bluuebunny/crossref_metadata_2025_split huggingface dataset:
Recent investigations of small‐scale plasticity mechanisms in 3D and small‐sized systems using advanced in‐situ
TEM
nanomechanical testing
Authors: Idrissi Hosni 
Published: December 2016

Recently, the development of a new generation of advanced instruments for in‐situ TEM nanomechanical testing has allowed establishing a one‐to‐one relationship between load‐displacement characteristics and stress‐induced microstructure evolution in the transmission electron microscope (TEM). In the present work, it will be demonstrated that a step forward in the investigation of structural defects and small‐scale plasticity mechanisms can be made by combining commercial [1] and in‐house developed lab‐on‐chip [2] nanomechanical testing techniques with advanced TEM techniques.
High resolution aberration corrected TEM and STEM as well as automated crystallographic orientation and phase mapping in TEM (ACOM‐TEM) have been used to reveal the nanoscale plasticity mechanisms controlling the mechanical responses of nanocrystalline (nc) metallic Al and Pd freestanding thin films. Special attention was paid to the strain rate sensitivity of nanostructured metallic materials involving thermally activated plasticity mechanisms [3] (Figure 1). Furthermore, the microstructure of nc Pd thin films subjected to hydriding cycles has been investigated in order to unravel the interaction mechanisms of hydrogen with dislocations and interfaces [4] as well as the influence of hydrogen cycling on the mechanical properties of the Pd films. An original method combining the measurement of dislocation mobility using in‐situ TEM nanomechanical testing and dislocation dynamic (DD) simulations has also been used to investigate the plasticity of olivine small‐sized crystals at low temperature. It demonstrates for the first time the possibility of characterizing the mechanical properties of specimens, which could be available in the form of micron‐sized samples only [5].
More recently, quantitative nanobeam electron diffraction (NBED) was used to investigate the relationship between the local atomic order and the activation of shear transformation zones (STZs) in nanostructured ZrNi metallic glasses freestanding thin films. These films exhibit outstanding mechanical properties involving very large homogenous plastic deformation and giant ductility without the observation of mature shear bands until fracture. The basic principle of NBED is shown in Figure 2, consisting of a coherent electron beam with diameter of around 0.4 nm in order to produce two‐dimensional diffraction patterns from atomic clusters with comparable size. Furthermore, high resolution HAADF‐STEM and EELS revealed a heterogeneous microstructure with Ni‐rich and Zr‐rich regions exhibiting different atomic densities with characteristic length of 2‐3 nm. The role of such behaviour in the absence of shear bands and the delay of fracture in the ZrNi thin films is discussed.
Quantitative Nanoplasmonics in the
TEM
Authors: Bosman Michel 
Published: December 2016

Quantitative Nanoplasmonics in the TEM

Michel Bosman
1

1
Institute of Materials Research and Engineering, A*STAR (Agency for Science, Technology and Research), 2 Fusionopolis Way, #08‐03 Innovis, Singapore 138634
.
michel.bosman@gmail.com

Scanning TEM (STEM)‐based surface plasmon characterization will be discussed in this paper, and it will be argued that some plasmon properties can be measured quantitatively in the STEM, by carefully analyzing the local spectral response. Examples will be given that demonstrate the unique capability of STEM‐based plasmon analysis in comparison with other experimental techniques.
Some very impressive experimental results have recently been published in which plasmons on subwavelength metal nanostructures were mapped with 1‐100 fs time resolution. These include time‐domain techniques based on photoelectron emission microscopy (PEEM) [1, 2], time‐resolved scanning near‐field optical microscopy (SNOM) [3], ultrafast TEM [4] and even plasmon mapping with free electron lasers [5].
STEM‐based electron energy‐loss spectroscopy (EELS) is performed in the energy/frequency domain, and it has a long history as an experimental technique for surface plasmon characterization [6‐17]. However, since surface plasmon resonances are damped harmonic oscillators, it is possible to interpret the EELS plasmon spectra as Fourier‐transforms of oscillations in the time‐domain [18].
This paper will explore the implications of this approach, and will apply it to quantify local dynamic materials properties [19].
Correlative immuno Light Electron Microscopy (
CLEM
) of subcellular compartments
Authors: Klumperman Judith 
Published: December 2016

