Tailoring the NIR-II Photoluminescence of Single Thiolated Au25 Nanoclusters by Selective Binding to Proteins.

Atomically precise gold nanoclusters are a fascinating class of nanomaterials that exhibit molecule-like properties and have outstanding photoluminescence (PL). Their ultrasmall size, molecular chemistry, and biocompatibility make them extremely appealing for selective biomolecule labeling in investigations of biological mechanisms at the cellular and anatomical levels. In this work, we report a simple route to incorporate a preformed Au25 nanocluster into a model bovine serum albumin (BSA) protein. A new approach combining small-angle X-ray scattering and molecular modeling provides a clear localization of a single Au25 within the protein to a cysteine residue on the gold nanocluster surface. Attaching Au25 to BSA strikingly modifies the PL properties with enhancement and a redshift in the second near-infrared (NIR-II) window. This study paves the way to conrol the design of selective sensitive probes in biomolecules through a ligand-based strategy to enable the optical detection of biomolecules in a cellular environment by live imaging.

Sub-100 nanometer silver doped gold-cysteine supramolecular assemblies with enhanced nonlinear optical properties.

The ability of gold(i) thiolates to self-assemble into supramolecular architectures opens the route for a new class of nanomaterials with a unique structure-optical property relationship. However, for a confirmed structure-optical property relationship, a control of the supramolecular architectures is required. In this work, we report a simple synthesis of sub-100 nanometer gold-cysteine and silver doped gold-cysteine supramolecular assemblies. We explore in particular silver-doping as a strategy to enhance the optical properties of these supramolecular assemblies. By an accurate characterization of as-synthesized supramolecular nanoparticles, we have been able to measure for the first time, their absolute two-photon absorption cross-section, two-photon excited fluorescence cross-section and first hyperpolarizabilities at different near-IR wavelengths. Huge values are obtained for silver doped gold-cysteine supramolecular assemblies, as compared to their corresponding undoped counterpart. In addition, we employ DFT and TD-DFT methods to study the geometric and electronic structures of model gold-cysteine and silver doped gold-cysteine compounds in order to address the structure-linear/nonlinear optical property relationship. The aim is to gain insights into the origin of the nonlinear optical enhancement of silver-doped gold supramolecular assemblies.

Ligand shell size effects on one- and two-photon excitation fluorescence of zwitterion functionalized gold nanoclusters.

Gold nanoclusters (Au NCs) are an emerging class of luminescent nanomaterials but still suffer from moderate photoluminescence quantum yields. Recent efforts have focused on tailoring their emission properties. Introducing zwitterionic ligands to cap the metallic kernel is an efficient approach to enhance their one-photon excitation fluorescence intensity. In this work, we extend this concept to the nonlinear optical regime, i.e., two-photon excitation fluorescence. For a proper comparison between theory and experiment, both ligand and solvent effects should be considered. The effects of ligand shell size and of aqueous solvent on the optical properties of zwitterion functionalized gold nanoclusters have been studied by performing quantum mechanics/molecular mechanics (QM/MM) simulations.

Ligand impact on reactive oxygen species generation of Au10 and Au25 nanoclusters upon one- and two-photon excitation.

In photodynamic therapy (PDT), light-sensitive photosensitizers produce reactive oxygen species (ROS) after irradiation in the presence of oxygen. Atomically-precise thiolate-protected gold nanoclusters are molecule-like nanostructures with discrete energy levels presenting long lifetimes, surface biofunctionality, and strong near-infrared excitation ideal for ROS generation in PDT. We directly compare thiolate-gold macromolecular complexes (Au10) and atomically-precise gold nanoclusters (Au25), and investigate the influence of ligands on their photoexcitation. With the ability of atomically-precise nanochemistry, we produce Au10SG10, Au10AcCys10, Au25SG18, and Au25AcCys18 (SG: glutathione; AcCys: N-acetyl-cysteine) fully characterized by high-resolution mass spectrometry. Our theoretical investigation reveals key factors (energetics of excited states and structural influence of surface ligands) and their relative importance in singlet oxygen formation upon one- and two-photon excitation. Finally, we explore ROS generation by gold nanoclusters in living cells with one- and two-photon excitation. Our study presents in-depth analyses of events within gold nanoclusters when photo-excited both in the linear and nonlinear optical regimes, and possible biological consequences in cells.

Four orders-of-magnitude enhancement in the two-photon excited photoluminescence of homoleptic gold thiolate nanoclusters following zinc ion-induced aggregation.

Herein we report unprecedented determination of the molar mass of zinc mediated assemblies of homoleptic gold nanoclusters, based on charge detection mass spectrometry measurements. The accurate determination of the molar mass of zinc coordinated assemblies of gold clusters has further allowed unambiguous determination of the two-photon excited photoluminescence cross section of the same. Furthermore, in line with one-photon excited photoluminescence measurements, four orders-of-magnitude enhancement in two-photon excited photoluminescence of gold nanoclusters has been observed following complexation with zinc ions. The study reported herein is envisioned to not only deepen the fundamental understanding of the multiphoton excitation properties of atomic clusters but also widen their application potential as luminescence markers.

Size and ligand effects of gold nanoclusters in alteration of organellar state and translocation of transcription factors in human primary astrocytes.

