In Situ Observation of Chemically Induced Protein Denaturation at Solvated Interfaces.

Proteins unfold in chaotropic salt solutions, a process that is difficult to observe at the single protein level. The work presented here demonstrates that a liquid-based atomic force microscope and graphene liquid-cell-based scanning transmission electron microscope make it possible to observe chemically induced protein unfolding. To illustrate this capability, ferritin proteins were deposited on a graphene surface, and the concentration-dependent urea- or guanidinium-induced changes of morphology were monitored for holo-ferritin with its ferrihydrite core as well as apo-ferritin without this core. Depending on the chaotropic agent the liquid-based imaging setup captured an unexpected transformation of natively folded holo-ferritin proteins into rings after urea treatment but not after guanidinium treatment. Urea treatment of apo-ferritin did not result in nanorings, confirming that nanorings are a specific signature of denaturation of holo-ferritins after exposture to sufficiently high urea concentrations. Mapping the in situ images with molecular dynamics simulations of ferritin subunits in urea solutions suggests that electrostatic destabilization triggers denaturation of ferritin as urea makes direct contact with the protein and also disrupts the water H-bonding network in the ferritin solvation shell. Our findings deepen the understanding of protein denaturation studied using label-free techniques operating at the solid-liquid interface.

Operando Electrochemical Liquid Cell Scanning Transmission Electron Microscopy Investigation of the Growth and Evolution of the Mosaic Solid Electrolyte Interphase for Lithium-Ion Batteries.

The solid electrolyte interphase (SEI) is a key component of a lithium-ion battery forming during the first few dischage/charge cycles at the interface between the anode and the electrolyte. The SEI passivates the anode-electrolyte interface by inhibiting further electrolyte decomposition, extending the battery's cycle life. Insights into SEI growth and evolution in terms of structure and composition remain difficult to access. To unravel the formation of the SEI layer during the first cycles, operando electrochemical liquid cell scanning transmission electron microscopy (ec-LC-STEM) is employed to monitor in real time the nanoscale processes that occur at the anode-electrolyte interface in their native electrolyte environment. The results show that the formation of the SEI layer is not a one-step process but comprises multiple steps. The growth of the SEI is initiated at low potential during the first charge by decomposition of the electrolyte leading to the nucleation of inorganic nanoparticles. Thereafter, the growth continues during subsequent cycles by forming an island-like layer. Eventually, a dense layer is formed with a mosaic structure composed of larger inorganic patches embedded in a matrix of organic compounds. While the mosaic model for the structure of the SEI is generally accepted, our observations document in detail how the complex structure of the SEI is built up during discharge/charge cycling.

Li7La3Zr2O12 Protonation as a Means to Generate Porous/Dense/Porous-Structured Electrolytes for All-Solid-State Lithium-Metal Batteries.

Ceramic Li7La3Zr2O12 (LLZO) represents a promising candidate electrolyte for next-generation all-solid-state lithium-metal batteries. However, lithium-metal batteries are prone to dendrite formation upon fast charging. Porous/dense and porous/dense/porous LLZO structures were proposed as a solution to avoid or at least delay the formation of lithium-metal dendrites by increasing the electrode/electrolyte contact area and thus lowering the local current density at the interface. In this work, we show the feasibility of producing porous/dense/porous LLZO by a new and scalable method. The method consists of LLZO chemical deep protonation in a protic or acidic solvent, followed by thermal deprotonation at high temperatures to create the porous structure by water and lithium oxide elimination. We demonstrate that the produced structure extends the lifetime of Li/LLZO/Li symmetric cells by a factor of 8 compared to a flat LLZO at a current density of 0.1 mA/cm2 and with a capacity of 1 mAh/cm2 per half-cycle. We also show clear improvement of the Li/LLZO/LiFePO4 full cell performance with a thermally deprotonated LLZO.

Direct Imaging of the Atomic Mechanisms Governing the Growth and Shape of Bimetallic Pt-Pd Nanocrystals by In Situ Liquid Cell STEM.