Correlative light and electron microscopy (CLEM) methods integrate light and electron microscopy on a single sample, literally bridging the gap between these two microscopy techniques. Most methods use fluorescent microscopy of thin or semi‐thin sections to define a region‐of‐interest, which then is traced back in the EM to provide subcellular context information (e.g. membrane organization, non‐labeled surroundings of the fluorescent structure). The most powerful application of CLEM is correlative live cell imaging‐EM, by which dynamic information is inferred to structures seen in static EM pictures. By merging the strengths of the two techniques a novel and integrated type of image is created that combines parameters that cannot ‐ or not easily ‐ be obtained when using separate images of related events. However, because imaging requirements are intrinsically different between light and electron microscopy, creating conditions that are ideal for both modalities is a challenging process. Moreover, correlating fluorescent labeling to the cellular architecture seen in the EM is not always straightforward. The most pressing challenges in CLEM are currently therefore 1. development of sample preparation methods that are appropriate for both LM and EM imaging. 2. development of bi‐modal probes that are visible in both LM and EM and 3. development of software that allows for rapid and accurate correlation of LM and EM images.
The focus of our research is to develop new probes and CLEM pipelines that allow us to efficiently and with high accuracy define the three‐dimensional (3D) ultrastructural context of fluorescently‐tagged proteins previously localized in fixed or living cells. We apply our technologies to study the cellular pathways and mechanisms that control the cell's digestive system – i.e. the endo‐lysosomal system ‐ in health and disease conditions. The main CLEM technology that we use in the lab is based on the use of immunogold labeling of ultrathin cryosections (the Tokuyasu technique) [1]. Most CLEM approaches, however, are restricted in their EM approach by the lack of 3D structural information. To overcome this limitation, we apply Focused Ion Beam Scanning Electron Microscopy (FIB‐SEM) as 3D‐EM approach in a live cell‐CLEM set up. To visualize endo‐lysosomes in live cells we combine fluorescent tagged endo‐lysosomal proteins (such as LAMP1‐mGFP) with endocytic tracers (such as fluorescently labeled dextran). This approach enables live‐cell tracking of specific endo‐lysosomal compartments, after which the samples are fixed, stained and resin‐embedded for FIB‐SEM imaging. Figure 1 presents an example of Dextran‐Alexa646 and/or LAMP1‐mGFP labeled endo‐lysosomal compartments in live cells (A) and in 3D‐EM (D), providing the cellular context at ultrastructural resolution. In my presentation I will show various examples of both immunoEM and FIB.SEM‐based CLEM approaches, which are designed to optimally image individual, membrane‐bounded compartments.
In situ
TEM
for understanding electrical and thermal transport properties on nano and atomic scales
Authors: Olsson Eva 
Published: December 2016

In situ electron microscopy allows studies of transport of charges and matter in complex structures as well as thermal properties. We can study mechanically and thermally induced changes of charge transport properties using holders designed to enable different stimuli allowing the direct observation and correlation between material structure and properties. The direct correlation between structure and properties on the small scale involving individual interfaces, defects and atoms provides access to new information about which microstructural constituents that are active in determining the material properties on the macro, micro, nano and atomic scale. This talk addresses examples of in situ electrical, mechanical and thermal studies. A few examples are briefly described below.
The nanoscale dimensions of semiconducting nanowires (NWs) provide extended strain relaxation capability between lattice‐mismatched materials and enable the fabrication of single‐NW p‐i‐n junction solar cells on low‐cost substrates. Due to its sub‐wavelength dimension, semiconductor NWs can function as optical antennas and exhibit a “self‐concentrating” effect that enhances optical absorption. The strain relaxation capability and the enhanced absorption cross section make NWs potential candidates as highly efficient and low‐cost solar cells. Due to the effects of elastic strain applied on the electronic band structure the strain can be used to achieve NW‐based photovoltaic devices with new functionality. We have studied the effect of mechanical strain on the electrical resistance of nanowires. Electron energy loss spectroscopy was used to study the effect of strain on the electronic structure with emphasis on the low energy loss interval of 0 to 50 eV. Electron beam induced current measurements were also performed to study the effect of strain on the diffusion length of the charge carriers [1].
Heating of a transmission electron microscopy (TEM) specimen can be performed in several parallel modes and this talk will address three types of heating modes and show experimental results from nanostructured materials. One mode is by resistive heating of a ring shaped support in contact with the circumference of the entire TEM sample. An additional mode is by use of a heating wire patterned on the TEM sample where the wire is contacted by leads fed through the TEM sample holder. The third mode is by active Joule heating of the nanostructure of study, such as carbon nanotubes, graphene, or metal nanowires. The purpose of having several parallel modes of heating is to enable the separation of temperature dependence, effects of self‐Joule heating, effects of radiative heating and thermal transport. It is also important to be able to extract the three dimensional information about the geometry of the investigated structures [2].
Scanning Probe Microscopy goes Live: seeing dynamic phenomena with
STM
Authors: Frenken Joost 
Published: December 2016

Scanning Tunneling Microscopy (STM) and other forms of Scanning Probe Microscopy (SPM), are traditionally applied mainly to static structures that are investigated mainly under relatively artificial conditions, such as ultrahigh vacuum (UHV). This is surprising in the light of the relative insensitivity of the operation mechanism of STM and that of most other types of SPM to such aspects as temperature or gas pressure or even the presence of liquids.
In this talk, I will demonstrate that it is possible to apply SPM techniques without compromising their atomic resolution even under harsh conditions [1‐4]. Extra attention is required to construct the SPM instrumentation such that it avoids the complications that are introduced by these conditions, such as excessive thermal drift or damage to delicate components. This is, in principle, a straightforward engineering task, which leads typically to dedicated designs for specific classes of imaging conditions [1‐4].

