On-chip light sheet illumination for nanoparticle tracking in microfluidic channels.

A simple and inexpensive method is presented to efficiently integrate light sheet illumination in a microfluidic chip for dark-field microscopic tracking and sizing of nanoparticles. The basic idea is to insert an optical fiber inside a polydimethylsiloxane (PDMS) elastomer microfluidic chip and use it as a cylindrical lens. The optical fiber is in this case no longer seen as only an optical waveguide but as a ready-made micro-optical component that is inexpensive and easy to source. Upon insertion, the optical fiber stretches the PDMS microchannel walls, which has two effects. The first effect is to tone down the intrinsic ripples in the PDMS that would otherwise create inhomogeneities in the light sheet illumination. The second effect is to remove any obliqueness of the channel wall and constrain it to be strictly perpendicular to the propagation of the illumination, avoiding the formation of a prismatic diopter. Through calculations, numerical simulations and measurements, we show that the optimal configuration consists in creating a slowly converging light sheet so that its axial thickness is almost uniform along the tracked area. The corresponding thickness was estimated at 12 µm, or 10 times the depth of field of the optical system. This leads to an at least six-fold increase in the signal-to-noise ratio compared to the case without the cylindrical lens. This original light-sheet configuration is used to track and size spherical gold nanoparticles with diameters of 80 nm and 50 nm.

Superpixels meet essential spectra for fast Raman hyperspectral microimaging.

In the context of spectral unmixing, essential information corresponds to the most linearly dissimilar rows and/or columns of a two-way data matrix which are indispensable to reproduce the full data matrix in a convex linear way. Essential information has recently been shown accessible on-the-fly via a decomposition of the measured spectra in the Fourier domain and has opened new perspectives for fast Raman hyperspectral microimaging. In addition, when some spatial prior is available about the sample, such as the existence of homogeneous objects in the image, further acceleration for the data acquisition procedure can be achieved by using superpixels. The expected gain in acquisition time is shown to be around three order of magnitude on simulated and real data with very limited distortions of the estimated spectrum of each object composing the images.

Schiff-base [4]helicene Zn(II) complexes as chiral emitters.

The controlled preparation of chiral emissive transition metal complexes is fundamental in the field of circularly polarized luminescence (CPL) active molecular materials. For this purpose, enantiopure Zn(ii) complexes 1 and 2 based on a tetradentate salen ligand surrounded by [4]helicene moieties, together with their racemic counterpart 3, have been herein synthesized. Chirality is primarily brought about by chiral 1,2-cyclohexane-diamines. Alternatively, achiral complex 4 based on ortho-phenylene-diamine has been prepared as well. Single crystal X-ray diffraction analyses have been performed on helicenic intermediates 8 and 9 and complexes 1 and 4. Complexes 1 and 4 display the typical tetradentate O,N,N,O coordination around Zn(ii) characteristic of salen ligands, and bear two [4]helicene moieties. The zinc complexes are luminescent in the visible range around 560 nm at room temperature in aerated solutions with the QY reaching ca. 15% for a luminescence lifetime of 5.5 ns. The optical activities of these complexes have been assessed by CD and CPL, and compared to DFT calculations.

Single-Particle Tracking with Scanning Non-Linear Microscopy.

This study describes the adaptation of non-linear microscopy for single-particle tracking (SPT), a method commonly used in biology with single-photon fluorescence. Imaging moving objects with non-linear microscopy raises difficulties due to the scanning process of the acquisitions. The interest of the study is based on the balance between all the experimental parameters (objective, resolution, frame rate) which need to be optimized to record long trajectories with the best accuracy and frame rate. To evaluate the performance of the setup for SPT, several basic estimation methods are used and adapted to the new detection process. The covariance-based estimator (CVE) seems to be the best way to evaluate the diffusion coefficient from trajectories using the specific factors of motion blur and localization error.

Photoluminescence spectra and quantum yields of gold nanosphere monomers and dimers in aqueous suspension.

The intrinsic one-photon excited photoluminescence (PL) of dimers and monomers of gold spheres suspended in water was studied by combining photon time-of-flight spectroscopy (PTOFS) and light scattering fluctuation correlation spectroscopy (LS-FCS). The samples are obtained by precisely controlling the dimerization of aqueous colloidal systems based on 50 and 80 nm gold nanospheres. The combination of PTOFS and LS-FCS enables the separate spectroscopic study of monomers and dimers even though they exist as a mixture in the samples. PL emission spectra and diffusional dynamics are obtained simultaneously through measurement at the single particle level. The PL spectra resemble the light scattering spectra, indicating the plasmon-assisted character of the photoluminescence process. We determine the intrinsic PL quantum yields of the dimers and the monomers. It is often not possible to measure such very low quantum yields in solution using conventional techniques, and we show here that PTOFS provides access to this information. The quantum yield of the dimers was found to be of same order of magnitude as that of the monomers, of the order of 10-6, which indicates that the interparticle 'electromagnetic hot-spots' do not play a major role in the luminescence emission mechanism in such plasmonic molecules.

