleophilicity of the superoxide is reduced by its interactions with the cations of the ionic liquid. A negligible vapor pressure of [EMIM][B(CN)<inf>4</inf>] and a sufficient enrichment of Fe(P8+) in a close proximity to the surface enabled XPS measurements as a case study for monitoring direct changes in the electronic structure of the metal centers during redox processes in solution and at liquid/solid interfaces. (Figure Presented). © 2015 American Chemical Society.

Metal nanoclusters, sometimes called metamolecules or plasmonic oligomers, exhibit interesting optical properties such as Fano resonances and optical chirality. These properties promise a variety of practical applications, particularly in ultrasensitive biochemical sensing. Here we investigate experimentally the sensitivities of plasmonic pentamers and quadrumers to the adsorption of self-assembled nanometer-thick alkanethiol monolayers. The monolayer sensitivity of such oligomers is found to be significantly higher than that of single plasmonic nanoparticles and depends on the nanocluster arrangement, constituent nanoparticle shape, and the plasmon resonance wavelength. Together with full-wave numerical simulation results and the electromagnetic perturbation theory, we unveil a direct correlation between the sensitivity and the near-field intensity enhancement and spatial localization in the plasmonic hot spots generated in each nanocluster. Our observation is beyond conventional considerations (such as optimizing nanoparticle geometry or narrowing resonance line width) for improving the sensing performance of metal nanoclusters-based biosensors and opens the possibilities of using plasmonic nanoclusters for single-molecule detection and identification. © 2014 American Chemical Society.

Silica-coated surface-enhanced Raman scattering (SERS) labels with a self-assembled monolayer (SAM) on the entire surface of the metal colloid combine high chemical and mechanical stability with bright and reproducible Raman signals due to the complete surface coverage and uniform molecular orientation within the SAM. Currently available chemical syntheses are either based on the direct encapsulation of covalently bound silane precursors or comprise several steps, such as the sequential addition of noncovalently bound polyelectrolytes to render the surface vitreophilic. Here, a generic approach for the direct and fast silica encapsulation of commercially available Raman reporter molecules with polar head groups by noncovalently bound silane precursors is reported. The formation of highly SERS-active silica-coated clusters during silica encapsulation is determined by several parameters, in particular the type of Raman reporter molecule, the solvent, and the type and amount of the silane precursor. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Quantitative and accurate detection of multiple biomarkers would allow for the rapid diagnosis and treatment of diseases induced by pathogens. Monoclonal antibodies are standard affinity reagents applied for biomarkers detection; however, their production is expensive and labor-intensive. Herein, we report on newly developed nanoyeast single-chain variable fragments (NYscFv) as an attractive alternative to monoclonal antibodies, which offers the unique advantage of a cost-effective production, stability in solution, and target-specificity. By combination of surface-enhanced Raman scattering (SERS) microspectroscopy using glass-coated, highly purified SERS nanoparticle clusters as labels, with a microfluidic device comprising multiple channels, a robust platform for the sensitive duplex detection of pathogen antigens has been developed. Highly sensitive detection for individual Entamoeba histolytica antigen EHI115350 (limit of detection = 1 pg/mL, corresponding to 58.8 fM) and EHI182030 (10 pg/mL, corresponding 453 fM) with high specificity has been achieved, employing the newly developed corresponding NYscFv as probe in combination with SERS microspectroscopy at a single laser excitation wavelength. Our first report on SERS-based immunoassays using the novel NYscFv affinity reagent demonstrates the flexibility of NYscFv fragments as viable alternatives to monoclonal antibodies in a range of bioassay platforms and paves the way for further applications. © 2014 American Chemical Society.

Rapid parallel detection of two cytokines (IL-6 and IL-8) with femtogram sensitivity in a simple direct dot-blot assay is demonstrated. The microspectroscopic SERS acquisition scheme employs rationally designed, hydrophilically stabilized Au-Ag nanoshells as SERS labels, which are optimized for signal enhancement upon red laser excitation. © 2014 The Royal Society of Chemistry.