Understanding the atomic mechanisms governing the growth of bimetallic nanoalloys is of great interest for scientists. As a promising material for photocatalysis applications, Pt-Pd bimetallic nanoparticles (NPs) have been in the spotlight for many years due to their catalytic performance, which is typically superior to that of pure Pt NPs. In this work, we use in situ liquid cell scanning transmission electron microscopy to track the exact atomic mechanisms governing the formation of bimetallic Pt-Pd NPs. We find that the formation process of the bimetallic Pt-Pd is divided into three stages. First, the nucleation and growth of ultrasmall primary nanoclusters are formed by the agglomeration of Pt and Pd atoms. Second, the primary nanoclusters are involved in a coalescence process to form two types of bigger agglomerates, namely, amorphous (a-NC) and crystalline (c-NC) nanoclusters. In the third stage, these clusters undergo a coalescence process leading to the formation of Pt-Pd NPs, while, in parallel, monomer attachment continues. We found that the third stage contains three types of coalescence processes, a-NC-a-NC, a-NC-c-NC, and c-NC-c-NC coalescence, which eventually give rise to crystalline bimetallic alloys. However, each type of coalescence gave distinct NPs in terms of shape and defects. Our results thus reveal the exact growth mechanisms of bimetallic alloys on the atomic scale, unravel the origin of their structure, and overall are of key interest to tailor the structure of bimetallic NPs.

Multi-step atomic mechanism of platinum nanocrystals nucleation and growth revealed by in-situ liquid cell STEM.

The understanding of crystal growth mechanisms has broadened substantially. One significant advancement is based in the conception that the interaction between particles plays an important role in the growth of nanomaterials. This is in contrast to the classical model, which neglects this process. Direct imaging of such processes at atomic-level in liquid-phase is essential for establishing new theoretical models that encompass the full complexity of realistic scenarios and eventually allow for tailoring nanoparticle growth. Here, we investigate at atomic-scale the exact growth mechanisms of platinum nanocrystals from single atom to final crystals by in-situ liquid phase scanning transmission electron microscopy. We show that, after nucleation, the nanocrystals grow via two main stages: atomic attachment in the first stage, where the particles initially grow by attachment of the atoms until depletion of the surrounding zone. Thereafter, follows the second stage of growth, which is based on particle attachment by different atomic pathways to finally form mature nanoparticles. The atomic mechanisms underlying these growth pathways are distinctly different and have different driving forces and kinetics as evidenced by our experimental observations.

Atomic Mechanisms of Nanocrystallization via Cluster-Clouds in Solution Studied by Liquid-Phase Scanning Transmission Electron Microscopy.

The formation of nanocrystals is at the heart of various scientific disciplines, but the atomic mechanisms underlying the early stages of crystallization from supersaturated solutions are still rather unclear. Here, we used in situ liquid-phase scanning transmission electron microscopy to study at the atomic level the very early stages of gold nanocrystal growth, and the evolution of its crystallinity. We found that the nucleation is initiated by the formation of poorly crystalline nanoparticles. These are transformed into monocrystals via nanocrystallization governed by a complex process of multiple out-and-in exchanges of matter between a crystalline-core and a disordered-shell, referred to as the cluster-cloud. Our observations at the crystal/cluster-cloud interface during growth demonstrate that the initially formed nanocrystals expel the poorly crystallized phases as nanoclusters into the cluster-cloud, then readsorb it by two distinct pathways, namely, by (i) monomer attachments and (ii) nanocluster coalescence. This growth process eventually leads to the formation of monocrystalline nanoparticles.

In Situ Electron Diffraction Tomography Using a Liquid-Electrochemical Transmission Electron Microscopy Cell for Crystal Structure Determination of Cathode Materials for Li-Ion batteries.