Ultra-small gold nanoclusters (AuNCs) with designed sizes and ligands are gaining popularity for biomedical purposes and ultimately for human imaging and therapeutic applications. Human non-tumor brain cells, astrocytes, are of particular interest because they are abundant and play a role in functional regulation of neurons under physiological and pathological conditions. Human primary astrocytes were treated with AuNCs of varying sizes (Au10, Au15, Au18, Au25) and ligand composition (glutathione, polyethylene glycol, N-acetyl cysteine). Concentration and time-dependent studies showed no significant cell loss with AuNC concentrations <10 µM. AuNC treatment caused marked differential astrocytic responses at the organellar and transcription factor level. The effects were exacerbated under severe oxidative stress induced by menadione. Size-dependent effects were most remarkable with the smallest and largest AuNCs (10, 15 Au atoms versus 25 Au atoms) and might be related to the accessibility of biological targets toward the AuNC core, as demonstrated by QM/MM simulations. In summary, these findings suggest that AuNCs are not inert in primary human astrocytes, and that their sizes play a critical role in modulation of organellar and redox-responsive transcription factor homeostasis.

Functionalized Au15 nanoclusters as luminescent probes for protein carbonylation detection.

Atomically precise, ligand-protected gold nanoclusters (AuNCs) attract considerable attention as contrast agents in the biosensing field. However, the control of their optical properties and functionalization of surface ligands remain challenging. Here we report a strategy to tailor AuNCs for the precise detection of protein carbonylation-a causal biomarker of ageing. We produce Au15SG13 (SG for glutathione) with atomic precision and functionalize it with a thiolated aminooxy moiety to impart protein carbonyl-binding properties. Mass spectrometry and molecular modelling reveal the key structural features of Au15SG12-Aminooxy and its reactivity towards carbonyls. Finally, we demonstrate that Au15SG12-Aminooxy detects protein carbonylation in gel-based 1D electrophoresis by one- and two-photon excited fluorescence. Importantly, to our knowledge, this is the first application of an AuNC that detects a post-translational modification as a nonlinear optical probe. The significance of post-translational modifications in life sciences may open avenues for the use of Au15SG13 and other nanoclusters as contrast agents with tailored surface functionalization and optical properties.

Gold nanoclusters elicit homeostatic perturbations in glioblastoma cells and adaptive changes of lysosomes.

Unique physicochemical features place gold nanoclusters at the forefront of nanotechnology for biological and biomedical applications. To date, information on the interactions of gold nanoclusters with biological macromolecules is limited and restricts their use in living cells. Methods: Our multidisciplinary study begins to fill the current knowledge gap by focusing on lysosomes and associated biological pathways in U251N human glioblastoma cells. We concentrated on lysosomes, because they are the intracellular destination for many nanoparticles, regulate cellular homeostasis and control cell survival. Results: Quantitative data presented here show that gold nanoclusters (with 15 and 25 gold atoms), surface-modified with glutathione or PEG, did not diminish cell viability at concentrations ≤1 µM. However, even at sublethal concentrations, gold nanoclusters modulated the abundance, positioning, pH and enzymatic activities of lysosomes. Gold nanoclusters also affected other aspects of cellular homeostasis. Specifically, they stimulated the transient nuclear accumulation of TFEB and Nrf2, transcription factors that promote lysosome biogenesis and stress responses. Moreover, gold nanoclusters also altered the formation of protein aggregates in the cytoplasm. The cellular responses elicited by gold nanoclusters were largely reversible within a 24-hour period. Conclusions: Taken together, this study explores the subcellular and molecular effects induced by gold nanoclusters and shows their effectiveness to regulate lysosome biology. Our results indicate that gold nanoclusters cause homeostatic perturbations without marked cell loss. Notably, cells adapt to the challenge inflicted by gold nanoclusters. These new insights provide a framework for the further development of gold nanocluster-based applications in biological sciences.

Second harmonic scattering from mass characterized 2D graphene oxide sheets.

In this communication, we report the second harmonic scattering from mass characterized 2D graphene oxide sheets dispersed in an aqueous suspension, in the femtosecond regime at 800 nm laser excitation. Charge-detection mass-spectrometry, performing at the single sheet level, allows for an exhaustive molar mass distribution and thus concentration for these 2D nanomaterials samples. The orientation-averaged hyperpolarizability value is (1.36 ± 0.15) × 10-25 esu as determined by the concentration-dependent harmonic scattering signal. In addition, the multi-photon excited fluorescence spectrum is characterized by a broad band in the visible range between 350 and 700 nm centered at about 500 nm.

Templating S100A9 amyloids on Aß fibrillar surfaces revealed by charge detection mass spectrometry, microscopy, kinetic and microfluidic analyses.

The mechanism of amyloid co-aggregation and its nucleation process are not fully understood in spite of extensive studies. Deciphering the interactions between proinflammatory S100A9 protein and Aß42 peptide in Alzheimer's disease is fundamental since inflammation plays a central role in the disease onset. Here we use innovative charge detection mass spectrometry (CDMS) together with biophysical techniques to provide mechanistic insight into the co-aggregation process and differentiate amyloid complexes at a single particle level. Combination of mass and charge distributions of amyloids together with reconstruction of the differences between them and detailed microscopy reveals that co-aggregation involves templating of S100A9 fibrils on the surface of Aß42 amyloids. Kinetic analysis further corroborates that the surfaces available for the Aß42 secondary nucleation are diminished due to the coating by S100A9 amyloids, while the binding of S100A9 to Aß42 fibrils is validated by a microfluidic assay. We demonstrate that synergy between CDMS, microscopy, kinetic and microfluidic analyses opens new directions in interdisciplinary research.