Optical extinction and scattering cross sections of plasmonic nanoparticle dimers in aqueous suspension.

Absolute extinction and scattering cross sections for gold nanoparticle dimers were determined experimentally using a chemometric approach involving singular-value decomposition of the extinction and scattering spectra of slowly aggregating gold nanospheres in aqueous suspension. Quantitative spectroscopic data on plasmonic nanoparticle assemblies in liquid suspension are rare, in particular for particles larger than 40 nm, and in this work we demonstrate how such data can be obtained directly from the aggregating suspension. Our method can analyse, non invasively, the evolution of several sub-populations of nanoparticle assemblies. It may be applied to other self-assembling nanoparticle systems with an evolving optical response. The colloidal systems studied here are based on 20, 50 and 80 nm gold nanospheres in aqueous solutions containing sodium lipoate. In these systems, the reversible dimerisation process can be controlled using pH and ionic strength, and this control is rationalised in terms of DLVO theory. The dimers were identified in suspension by their translational and rotational diffusion through scattering correlation spectroscopy. Moreover, their gigadalton molecular weight was measured using electrospray charge-detection mass spectrometry, demonstrating that mass spectrometry can be used to study nanoparticles assemblies of very high molecular mass. The extinction and scattering cross sections calculated in the discrete-dipole approximation (DDA) agree very well with those obtained experimentally using our approach.

The intrinsic luminescence of individual plasmonic nanostructures in aqueous suspension by photon time-of-flight spectroscopy.

We have studied the intrinsic one-photon excited luminescence of freely diffusing gold nanoparticles of different shapes in aqueous suspension. Gold nanospheres were used as a reference, since their luminescence has been investigated previously and their light absorption and scattering properties are described analytically by Mie theory. We then studied gold nanobipyramids and nanostars that have recently gained interest as building blocks for new plasmonic nanosensors. The aim of our study is to determine whether the luminescence of gold nanoparticles of complex shape (bipyramids and nanostars) is a plasmon-assisted process, in line with the conclusions of recent spectroscopic studies on spheres and nanorods. Our study has been performed on particles in suspension in order to avoid any artefact from the heterogeneous environment created when particles are deposited on a substrate. We employ a recently developed photon time-of-flight method in combination with correlation spectroscopy of the light scattered by the particles to probe the luminescent properties of individual particles based on a particle-by-particle spectral analysis. Furthermore, we have performed resonant light scattering spectroscopic measurements on the same samples. Our work demonstrates the power of our time-of flight method for uncovering the plasmonic signatures of individual bipyramids and nanostars during their brief passage in the focal volume of a confocal set-up. These spectral features of individual particles remain hidden in macroscopic measurements. We find that the intrinsic photoluminescence emission of gold bipyramids and gold nanostars is mediated by their localized surface plasmons.

One-photon excited luminescence of single gold particles diffusing in solution under pulsed illumination.

Here we report on the visible luminescence of single gold nanospherical particles diffusing in water excited by a pulsed-laser at 488 nm. The signal studied by fluorescence correlation spectroscopy does not display the expected characteristics. The main deviation is obtained for the diffusion time that depends on the laser irradiance. Additional advanced methods of measurements have been implemented to further characterize the emission. These methods have allowed us to demonstrate that the luminescence does not blink even in the picosecond domain and that it does not photobleach either. The comparison between the signals obtained under pulsed and continuous excitations at the same wavelength suggests that the increase in the particle temperature plays a role in the non-linear increase in the luminescence intensity with the excitation power. As in the case of two-photon excited luminescence, it implies that a model describing the luminescence process for a single particle diffusing in liquid must take into account the whole system composed of the gold particle, the capping ligands and the surrounding water.

Time-of-flight photon spectroscopy: a simple scheme to monitor simultaneously spectral and temporal fluctuations of emission on single nanoparticles.

Here we report on a novel scheme for spectral analysis that exploits the wavelength dependence of the time-of-flight of a photon in a dispersive medium. This versatile and cost-effective method, named time-of-flight photon spectroscopy (TOFPS), has the major advantage of being compatible with time-correlated single-photon counting experiments. Consequently, each photon acquired during an experiment is characterized by two parameters, its absolute time of arrival and its color, respectively. As a result, the spectral and temporal fluctuations of the emission of a single nano-object can be derived from a single measurement. As a proof of the concept, we demonstrate in the paper that the method can be used to perform Raman spectroscopy as well as fluorescence spectroscopy. We emphasize that TOFPS proves to be very efficient for improving signal-to-noise ratio in fluorescence correlation spectroscopy measurements by subsequent spectral filtering and to record luminescence spectra from single metallic particles. We demonstrate that the opportunity of simultaneously recording spectral and temporal fluctuations could be used to sort particles of different shapes inside a sample. TOFPS furthermore allows developing a new type of time interval distribution analysis which correlates the time interval between two photons and their corresponding color shift. It is applied to the analysis of the two-photon excited luminescence of a single gold nanorod. This method has a potential for a broad range of applications, among which time-resolved SERS spectroscopy and analysis of the dynamics of emission processes can be handled with new statistical approaches based on the correlation of spectral and temporal fluctuations.

