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International audience; Due to their unique size-dependent properties, nanoparticles (NPs) have many industrial and biomedical applications. Although NPs are generally characterized based on the size or morphological analysis, the mass of whole particles can be of interest as it represents the total amount of material in the particle regardless of shape, density, or elemental composition. In addition, the shape of nonspherical NPs presents a conceptual challenge, making them difficult to characterize in terms of size or morphological characteristics. Here, we used a novel nano-electro-mechanical sensor mass spectrometry (NEMS-MS) technology to characterize the mass distributions of various NPs. For standard spherical gold NPs, mass distributions covered the range from ∼5 to 250 MDa (8 to ∼415 attograms). Applying the density of gold (19.3 g/cm3) and assuming perfect sphericity, these mass measurements were used to compute the equivalent diameters of the NPs. The sizes determined agreed well with the transmission electron microscopy (TEM) imaging data, with deviations of ∼1.4%. Subsequently, we analyzed the mass distribution of ∼50 nm synthetic silicon dioxide particles, having determined their size by electron microscopy (SEM and TEM). Their estimated density was in line with the literature values derived from differential mobility analyzer and aerosol particle mass analyzer data. Finally, we examined the intact gold nanotetrapods and obtained a mass distribution revealing their controlled polydispersity. The presence of polyethylene glycol coating was also quantified and corroborated nuclear magnetic resonance observations. Our results demonstrate the potential of NEMS-MS-based measurements as an effective means to characterize NPs, whatever their composition, shape or density.

When studying viruses, the most prevalent aspects that come to mind are their structural and functional features, but this leaves in the shadows a quite universal characteristic: their mass. Even if approximations can be derived from size and density measurements, the multi MDa to GDa mass range, featuring a majority of viruses, has so far remained largely unexplored. Recently, nano-electromechanical resonator-based mass spectrometry (NEMS-MS) has demonstrated the ability to measure the mass of intact DNA filled viral capsids in excess of 100 MDa. However, multiple factors have to be taken in consideration when performing NEMS-MS measurements. In this article, phenomena influencing NEMS-MS mass estimates are listed and discussed, including some particle's extraneous physical properties (size, aspect ratio, stiffness), and the influence of frequency noise and device fabrication defects. These factors being accounted for, we could begin to notice subtler effects linked with (e.g.) particle desolvation as a function of operating parameters. Graphical abstract.

International audience; Due to their physical properties, nanomechanical sensors (NEMS) can achieve mass measurements in the mega- to gigadalton range, which is hardly obtained with conventional mass-spectrometers. However, NEMS signals are subject to noise, causing a loss of mass resolution and thus emphasizing the need of noise control. We propose a denoising model that relies on a total variation formulation, which deals with different noise models (particularly colored noise) affecting NEMS. The model also takes into account the physics of NEMS, such as the non-linear coupling between signals of individual NEMS. The performance of the proposed model is tested on simulated data which parameters are chosen similar to true experimental conditions. The obtained results confirm the interest of our model with a mass-resolution increase over 20% compared to methods used in literature.

When studying viruses, the most prevalent aspects that come to mind are their structural and functional features, but this leaves in the shadows a quite universal characteristic: their mass. Even if approximations can be derived from size and density measurements, the multi MDa to GDa mass range, featuring a majority of viruses, has so far remained largely unexplored. Recently, nano-electromechanical resonator–based mass spectrometry (NEMS-MS) has demonstrated the ability to measure the mass of intact DNA filled viral capsids in excess of 100 MDa. However, multiple factors have to be taken in consideration when performing NEMS-MS measurements. In this article, phenomena influencing NEMS-MS mass estimates are listed and discussed, including some particle’s extraneous physical properties (size, aspect ratio, stiffness), and the influence of frequency noise and device fabrication defects. These factors being accounted for, we could begin to notice subtler effects linked with (e.g.) particle desolvation as a function of operating parameters.