We demonstrate that changes in the unit cell structure of lithium battery cathode materials during electrochemical cycling in liquid electrolyte can be determined for particles of just a few hundred nanometers in size using in situ transmission electron microscopy (TEM). The atomic coordinates, site occupancies (including lithium occupancy), and cell parameters of the materials can all be reliably quantified. This was achieved using electron diffraction tomography (EDT) in a sealed electrochemical cell with conventional liquid electrolyte (LP30) and LiFePO4 crystals, which have a well-documented charged structure to use as reference. In situ EDT in a liquid environment cell provides a viable alternative to in situ X-ray and neutron diffraction experiments due to the more local character of TEM, allowing for single crystal diffraction data to be obtained from multiphased powder samples and from submicrometer- to nanometer-sized particles. EDT is the first in situ TEM technique to provide information at the unit cell level in the liquid environment of a commercial TEM electrochemical cell. Its application to a wide range of electrochemical experiments in liquid environment cells and diverse types of crystalline materials can be envisaged.

Operando Monitoring of the Solution-Mediated Discharge and Charge Processes in a Na-O2 Battery Using Liquid-Electrochemical Transmission Electron Microscopy.

Although in sodium-oxygen (Na-O2) batteries show promise as high-energy storage systems, this technology is still the subject of intense fundamental research, owing to the complex reaction by which it operates. To understand the formation mechanism of the discharge product, sodium superoxide (NaO2), advanced experimental tools must be developed. Here we present for the first time the use of a Na-O2 microbattery using a liquid aprotic electrolyte coupled with fast imaging transmission electron microscopy to visualize, in real time, the mechanism of NaO2 nucleation/growth. We observe that the formation of NaO2 cubes during reduction occurs by a solution-mediated nucleation process. Furthermore, we unambiguously demonstrate that the subsequent oxidation of NaO2 of which little is known also proceeds via a solution mechanism. We also provide insight into the cell electrochemistry via the visualization of an outer shell of parasitic reaction product, formed through chemical reaction at the interface between the growing NaO2 cubes and the electrolyte, and suggest that this process is responsible for the poor cyclability of Na-O2 batteries. The assessment of the discharge-charge mechanistic in Na-O2 batteries through operando electrochemical transmission electron microscopy visualization should facilitate the development of this battery technology.

Reversible magnesium and aluminium ions insertion in cation-deficient anatase TiO2.

In contrast to monovalent lithium or sodium ions, the reversible insertion of multivalent ions such as Mg2+ and Al3+ into electrode materials remains an elusive goal. Here, we demonstrate a new strategy to achieve reversible Mg2+ and Al3+ insertion in anatase TiO2, achieved through aliovalent doping, to introduce a large number of titanium vacancies that act as intercalation sites. We present a broad range of experimental and theoretical characterizations that show a preferential insertion of multivalent ions into titanium vacancies, allowing a much greater capacity to be obtained compared to pure TiO2. This result highlights the possibility to use the chemistry of defects to unlock the electrochemical activity of known materials, providing a new strategy for the chemical design of materials for practical multivalent batteries.

Intracellular degradation of functionalized carbon nanotube/iron oxide hybrids is modulated by iron via Nrf2 pathway.

The in vivo fate and biodegradability of carbon nanotubes is still a matter of debate despite tremendous applications. In this paper we describe a molecular pathway by which macrophages degrade functionalized multi-walled carbon nanotubes (CNTs) designed for biomedical applications and containing, or not, iron oxide nanoparticles in their inner cavity. Electron microscopy and Raman spectroscopy show that intracellularly-induced structural damages appear more rapidly for iron-free CNTs in comparison to iron-loaded ones, suggesting a role of iron in the degradation mechanism. By comparing the molecular responses of macrophages derived from THP1 monocytes to both types of CNTs, we highlight a molecular mechanism regulated by Nrf2/Bach1 signaling pathways to induce CNT degradation via NOX2 complex activation and O2•-, H2O2 and OH• production. CNT exposure activates an oxidative stress-dependent production of iron via Nrf2 nuclear translocation, Ferritin H and Heme oxygenase 1 translation. Conversely, Bach1 was translocated to the nucleus of cells exposed to iron-loaded CNTs to recycle embedded iron. Our results provide new information on the role of oxidative stress, iron metabolism and Nrf2-mediated host defence for regulating CNT fate in macrophages.