Surface plasmon resonance properties of single elongated nano-objects: gold nanobipyramids and nanorods.

The spectral characteristics (wavelength and line width) and the optical extinction cross-section of the longitudinal localized surface plasmon resonance (LSPR) of individual gold nanobipyramids have been quantitatively measured using the spatial modulation spectroscopy technique. The morphology of the same individual nanoparticles has been determined by transmission electron microscopy (TEM). The experimental results are thus interpreted with a numerical model using the TEM measured sizes of the particles as an input, and either including the substrate or assuming a mean homogeneous environment. Results are compared to those obtained for individual nanorods and also show the importance of the local environment of the particle on the detailed description of its spectral position and extinction amplitude.

Influence of polarization and wavelength on two-photon excited luminescence of single gold nanospheres.

The Brownian rotation of a nearly spherical gold particle capped with ligands can be observed in the correlation profile of the intensity of the two-photon excited luminescence. Here we report on a multi-parameter study of the luminescence properties, including spectral and polarization analysis of the signal at the single particle level. First, the data confirm the role of the radiative de-excitation of the surface plasmons in the luminescence process. Secondly, the results obtained at low power indicate that the capped particle in water can be approximatively described as a spherical rotor acting in the far-field as a point-like absorption and emission dipole of fixed directions. In addition, we show that the dynamics of the ligands, induced by the heat transfer from the particle to its environment, can be partly controlled by the choice of excitation wavelength.

Fluorescence correlation spectroscopy reveals strong fluorescence quenching of FITC adducts on PEGylated gold nanoparticles in water and the presence of fluorescent aggregates of desorbed thiolate ligands.

Colloidal gold particles functionalised with oligoethylene-glycolated disulfide ligands and fluorescent moieties derived from fluorescein isothiocyanate (FITC) have been prepared and studied in aqueous suspension using fluorescence correlation spectroscopy (FCS). FCS probes the dynamics of the particles at the single object level, and reveals the desorption of fluorescent ligands which subsequently aggregate into larger (slower diffusing) objects. Cross-correlation spectroscopy of the FITC fluorescence and the Rayleigh-Mie scattering (RM-FCCS) of the gold cores shows that the only detectable fluorescent objects are free ligands and aggregates not associated with a gold particle. The fluorescence of bound fluorophores is quenched making their fluorescence too weak to be detected. FCS and RM-FCCS are useful tools for characterising functionalised noble metal particles in solution, under conditions similar to those used in optical bio-imaging. Desorption of thiolates from gold nanoparticles needs to be taken into account when working with these materials at low concentration.

Ligand-induced anisotropy of the two-photon luminescence of spherical gold particles in solution unraveled at the single particle level.

Here we report on the visible luminescence properties of individual spherical gold particles in solution, obtained by two-photon excited fluorescence correlation spectroscopy and by an original dual Rayleigh-fluorescence method, correlating the Rayleigh scattering and the luminescence fluctuations of the same particle. The results demonstrate that the power needed to observe the two-photon excited visible luminescence depends on the illuminated particle and that the corresponding emission is anisotropic at low power. These observations combined with the evolution of the dynamics of the luminescence with respect to excitation power are interpreted by the presence of unique emissive surface states that are randomly switched off and on by the heat-induced movement of the molecular coating. These characteristics, which remain hidden in macroscopic experiments, have important implications with respect to the potential use of the particles as labels in two-photon imaging in aqueous samples.

Probing the interactions between disulfide-based ligands and gold nanoparticles using a functionalised fluorescent perylene-monoimide dye.

The binding of disulfides to gold nanoparticles was investigated using fluorescence spectroscopy and a perylene-monoimide dye coupled to a dissymmetric disulfide via a tetraethyleneglycolalkyl chain (PMImSS). Quantum chemical calculations using the polarizable continuum model (PCM) predict a strong quenching of perylene-monoimide fluorescence by gold nanoparticles as a result of efficient excitation energy transfer from the dye to the particle. Such quenching is indeed observed when unfunctionalised gold nanoparticles are added to a solution of PMImSS. The fluorimetric titration curves show behaviour indicative of the existence of an equilibrium between free and bound ligands (association constant 5 x 10(5) M(-1)), whereas the affinity of thiols and disulfide for gold surfaces is in general assumed to be much higher. Gold nanoparticles fully functionalised with PMImSS were synthesised and purified. Fluorescence correlation spectroscopy shows the appearance of free PMImSS ligands in dilute (approx. pM) suspensions of these PMImSS-functionalised nanoparticles over a period of several days.