Micrometer scale colloidal particles experiencing ∼kT scale interactions and suspended in a fluid are relevant to a broad spectrum of applications. Often, colloidal particles are anisotropic, either by design or by nature. Yet, there are few techniques by which ∼kT scale interactions of anisotropic particles can be measured. Herein, we present the initial development of scattering morphology resolved total internal reflection microscopy (SMR-TIRM). The hypothesis of this work is that the morphology of light scattered by an anisotropic particle from an evanescent wave is a sensitive function of particle orientation. This hypothesis was tested with experiments and simulations mapping the scattered light from colloidal ellipsoids at systemically varied orientations. Scattering morphologies were first fitted with a two-dimensional (2D) Gaussian surface. The fitted morphology was parameterized by the morphology's orientation angle Mϕ and aspect ratio MAR. Data from both experiments and simulations show Mϕ to be a function of the particle azimuthal angle, while MAR was a sensitive function of the polar angle. This analysis shows that both azimuthal and polar angles of a colloidal ellipsoid could be resolved from scattering morphology as well or better than using bright-field microscopy. The integrated scattering intensity, which will be used for determining the separation distance, was also found to be a sensitive function of particle orientation. A procedure for interpreting these confounding effects was developed that in principle would uniquely determine the separation distance, the azimuthal angle, and the polar angle. Tracking these three quantities is necessary for calculating the potential energy landscape sampled by a colloidal ellipsoid.

Fluctuation correlation spectroscopy (FCS) is a well-established analytical technique traditionally used to monitor molecular diffusion in dilute solutions, the dynamics of chemical reactions, and molecular processes inside living cells. In this review, we present the recent use of FCS for measuring the size of colloidal nanoparticles in solution. We review the theoretical basis and experimental implementation of this technique and its advantages and limitations. In particular, we show examples of the use of FCS to measure the size of gold nanoparticles, monitor the rotational dynamics of gold nanorods, and investigate the formation of protein coronas on nanoparticles.

Bridging the mass gap Viruses and many large biomolecule complexes are in a mass range that is challenging to measure with conventional mass spectrometry methods. Nanomechanical resonators can determine masses of impacting molecules, but separation methods often lose too much of the sample to be efficient. Dominguez-Medina et al. used an aerodynamic lens that improved separation and focusing of nebulized molecules with increasing mass. The mass of both filled and empty viral capsids was determined with an array of 20 nanoresonators. Science , this issue p. 918

Circular dichroism spectroscopy is essential for structural characterization of proteins and chiral nanomaterials. Chiral structures from plasmonic materials have extraordinary strong circular dichroism effects compared to their molecular counterparts. While being extensively investigated, the comprehensive account of circular dichroism effects consistent with other plasmonic phenomena is still missing. Here we investigated the circular differential scattering of a simple chiral plasmonic system, a twisted side-by-side Au nanorod dimer, using single-particle circular dichroism spectroscopy complimented with electromagnetic simulations. This approach enabled us to quantify the effects of structural symmetry breaking, namely, size-mismatch between the constituent Au nanorods and large twist angles on the resulting circular differential scattering spectrum. Our results demonstrate that, if only scattering is considered as measured by dark-field spectroscopy, a homodimer of Au nanorods with similar sizes prod...

The response of living systems to nanoparticles is thought to depend on the protein corona, which forms shortly after exposure to physiological fluids and which is linked to a wide array of pathophysiologies. A mechanistic understanding of the dynamic interaction between proteins and nanoparticles and thus the biological fate of nanoparticles and associated proteins is, however, often missing mainly due to the inadequacies in current ensemble experimental approaches. Through the application of a variety of single molecule and single particle spectroscopic techniques in combination with ensemble level characterization tools, we identified different interaction pathways between gold nanorods and bovine serum albumin depending on the protein concentration. Overall, we found that local changes in protein concentration influence everything from cancer cell uptake to nanoparticle stability and even protein secondary structure. We envision that our findings and methods will lead to strategies to control the associated pathophysiology of nanoparticle exposure in vivo.

We report on the one-photon photoluminescence of gold nanorods with different aspect ratios. We measured photoluminescence and scattering spectra from 82 gold nanorods using single-particle spectroscopy. We found that the emission and scattering spectra closely resemble each other independent of the nanorod aspect ratio. We assign the photoluminescence to the radiative decay of the longitudinal surface plasmon generated after fast interconversion from excited electron-hole pairs that were initially created by 532 nm excitation. The emission intensity was converted to the quantum yield and was found to approximately exponentially decrease as the energy difference between the excitation and emission wavelength increased for gold nanorods with plasmon resonances between 600 and 800 nm. We compare this plasmon emission to its molecular analogue, fluorescence.