FT-Raman and FT-IR studies of the biomolecule 5-fluoroorotic acid in the solid state were carried out. The unit cell found in the crystal was simulated as a tetramer form by density functional calculations. They were performed to clarify wavenumber assignments of the experimental observed bands in the spectra. Correlations with the molecule of uracil were made, and specific scale equations were employed to scale the wavenumbers of 5-fluoroorotic acid. Good reproduction of the experimental wavenumbers is obtained and the % error is very small in the majority of the bands. This fact confirms our simplified solid state model. The molecular structure was fully optimized using DFT and MP2 methods. The relative stability of both the syn and anti conformations was investigated, and the anti-form was found to be slightly more stable, by 7.49 kJ/mol at the MP2 level. The structures of all possible tautomeric forms were determined. The keto-form appeared as the most stable one. The NBO atomic charges and several thermodynamic parameters were also calculated. © 2014 Elsevier B.V. All rights reserved.

Noble metal nanoparticles (NPs) are the most commonly employed plasmonic substrates in surface-enhanced Raman scattering (SERS) experiments. Computer simulations show that monomers of Ag and Au nanocrystals ("spherical" NPs) do not exhibit a notable plasmonic enhancement, i.e., they are essentially non-SERS-active. However, in experiments, SERS enhanced by spherical NP colloids has been frequently reported. This implies that the monomers do not have strong SERS activity, but detectable enhancement should more or less be there. Because of the gap between theory and practice, it is important to demonstrate experimentally how SERS-active the metal colloid actually is and, in case a SERS signal is observed, where it originates from. In particular the aggregation of the colloid, induced by high centrifugal forces in washing steps or due to a harsh ionic environment of the suspension medium, should be controlled since it is the very high SERS activity of NP clusters which dominates the overall SERS signal of the colloid. We report here the experimental evaluation of the SERS activity of 80 nm Au and Ag NP monomers. Instead of showing fancy nanostructures and super SERS enhancement, we present the method on how to obtain negative experimental data. In this approach, no SERS signal was obtained from the colloid with a Raman reporter on the metal surface when the NPs were encapsulated carefully within a thick silica shell. Without silica encapsulation, if a very low centrifugation speed is used for the washing steps, only a negligible SERS signal can be detected even at very high NP concentrations. In contrast, strong SERS signals can be detected when the NPs are suspended in acidic solutions. These results indicate that Au and Ag NP monomers essentially exhibit no SERS activity of practical relevance. © the Owner Societies 2015.

Rationally designed multifunctional plasmonic nanostructures efficiently integrate two or more functionalities into a single entity, for example, with both plasmonic and catalytic activity. This review article is focused on their synthesis and use in surface-enhanced Raman scattering (SERS) as a molecular spectroscopic technique with high sensitivity, fingerprint specificity, and surface selectivity. After a short tutorial on the fundamentals of Raman scattering and SERS in particular, applications ranging from chemistry (heterogeneous catalysis) to biology and medicine (diagnostics/imaging, therapy) are summarized. © 2014 IOP Publishing Ltd.

Surface-enhanced Raman scattering (SERS) has become a mature vibrational spectroscopic technique during the last decades and the number of applications in the chemical, material, and in particular life sciences is rapidly increasing. This Review explains the basic theory of SERS in a brief tutorial and - based on original results from recent research - summarizes fundamental aspects necessary for understanding SERS and provides examples for the preparation of plasmonic nanostructures for SERS. Chemical applications of SERS are the centerpiece of this Review. They cover a broad range of topics such as catalysis and spectroelectrochemistry, single-molecule detection, and (bio)analytical chemistry. Expanding vibrational spectroscopy: Since its first observation in 1973, surface-enhanced Raman scattering (SERS) has developed into a mature vibrational spectroscopic technique. The number of applications in chemistry as well as the material and life sciences is increasing rapidly. This Review summarizes the key concepts behind SERS and provides an overview of current applications in chemistry. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Abstract The site-specific pH is an experimental probe for assessing models of structural folding and function of a protein as well as protein-protein and protein-ligand interactions. It can be determined by various techniques such as NMR, FT-IR, fluorescence and EPR spectroscopy. The latter require the use of external labels, i.e., employ pH-dependent dyes and spin labels, respectively. In this contribution, we outline an approach to a label-free and site-specific method for determining the local pH using deep ultraviolet resonance Raman (UVRR) spectroscopic fingerprints of the aromatic amino acids histidine and tyrosine in combination with a robust algorithm that determines the pH value using three UVRR reference spectra and without prior knowledge of the pKa. © 2014 Elsevier B.V. All rights reserved.