The examples provided in this talk are all
live
STM observations of relevant dynamic surface phenomena. They range from model catalysts under the conditions of high temperatures and high pressures at which they are being used in the chemical industry, to the chemical vapor deposition of graphene on metal substrates and the atom‐by‐atom deposition or erosion of surfaces under the influence of atom and ion beams.
Metal Contaminated Environments And Interactions With Biota
Authors: Isaure Marie‐Pierre 
Published: December 2016

Contamination of the environment with metals has dramatically increased during the last century due to intense anthropogenic activities and increasing metal need. Living organisms present in soils and aquatic systems are affected by these metals, and some of them are able to develop some specific strategies to cope with metal toxicity. To decipher the impact of metals on these organisms, and particularly their trafficking and storage, the determination of their localization at the tissue and cell level as well as their chemical forms (speciation) are crucial.
In this context, we combine various imaging techniques including synchrotron micro X‐ray Fluorescence (µXRF), Transmission Electron Microscopy combined with Energy Dispersive X‐ray analysis (TEM‐EDX), and Nano‐ Secondary Ion Mass Spectrometry (nano‐SIMS) to locate metals, and X‐ray Absorption Spectroscopy with focused and non focused beam (µXAS and XAS) to clarify the nature of metal species.

This approach will be illustrated by two environmental studies. The first one is focused on the green micro‐algae
Chlamydomonas reinhardtii
, an eukaryotic photosynthetic model for metal stress.
Chlamydomonas reinhardtii
is tolerant to cadmium but the mechanisms involved in this tolerance are misknown. We will show that intracellular thiol ligands are involved in Cd binding at the intracellular level, and that these sulfur ligands are not the only tolerance mechanism implemented by the cell. The second environmental study is related to mining soils contaminated with Cd and Zn. Among the existing soil treatments, phytoremediation has emerged as an alternative sustainable technique for two decades. We will show here how pioneer plants used in a phytostabilization process interact with metals in the soil.

This presentation illustrates the interest of combining imaging and spectroscopy techniques to get a global view of metals in complex environmental systems.
mScarlet, a novel high quantum yield (71%) monomeric red fluorescent protein with enhanced properties for
FRET
‐ and super resolution microscopy
Authors: Haarbosch Lindsay , Bindels Daphne , Postma Marten , Hink Mark , Royant Antoine , Gadella Theodorus 
Published: December 2016

mScarlet, a novel red fluorescent protein was generated from a synthetic template based on a consensus amino acid sequence derived from naturally occurring red fluorescent proteins and purple chromoproteins and on consensus monomerization mutations. The encoded synthetic red fluorescent protein was optimized by molecular evolution through site directed and random mutagenesis. Improved variants were selected by quantitative multimode screening for increased fluorescence lifetime, increased photo stability, increased quantum yield and for increased chromophore maturation.
Very bright variants were obtained with high fluorescence lifetimes up to 3.8 ns, quantum yields >70 % and complete maturation. The monomeric status of the variants was confirmed by OSER analysis and with a‐tubulin fusions. The brightness of mScarlet is >2‐ fold increased as compared to bright red fluorescent proteins such as mCherry, mRuby2 and tagRFP‐T as was analyzed with quantitative (single plasmid with viral 2A sequence) coexpression with mTurquoise2 in mammalian cells.
mScarlet can be used as a bright red fluorescent fusion tag for staining various subcellular structures in live cells. Because of their efficient maturation and high quantum yield, mScarlet vastly outperforms existing monomeric red fluorescent proteins in ratiometric FRET‐microscopy applications due to seriously enhanced sensitized emission and lack of photochromism (figure 1).
During evolution mScarlet variants with substantially altered spectroscopic properties were generated including fluorescence lifetime variants, photo labile variants and strongly spectrally shifted variants. Besides looking at generally optimized properties such as increased lifetime, maturation and brightness, some of these variants were rescreened for blinking properties. An mScarlet variant (7Q2BMs‐K) was identified that produced high spontaneous blinking upon illumination at 561 nm. This variant was fused to life‐act and co‐expressed with mVenus‐Lifeact. With 488 and 561 nm excitation, dual life cell single molecule localization microscopy produced perfectly colocalized yellow and red actin structures in living cells (figure 2), demonstrating the usefulness of the novel red fluorescent proteins for life cell super‐resolution microscopy.
Eavesdropping on the molecular signatures of embryonic development
Authors: Fraser Scott 
Published: December 2016