A strong intrinsic signal is advantageous over labeling for optical detection of nanoparticles. Intense scattering and absorption by the surface plasmon resonance, which exceeds molecular cross sections, provides a direct method for visualizing noble metal nanoparticles. While two-photon luminescence in gold nanoparticles yields a strong signal, one-photon luminescence is generally regarded to be much weaker and has seldom been employed for optical nanoparticle detection. In this article we investigated one-photon luminescence of gold nanospheres and nanorods using single particle spectroscopy with excitation at 514 and 633 nm. We characterized the polarization dependence, determined the quantum yield, and present a mechanism describing one-photon luminescence. Our results suggest fast interconversion between surface plasmons and hot electron–hole pairs and show that the luminescence occurs via emission by a surface plasmon. Using the information obtained from the single particle studies, we were able to ...

Photoluminescent Au nanoparticles are appealing for biosensing and bioimaging applications because of their non-photobleaching and non-photoblinking emission. The mechanism of one-photon photoluminescence from plasmonic nanostructures is still heavily debated though. Here, we report on the one-photon photoluminescence of strongly coupled 50 nm Au nanosphere dimers, the simplest plasmonic molecule. We observe emission from coupled plasmonic modes as revealed by single-particle photoluminescence spectra in comparison to correlated dark-field scattering spectroscopy. The photoluminescence quantum yield of the dimers is found to be surprisingly similar to the constituent monomers, suggesting that the increased local electric field of the dimer plays a minor role, in contradiction to several proposed mechanisms. Aided by electromagnetic simulations of scattering and absorption spectra, we conclude that our data are instead consistent with a multistep mechanism that involves the emission due to radiative decay of surface plasmons generated from excited electron-hole pairs following interband absorption.

Plasmonic polymers are quasi one-dimensional assemblies of nanoparticles whose optical responses are governed by near-field coupling of localized surface plasmons. Through single particle extinction spectroscopy correlated with electron microscopy, we reveal the effect of the composition of the repeat unit, the chain length, and extent of disorder on the energies, intensities, and line shapes of the collective resonances of individual plasmonic polymers constructed from three different sizes of gold nanoparticles. Our combined experimental and theoretical analysis focuses on the superradiant plasmon mode, which results from the most attractive interactions along the nanoparticle chain and yields the lowest energy resonance in the spectrum. This superradiant mode redshifts with increasing chain length until an infinite chain limit, where additional increases in chain length cause negligible change in the energy of the superradiant mode. We find that, among plasmonic polymers of equal width comprising nanoparticles with different sizes, the onset of the infinite chain limit and its associated energy are dictated by the number of repeat units and not the overall length of the polymer. The intensities and linewidths of the superradiant mode relative to higher energy resonances, however, differ as the size and number of nanoparticles are varied in the plasmonic polymers studied here. These findings provide general guidelines for engineering the energies, intensities, and line shapes of the collective optical response of plasmonic polymers constructed from nanoparticles with sizes ranging from a few tens to one hundred nanometers.

This tutorial review surveys the optical properties of plasmonic nanoparticles studied by various single particle spectroscopy techniques. The surface plasmon resonance of metallic nanoparticles depends sensitively on the nanoparticle geometry and its environment, with even relatively minor deviations causing significant changes in the optical spectrum. Because for chemically prepared nanoparticles a distribution of their size and shape is inherent, ensemble spectra of such samples are inhomogeneously broadened, hiding the properties of the individual nanoparticles. The ability to measure one nanoparticle at a time using single particle spectroscopy can overcome this limitation. This review provides an overview of different steady-state single particle spectroscopy techniques that provide detailed insight into the spectral characteristics of plasmonic nanoparticles.

The retention and elution of proteins in ion-exchange chromatography is routinely controlled by adjusting the mobile phase salt concentration. It has repeatedly been observed, as judged from adsorption isotherms, that the apparent heterogeneity of adsorption is lower at more-eluting, higher ionic strength. Here, we present an investigation into the mechanism of this phenomenon using a single-molecule, super-resolution imaging technique called motion-blur Points Accumulation for Imaging in Nanoscale Topography (mbPAINT). We observed that the number of functional adsorption sites was smaller at high ionic strength and that these sites had reduced desorption kinetic heterogeneity, and thus narrower predicted elution profiles, for the anion-exchange adsorption of α-lactalbumin on an agarose-supported, clustered-charge ligand stationary phase. Explanations for the narrowing of the functional population such as inter-protein interactions and protein or support structural changes were investigated through kinetic analysis, circular dichroism spectroscopy, and microscopy of agarose microbeads, respectively. The results suggest the reduction of heterogeneity is due to both electrostatic screening between the protein and ligand and tuning the steric availability within the agarose support. Overall, we have shown that single molecule spectroscopy can aid in understanding the influence of ionic strength on the population of functional adsorbent sites participating in the ion-exchange chromatographic separation of proteins.