SERS microscopy is a novel staining technique in immunohistochemistry, which is based on antibodies labeled with functionalized noble metal colloids called SERS labels or nanotags for optical detection. Conventional covalent bioconjugation of these SERS labels cannot prevent blocking of the antigen recognition sites of the antibody. We present a rational chemical design for SERS label-antibody conjugates which addresses this issue. Highly sensitive, silica-coated gold nanoparticle clusters as SERS labels are non-covalently conjugated to primary antibodies by using the chimeric protein A/G, which selectively recognizes the Fc part of antibodies and therefore prevents blocking of the antigen recognition sites. In proof-of-concept two-color imaging experiments for the co-localization of p63 and PSA on non-neoplastic prostate tissue FFPE specimens, we demonstrate the specificity and signal brightness of these rationally designed primary antibody-protein A/G-gold nanocluster conjugates. © 2014 The Royal Society of Chemistry.

Conformational search using force field methods on complex biomolecular systems is a key factor in understanding molecular and structural properties. The reliability of such investigations strongly depends on the efficiency of the conformational search algorithm as well as the accuracy of the employed force field. In the present work we compared the performance of two different approaches: the Monte-Carlo multiple minimum/low mode sampling (MCMM/LM), in combination with the OPLS2005 (MCMM/LM//OPLS2005), and Tabu-Search combined with Basin Hopping (TS/BH), employing the original OPLS-AA implementation proposed by Jorgensen (TS/BH//OPLS-AA). We investigated their performance in locating energetically low-lying structures and the efficiency in scanning the conformational phase space of non-covalently bonded complexes. As test systems we employed complexes of the artificial peptide receptor CBS-KKF with four different tetrapeptide ligands. The reliability and the accuracy of both approaches were examined by re-optimising all low-energy structures employing density functional theory with empirical dispersion correction in combination with triple zeta basis sets. Solvent effects were mimicked by a continuum solvent model. In all the four-test systems, the TS/BH//OPLS-AA approach yielded structures that are much lower in energy after the DFT optimisation. Additionally, it provided many low-lying structures that were not identified by the MCMM/LM//OPLS2005 approach. © 2013 Taylor and Francis Group, LLC.

Label-free in situ surface-enhanced Raman scattering (SERS) monitoring of reactions catalyzed by small gold nanoparticles using rationally designed plasmonic superstructures is presented. Catalytic and SERS activities are integrated into a single bifunctional 3D superstructure comprising small gold satellites self-assembled onto a large shell-isolated gold core, which eliminates photocatalytic side reactions. © 2012 American Chemical Society.

This perspective article provides an overview of selected medical applications of surface-enhanced Raman scattering (SERS), highlighting recent developments and trends. The use of SERS for detection, analysis and imaging has attracted great interest in the past decade owing to its high sensitivity and molecular fingerprint specificity. SERS can deliver chemical and structural information from analytes rapidly and nondestructively in a label-free manner. Alternatively, SERS labels or nanotags, when conjugated to target-specific ligands, can be employed for the selective detection and localization of the corresponding target molecule. Biomedical applications based on both approaches are highlighted. © 2013 the Owner Societies.

Rationally designed gold/silver nanoshells (Au/Ag-NS) with plasmon resonances optimized for red laser excitation in order to minimize autofluorescence from clinical samples exhibit scattering cross-sections, which are ca. one order of magnitude larger compared with solid quasi-spherical gold nanoparticles (Au-NPs) of the same size. Hydrophilic stabilization and sterical accessibility for subsequent bioconjugation of Au/Ag-NS is achieved by coating their surface with a self-assembled monolayer (SAM) of rationally designed Raman reporter molecules comprising terminal mono- and tri-ethylene glycol (EG) spacers, respectively. The stability of the hydrophilically stabilized metal colloid was tested under different conditions. In contrast to metal colloids coated with a SAM without terminal EG spacers, the hydrophilically stabilized SERS particles do not aggregate under physiologically relevant conditions, i.e., buffer solutions with high ionic strength. Using these rationally designed SERS particles in conjunction with a microspectroscopic acquisition scheme, a sandwich immunoassay for the sensitive detection of interleukin-6 (IL-6) was developed. Several control experiments demonstrate the high specificity of the assay towards IL-6, with a lowest detectable concentration of ca. 1 pg mL -1. The signal strength of the Au/Ag-NS is at least one order of magnitude higher compared with hydrophilically stabilized, non-aggregated solid quasi-spherical Au-NPs of the same size. © The Royal Society of Chemistry.