Carbon Nanotube Degradation in Macrophages: Live Nanoscale Monitoring and Understanding of Biological Pathway.

Despite numerous applications, the cellular-clearance mechanism of multiwalled carbon nanotubes (MWCNTs) has not been clearly established yet. Previous in vitro studies showed the ability of oxidative enzymes to induce nanotube degradation. Interestingly, these enzymes have the common capacity to produce reactive oxygen species (ROS). Here, we combined material and life science approaches for revealing an intracellular way taken by macrophages to degrade carbon nanotubes. We report the in situ monitoring of ROS-mediated MWCNT degradation by liquid-cell transmission electron microscopy. Two degradation mechanisms induced by hydroxyl radicals were extracted from these unseen dynamic nanoscale investigations: a non-site-specific thinning process of the walls and a site-specific transversal drilling process on pre-existing defects of nanotubes. Remarkably, similar ROS-induced structural injuries were observed on MWCNTs after aging into macrophages from 1 to 7 days. Beside unraveling oxidative transformations of MWCNT structure, we elucidated an important, albeit not exclusive, biological pathway for MWCNT degradation in macrophages, involving NOX2 complex activation, superoxide production, and hydroxyl radical attack, which highlights the critical role of oxidative stress in cellular processing of MWCNTs.

Unravelling kinetic and thermodynamic effects on the growth of gold nanoplates by liquid transmission electron microscopy.

The growth of colloidal nanoparticles is simultaneously driven by kinetic and thermodynamic effects that are difficult to distinguish. We have exploited in situ scanning transmission electron microscopy in liquid to study the growth of Au nanoplates by radiolysis and unravel the mechanisms influencing their formation and shape. The electron dose provides a straightforward control of the growth rate that allows quantifying the kinetic effects on the planar nanoparticles formation. Indeed, we demonstrate that the surface-reaction rate per unit area has the same dose-rate dependent behavior than the concentration of reducing agents in the liquid cell. Interestingly, we also determine a critical supply rate of gold monomers for nanoparticle faceting, corresponding to three layers per second, above which the formation of nanoplates is not possible because the growth is then dominated by kinetic effects. At lower electron dose, the growth is driven by thermodynamic and the formation and shape of nanoplates are directly related to the twin-planes formed during the growth.

Competing Forces in the Self-Assembly of Coupled ZnO Nanopyramids.

Self-assembly (SA) of nanostructures has recently gained increasing interest. A clear understanding of the process is not straightforward since SA of nanoparticles is a complex multiscale phenomenon including different driving forces. Here, we study the SA between aluminum doped ZnO nanopyramids into couples by combining inorganic chemistry and advanced electron microscopy techniques with atomistic simulations. Our results show that the SA of the coupled nanopyramids is controlled first by morphology, as coupling only occurs in the case of pyramids with well-developed facets of the basal planes. The combination of electron microscopy and atomistic modeling reveals that the coupling is further driven by strong ligand-ligand interaction between the bases of the pyramids as dominant force, while screening effects due to Al doping or solvent as well as core-core interaction are only minor contributions. Our combined approach provides a deeper understanding of the complex interplay between the interactions at work in the coupled SA of ZnO nanopyramids.

Growth of Pt-Pd Nanoparticles Studied In Situ by HRTEM in a Liquid Cell.

The growth of Pt-Pd nanoparticles from organometallic precursors is studied in situ in real time by HRTEM in a graphene oxide liquid cell. The reduction of the metal precursors is induced by the electron beam. During the growth, the particles rearrange their internal structure to form faceted single crystals. The growth is compatible with the Lifshitz-Slyozov-Wagner (LSW) mechanism in the limiting case of a reaction-limited process. The same particles are also synthesized ex situ by using a chemical reducing agent and observed in HRTEM.