We find that citrate-stabilized gold nanoparticles aggregate and precipitate in saline solutions below the NaCl concentration of many bodily fluids and blood plasma. Our experiments indicate that this is due to complexation of the citrate anions with Na+ cations in solution. A dramatically enhanced colloidal stability is achieved when bovine serum albumin is adsorbed to the gold nanoparticle surface, completely preventing nanoparticle aggregation under harsh environmental conditions where the NaCl concentration is well beyond the isotonic point. Furthermore, we explore the mechanism of the formation of this albumin ‘corona’ and find that monolayer protein adsorption is most likely ruled by hydrophobic interactions. As for many nanotechnology-based biomedical and environmental applications, particle aggregation and sedimentation are undesirable and could substantially increase the risk of toxicological side-effects, the formation of the BSA corona presented here provides a low-cost bio-compatible strategy for nanoparticle stabilization and transport in highly ionic environments.

Here we present in situ observations of adsorption of bovine serum albumin (BSA) on citrate-stabilized gold nanospheres. We implemented scattering correlation spectroscopy as a tool to quantify changes in the nanoparticle Brownian motion resulting from BSA adsorption onto the nanoparticle surface. Protein binding was observed as an increase in the nanoparticle hydrodynamic radius. Our results indicate the formation of a protein monolayer at similar albumin concentrations as those found in human blood. Additionally, by monitoring the frequency and intensity of individual scattering events caused by single gold nanoparticles passing the observation volume, we found that BSA did not induce colloidal aggregation, a relevant result from the toxicological viewpoint. Moreover, to elucidate the thermodynamics of the gold nanoparticle-BSA association, we measured an adsorption isotherm which was best described by an anti-cooperative binding model. The number of binding sites based on this model was consistent with a BSA monolayer in its native state. In contrast, experiments using poly-ethylene glycol capped gold nanoparticles revealed no evidence for adsorption of BSA.

A surface plasmon is the coherent oscillation of the conduction band electrons. When a metal nanoparticle is excited to produce surface plasmons, incident light is both scattered and absorbed, giving rise to brilliant colors. One available technique for measuring these processes, ensemble extinction spectroscopy, only measures the sum of scattering and absorption. Although the spectral responses of these processes are closely related, their relative efficiencies can differ significantly as a function of nanoparticle size and shape. For some applications, researchers may need techniques that can quantitatively measure absorption or scattering alone. Through advances in single particle spectroscopy, researchers can overcome this problem, separately determining the radiative (elastic and inelastic scattering) and nonradiative (absorption) properties of surface plasmons. Furthermore, because we can use the same sample preparation for both single particle spectroscopy measurements and electron microscopy, this technique provides detailed structural information and a direct correlation between optical properties and nanostructure morphology. In this Account, we present our quantitative investigations of both radiative (scattering and one-photon luminescence) and nonradiative (absorption) properties of the same individual plasmonic nanostructures employing different single particle spectroscopy techniques. In particular, we have used a combined setup to study the same structure with dark-field scattering spectroscopy, photothermal heterodyne imaging, confocal luminescence microscopy, and scanning electron microscopy. While Mie theory thoroughly describes the overall size dependence of scattering and absorption for nanospheres, our real samples deviate significantly from the predicted trend: their particle shape is not perfectly spherical, especially when supported on a substrate. Because of the high excitation rate in laser based single particle measurements, we can efficiently detect one-photon luminescence despite a low quantum yield. For gold nanoparticles, the luminescence spectrum follows the scattering response, and therefore we assigned it to the emission of a plasmon. Due to strong near-field interactions the plasmonic response of closely spaced nanoparticles deviates significantly from that of the constituent nanoparticles. This response arises from coupled surface plasmon modes that combine those of the individual nanoparticles. Our correlated structural and optical imaging strategy is especially powerful for understanding these collective modes and their dependence on the assembly geometry.