Immuno-SERS microscopy is a novel imaging technique in nano-biophotonics, which employs antibodies labeled with SERS-active nanoparticles in conjunction with Raman microscopy. Rapid data acquisition is of central importance for screening large areas of tissue specimens. Here, we first discuss the role of SERS labels with single-particle sensitivity in immuno-SERS microscopy, in particular with respect to false-negative results. In combined single-particle experiments (SERS microscopy/dark-field microscopy/HR-SEM), we then demonstrate that small glass-coated clusters (dimers and trimers) of gold nanospheres exhibit the desired single-particle SERS sensitivity, even at acquisition times as short as 30 msec per pixel, while monomers do not. The proof-of-concept for rapid immuno-SERS microscopy with 30 msec acquisition time per pixel for selective imaging of the p53 family member p63 in prostate tissue sections is demonstrated. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

SERS labels are a new class of nanotags for optical detection based on Raman scattering. Central advantages include their spectral multiplexing capacity due to the small line width of vibrational Raman bands, quantification based on spectral intensities, high photostability, minimization of autofluorescence from biological specimens via red to near-infrared (NIR) excitation, and the need for only a single laser excitation line. Current concepts for the rational design and synthesis of SERS labels are summarized in this review. Chemical constituents of SERS labels are the plasmonically active metal colloids for signal enhancement upon resonant laser excitation, organic Raman reporter molecules for adsorption onto the metal surface for identification, and an optional protective shell. Different chemical approaches towards the synthesis of rationally designed SERS labels are highlighted, including also their subsequent bioconjugation.

Dimers of metal nanospheres are well-known for their characteristic anisotropic optical response. Here, we demonstrate in single-particle SERS experiments that individual gold trimers and 3D superstructures exhibit a polarization-independent SERS response. This optical behavior of single particle clusters provides constant SERS signals, independent of the mutual orientation of the incident laser polarization and the plasmonic nanostructure, which is desired or even required in many SERS applications. © 2013 The Royal Society of Chemistry.

In order to gain insights into nucleotide-protein interaction, the molecular interaction of glycine (Gly) with 2′-deoxyguanosine 5′-monophosphate (dGMP) was monitored in aqueous media through Raman spectroscopic measurements and density functional theory (DFT) calculations. Raman spectra of dGMP, glycine and their binary mixtures (dGMP + Gly) in aqueous media at 10 different concentrations corresponding to the dGMP:Gly molar ratios varying from 1:1 to 1:10 were recorded in the fingerprint region 2000-400 cm-1. Raman spectra of the dGMP:Gly mixture with a molar ratio of 1:3 in aqueous media at 10 different pH values starting from 1 to 10 at an interval of 1 were also recorded. The DFT calculations were performed on dGMP, glycine and their various complexes with varying number of H2O molecules in gas phase as well as in solvation using hybrid functional B3LYP employing the high level basis set 6-311+G(d,p). The variations in the observed spectral features were explained in terms of calculated optimized structures of dGMP, glycine and their various complexes with water molecules and a good spectra-structure correlation was obtained. Raman spectra of glycine in 1.2 M aqueous solution were also recorded at three pH values, 2, 6 and 10, and the subtle differences in the spectral features were correlated with the calculated Raman spectra of protonated, deprotonated and zwitterionic forms of glycine. The results also give experimental evidence rather convincingly that the zwitterionic and protonated forms of glycine are formed at pH = 6 and 2, respectively, but even at pH = 10, deprotonated glycine is not formed. An important aspect of this study is the monitoring of the interaction of dGMP with the zwitterionic form of glycine giving insightful details regarding the binding site. © 2012 the Owner Societies.

A simple two-step seed-mediated synthesis of monodisperse quasi-spherical silver nanoparticles by citrate and ascorbic acid reduction is presented. Control over monodispersity is achieved by a variety of compounds with hydroxyl groups such as glycerol, ethylene glycol, agarose, or sucrose. The latter can also be used as a matrix for storage. © 2012 The Royal Society of Chemistry.

Optimal control of coherent anti-Stokes Raman scattering (CARS) image contrast is reported. The setup combines an evolutionary strategy and a closed-loop feedback with a liquid-crystal spatial modulator to control the spectrum of the Stokes pulse within a CARS scheme to optimize the vibrational contrast of CARS images. The CARS excitation spectrum is optimized for image contrast at a pre-determined wavenumber position. The optimization feedback uses an image-contrast parameter generated from the image itself as the experimentally imposed fitness parameter. This strategy allows for enhancing the image contrast by a factor of up to 2.6. © 2012 American Institute of Physics.

Vibrational spectroscopic investigations on molecular recognition processes are surprisingly rare, even at the qualitative level. In this first comparative study, we employ Fourier-transform infrared (FT-IR) and UV resonance Raman (UVRR) spectroscopy for quantitative label-free monitoring of molecular recognition processes. Specifically, the complexation of two different tetrapeptide ligands by an artificial receptor is investigated. The central advantage of UVRR is its capability to selectively probe the binding site of the receptor in the free/unbound and complexed form. In contrast, FT-IR probes the entire receptor-ligand complex without spectral selectivity, thereby providing complementary vibrational information. Multivariate analysis of the experimental IR/UVRR binding studies is required for determining association constants and the vibrational spectrum of the complex, which is not directly accessible. Both FT-IR and UVRR spectroscopy provide similar association constants for the two different tetrapeptide ligands. Complementary DFT calculations support the interpretation of the observed spectral changes upon complexation, which is a prerequisite for extracting structural information from vibrational binding studies. © 2012 The Royal Society of Chemistry.

The synthesis of 3D self-assembled plasmonic superstructures of gold nanospheres as well as the characterization of their structural and optical properties at the single-particle level is presented. This experimental work is complemented by FEM (finite element method) simulations of elastic scattering spectra and the spatial |E| 4 distribution for establishing structure-activity correlations in these complex 3D nanoclusters. © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Deep UVRR spectra of the aromatic amino acids Phe and Tyr in the wavenumber range 800-1800 cm-1 with λexc = 195-208 nm exhibit a selective enhancement of signals arising from vibrations localized in the aromatic ring. For λexc &gt; 198 nm, the UVRR spectra of Phe and Tyr are dominated by contributions from the in-plane ring stretching modes v8a and v8b at ∼ 1600 cm-1. For λexc ≤ 198 nm, intense signals from the symmetric ring stretching, in-plane C-H bending and phenyl-C stretching vibrations below 1400 cm-1 are observed. Excellent stray light rejection is achieved by a triple monochromator, which can be used either in the additive or subtractive mode for high-resolution and low-wavenumber measurements, respectively. A home-built circulating free-flow system allows the investigation of sample volumes as small as 1 mL. © by Oldenbourg Wissenschaftsverlag, München.

The design and synthesis of Raman reporter molecules comprising olefin or alkyne moieties with strong and characteristic vibrational Raman bands is presented. Chemisorption onto the surface of colloidal Au/Ag shells yields a self-assembled monolayer. Hydrophilic stabilization of such SERS labels can be achieved by short terminal ethylene glycol units attached to the Raman reporter. Encapsulation by silica with subsequent functionalization of the glass surface allows the conjugation to biomolecules such as antibodies. We demonstrate the use of SERS-labeled antibodies for tissue imaging of the tumor suppressor p63 in prostate biopsies. © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A multilayer quasi-continuous density gradient centrifugation method for separating 20-250 nm metal colloids with high size resolution while maintaining particle stability is presented. Colloidal mixtures containing monodisperse gold nanospheres and clusters thereof, in particular, gold dimers, are purified with yields up to 94%. The rapid method uses standard laboratory equipment. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

We report on the hydrogen bonding between pyrimidine (Pd) and methanol (M) as H-donor in this study. Hydrogen bonds between pyrimidine and methanol molecules as well as those between different methanol molecules significantly influence the spectral features at high dilution. The ring-breathing mode ν1 of the reference system Pd was chosen as a marker band to probe the degree of hydrogen bonding. Polarized Raman spectra in the region 970-1020 cm-1 for binary mixtures of (pyrimidine + methanol) at 28 different mole fractions were recorded. A Raman line shape analysis of the isotropic Raman line profiles at all concentrations revealed three distinct spectral components at mole fractions of Pd below 0.75. The three components are attributed to three distinct groups of species: 'free Pd' (pd), 'Pd with low methanol content' (pd1) and 'Pd with high-methanol content' (pd2). The two latter species differ considerably in the pattern and the strengths of the hydrogen bonds. The results of density functional theory calculations on structures and vibrational spectra of neat Pd and eight Pd/M complexes with varying methanol content support our interpretations of the experimental results. A nice spectra-structure correlation for the different cluster subgroups was obtained, similar to earlier results obtained for Pd and water. Apart from N⋯H and O⋯H hydrogen bonds between pyrimidine and methanol, O⋯H hydrogen bonds formed among the methanol molecules in the cluster at high methanol content also play a crucial role in the interpretation of the experimental results. Vibrational spectroscopic analysis on hydrogen bonding between pyrimidine (Pd) with methanol (M) comprises both experimental Raman spectra and density functional theory (DFT) calculations on structures and vibrational spectra of various Pd/M complexes with varying solvent content. Polarized Raman spectra in the region 970-1020 cm-1 for binary mixtures of (Pd + M) at 28 different mole fractions were recorded. The results of DFT calculations on structures and vibrational spectra of neat Pd and eight Pd/M complexes were used for interpreting the experimental results. Finally, a spectra-structure correlation for different cluster subgroups was obtained. Copyright © 2010 John Wiley & Sons, Ltd.

A biocompatible, seed-mediated synthesis of monodisperse ∼60 nm gold nanostars, followed by hydrophilic stabilization with ethylene glycol-modified Raman reporter molecules, is presented. Their application as SERS labels for imaging of the tumor suppressor p63 in prostate biopsies by immuno-SERS microscopy is demonstrated. © 2011 The Royal Society of Chemistry.

Immunohistochemistry (IHC) is one of the most widely used staining techniques for diagnostic purposes. The selective localization of target proteins in tissue specimens by conventional IHC is achieved with dye- or enzyme-labeled antibodies in combination with light microscopy. In this contribution, we demonstrate the proof-of-principle for IHC based on surface-enhanced coherent Raman scattering for contrast generation. Specifically, antibody-labeled metallic nanoshells in conjunction with surface-enhanced coherent anti-Stokes Raman scattering (SECARS) microscopy are employed for the selective, sensitive, and rapid localization of the basal cell protein p63 in normal prostate tissue. Negative control experiments were performed in order to confirm the selective binding of the target-specific metal nanoprobes and to disentangle the role of plasmonic (metal) and molecular (Raman reporter) resonances in this plasmon-assisted four-wave mixing technique. © 2011 American Chemical Society.

Self-assembly of gold nanospheres with a very thin glass shell onto the surface of beads yields a plasmonically active micron-sized substrate for integrated solid-phase synthesis and label-free SERS analysis. The proof-of-principle of this approach is demonstrated by the vibrational spectroscopic discrimination of three distinct amino acids and a dipeptide. © 2011 The Royal Society of Chemistry.

The introduction of carbon-deuterium (C-D) bonds into drug compounds by organic synthesis is a non-invasive labelling approach, which does not alter the chemical and physiological properties of the drug itself. C-deuterated drugs exhibit characteristic vibrational signatures in the C-D stretching region around 2100-2300 cm -1, which avoids spectral interference with contributions from a complex biological environment. In this paper, the quantitative detection of C-deuterated drugs by Raman microspectroscopy and single-band CARS microscopy is examined. Concentration-dependent studies on drugs with aliphatic and aromatic C-D moieties were performed in a two-channel microfluidic chip, using the corresponding non-deuterated (C-H) isotopologues as an internal reference. © 2011 The Royal Society of Chemistry.

A site-specific and quantitative approach for label-free monitoring of molecular recognition is presented. Specifically, the binding site of an artificial receptor is probed selectively by UVRR spectroscopy. The ligand binding constant can be determined by non-negative matrix factorization. © 2011 The Royal Society of Chemistry.

The Raman spectra of neat propionaldehyde [CH3CH2CHO or propanal (Pr)] and its binary mixtures with hydrogen-donor solvents, water (W) and methanol (M), [CH3CH2CHO + H2O] and CH3CH2CHO + CH3OH] with different mole fractions of the reference system, Pr varying from 0.1 to 0.9 at a regular interval of 0.1, were recorded in the ν(Ci=O) stretching region, 1600-1800 cm-1. The isotropic parts of the Raman spectra were analyzed for both the cases. The wavenumber positions and line widths of the component bands were determined by a rigorous line-shape analysis, and the peaks corresponding to self-associated and hydrogen-bonded species were identified. Raman peak at ∼1721 cm-1 in neat Pr, which has been attributed to the self-associated species, downshifts slightly (∼1 cm-1) in going from mole fraction 0.9 to 0.6 in (Pr + W) binary mixture, but on further dilution it shows a sudden downshift of ∼7 cm-1. This has been attributed to the low solubility of Pr in W (∼30%), which does not permit a hydrogen-bonded network to form at higher concentrations of Pr. A significant decrease in the intensity of this peak in the Raman spectra of Pr in a nonpolar solvent, n-heptane, at high dilution (C = 0.05) further confirms that this peak corresponds to the self-associated species. In case of the (Pr + M) binary mixture, however, the spectral changes with concentration show a rather regular trend and no special features were observed. The Raman spectra of neat propionaldehyde (propanal) and its binary mixtures with the hydrogen-donor solvents water and methanol with different mole fractions of propanal were recorded in the ν(Ci= O) stretching region 1600-1800 cm-1. Raman peaks corresponding to self-associated and hydrogen-bonded species were identified and the peak at ∼1721 cm-1 was attributed to the self-associated species, which was confirmed further by measuring the Raman spectra of propanal in a nonpolar solvent, n-heptane, at high dilution (C = 0.05), whereupon the intensity of this band diminished considerably. Copyright © 2010 John Wiley & Sons, Ltd.

Supercontinuum fiber light sources with their extremely wide wavelength spectrum can provide new options for achieving specific wavelength distributions in a very flexible way. Two concepts for the combination of such supercontinuum light sources with a spectrally dispersive optical system and an interactive filter for modulating the light spectrum are discussed. These concepts provide the possibility to achieve laser-like light sources with an almost arbitrary shape of the spectrum, great flexibility in interactive tuning of the spectral properties and covering wavelengths from UV to the infrared range. © 2010 Elsevier B.V. All rights reserved.

The synthesis of bifunctional Au/Pt/Au nanoraspberries for use in quantitative in situ monitoring of platinum-catalyzed reactions by surface-enhanced Raman scattering (SERS) is presented. Highly convolved SERS spectra of reaction mixtures can be decomposed into the contributions of distinct molecular species by multivariate data analysis. © 2011 American Chemical Society.

A simplified setup for coherent anti-Stokes Raman scattering (CARS) microscopy is introduced, which allows for recording CARS images with 30 cm-1 excitation band-width for probing Raman bands between 500 and 900 cm-1 with minimal requirements for alignment. The experimental arrangement is based on electronic switching between CARS images recorded at different Raman resonances by combining a photonic crystal fiber (PCF) as broadband light source and an acousto-optical programmable dispersive filter (AOPDF) as tunable wavelength filter. Such spatial light modulator enables selection of a narrow-band spectrum to yield high vibrational contrast and hence chemical contrast in the resultant CARS images. Furthermore, an experimental approach to reconstruct spectral information from CARS image contrast is introduced. © 2011 by Astro Ltd., published exclusively by WILEY-VCH Verlag GmbH & Co. KGaA.

Density functional calculations (DFT) by various methods were performed to clarify wavenumber assignments of the experimental observed bands. A comparison with the molecule of uracil was made, and specific scale factors were deduced and employed in the predicted wavenumbers of 5-IU. Comparisons were also performed with other halo-uracil derivatives. The scaled wavenumbers were compared with IR and Raman experimental data. Good reproduction of the experimental wavenumbers is obtained and the % error is very small in the majority of cases. The equilibrium geometry of 5-IU was also calculated at several levels, as well as the atomic charges and several thermodynamic parameters. All the tautomer forms of 5-iodouracil were determined and optimized. Several general conclusions were underlined. © 2009.

High-resolution Raman spectra of pyrimidine (PD) and formamide (FA)mixtures with different compositions recorded in the ring breathing region of PD(ν1 ∼ 991 cm-1) are presented. The dilution of PDwith FA leads to the appearance of a new band at ν1' ∼ 994cm-1, which is assigned to hydrogen-bonded PD:FA species. From aquantitative analysis of the concentration-dependent Raman spectra, the averagenumber of FA molecules in the first solvation sphere of PD is determined asbeing equal to 2. This value is supported by density functional theory (DFT)calculations: a symmetric 1:2 complex is the most stable species among varioushydrogen-bonded PD:FA clusters with stoichiometries ranging from 1:1 to 1:4. Aqualitative explanation for the blue shift of the ν1 mode uponcomplexation is given. Additionally, we have observed not only similarities butalso some differences with respect to the PD:water system. Copyright © 2010John Wiley & Sons, Ltd.

Nanoparticle probes for use in targeted detection schemes and readout by surface-enhanced Raman scattering (SERS) comprise a metal core, Raman reporter molecules and a protective shell. One design of SERS labels specifically optimized for biomedical applications in conjunction with red laser excitation is based on tunable gold/silver nanoshells, which are completely covered by a self-assembled monolayer (SAM) of Raman reporters. A shell around the SAMcoated metal core stabilizes the colloid and prevents particle aggregation. The optical properties and SERS efficiencies of these plasmonic nanostructures are characterized both experimentally and theoretically. Subsequent bioconjugation of SERS probes to ligands such as antibodies is a prerequisite for the selective detection of the corresponding target molecule via the characteristic Raman signature of the label. Biomedical imaging applications of SERS-labeled antibodies for tumor diagnostics by SERS microscopy are presented, using the localization of the tumor suppressor p63 in prostate tissue sections as an example. © 2010 SPIE.

Site-specific pKa determination of the carboxylate-binding subunit in artificial peptide receptors is achieved by pH-dependent UV resonance Raman scattering with subsequent matrix factorization. This quantitative methodology allows to determine accurately the local acidity of a binding site based on the complete vibrational spectrum and, most importantly, without the need of selecting any particular peak. © The Royal Society of Chemistry.

Covering everything from the basic theoretical and practical knowledge to new exciting developments in the field with a focus on analytical and life science applications, this monograph shows how to apply surface-enhanced Raman scattering (SERS) for solving real world problems. © 2011 Wiley-VCH Verlag GmbH & Co. KGaA.

A short synthesis route to silica-encapsulated nanoparticles coated with a self-assembled monolayer (SAM) is presented. The organic molecules within the SAM contain a SiO2 precursor to render the surface vitreophilic. Due to the high mechanical and chemical stability of a glass shell, such particles can be used as probes in targeted research with surface-enhanced Raman scattering as the read-out method. © 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

In this letter we present an approach to CARS microscopy, which compromises between fast acquisition rates and the amount of chemical information obtained. By using a light modulator as tunable filter in concert with narrowband pump and broadband Stokes pulses, we demonstrate an experimental arrangement, which allows for fast electronic switching between CARS images recorded at different Raman resonances without the need for any optical adjustment. © 2010 by Astro Ltd.

We discuss the combination of a CARS-imaging system with microfluidics. Such system is a versatile tool to quantify the relative contributions of resonant and non-resonant scattering at the CARS frequency. We will show that the twochannel microfluidic chip employed in combination with deuterated isotopomers as an internal standard allows for fast and quantitative detection of organic molecules by CARS microscopy. The experimental design enables the simultaneous measurement of both the chemically relevant Raman-resonant signal and the non-Raman-resonant background. © 2010 SPIE.