The study of embryonic development has been dramatically advanced by the wealth of high‐throughput molecular studies that have defined the genes and proteins involved. This wealth of data now presents the challenge of integrating a working knowledge of how these molecular components, often present at vanishingly small concentrations, generate reliable patterns of cell migration and cell differentiation. In typical cell biology approaches, cultures of isolated cells have been used reveal mechanism. What is needed to understand development is to carry out studies on cells in their normal context interacting with other cells and signals in the intact embryo. Key events of embryonic development take place over dimensions of less than 500um in less than 5 hours, making it tractable for light imaging tools to be used to answer this challenge.
Imaging techniques are challenged by major tradeoffs between spatial resolution, temporal resolution, and the limited photon budget. We are attempting to advance this tradeoff by constructing light sheet microscopes that maintain subcellular resolution in thick and scattering specimens. Our two‐photon light‐sheet microscope, combines the deep penetration of two‐photon microscopy and the speed of light sheet microscopy to generate images with more than ten‐fold improved imaging speed and sensitivity. As with other light sheet technologies, the collection of an entire 2‐D optical section in parallel offers dramatically speeds acquisition rates. By adopting two‐photon SPIM is far less subject to light scattering, permitting subcellular resolution to be maintained far better than conventional light sheet microscopes. The combination of attributes permits cell and molecular imaging with sufficient speed and resolution to generate unambiguous tracing of cells and signals in intact systems, which presents a major challenge in data management, processing and analysis.
Multispectral imaging offers the chance of asking multiple questions of the same embodied cells. Multiplex analyses permit the variance and the “noise” in a system to be exploited by asking about the analytes that co‐vary with a selected gene product. Transforming fluorescent spectra to a point on a 2D‐plot with sinusoidal functions (phasors) provides a powerful tool for the cumbersome tasks of visualizing and analyzing hyper‐spectral imaging data. This technique offers excellent performance in the face of biological and instrumental uncertainties. Our results on live zebrafish embryos document the accuracy of hyper‐spectral phasors and demonstrates their capability to distinguish multiple spectrally overlapping fluorophores in low signal‐to‐noise and fast analysis time.
Combined, these tools offer the multi‐dimensional imaging required to follow key events in embryos as they take place, and allow us to use variance as an experimental tool rather than a limitation.
Possibility to Predict Extrusion Die Incidental Fracture by Finite Element Simulation
Authors: Bevacqua Alessandro , Medvedev Alexander E. , Molotnikov Andrey , Axe Richard , Lapovok Rimma 
Published: December 2016

Finite element (FE) simulation of the extrusion process can minimize the time and effort in die design and assist with the optimization of technological parameters for obtaining high quality profiles. However, there are cases when extrusion die fails unexpectedly despite the simulations predicting a positive outcome.In this paper the incidental failure of the die used by Capral Ltd for extrusion of the T‐shape profile from AA6082 aluminium alloy is analysed. FE simulation complemented with a damage modelling is shown to be valuable tool for establishing the reasons for the die failure and for developing the strategy to prevent such an event.
Mössbauer spectroscopy evidence of intrinsic non‐stoichiometry in iron telluride single crystals
Authors: Kiiamov Airat G. , Lysogorskiy Yury V. , Vagizov Farit G. , Tagirov Lenar R. , Tayurskii Dmitrii A. , Croitori Dorina , Tsurkan Vladimir , Loidl Alois 
Published: December 2016

The FeTe parent compound for iron‐superconductor chalcogenides was studied applying Mössbauer spectroscopy accompanied by ab initio calculations of electric field gradients at the iron nuclei. Room‐temperature (RT) Mössbauer spectra of single crystals have shown asymmetric doublet structure commonly ascribed to contributions of over‐stoichiometric iron or impurity phases. Low‐temperature Mössbauer spectra of the magnetically ordered compound could be well described by four hyperfine‐split sextets, although no other foreign phases different from Fe1.05Te were detected by XRD and microanalysis within the sensitivity limits of the equipment. Density functional ab initio calculations have shown that over‐stoichiometric iron atoms significantly affect electron charge and spin density up to the second coordination sphere of the iron sub‐lattice, and, as a result, four non‐equivalent groups of iron atoms are formed by their local environment. The resulting four‐group model consistently describes the angular dependence of the single crystals Mössbauer spectra as well as intensity asymmetry of the doublet absorption lines in powdered samples at RT. We suppose that our approach could be extended to the entire class of FeSeTex compounds, which contain excess iron atoms.
image
Polymerase‐Mediated Site‐Specific Incorporation of a Synthetic Fluorescent Isomorphic G Surrogate into RNA
Authors: Li Yao , Fin Andrea , McCoy Lisa , Tor Yitzhak 
Published: December 2016

AbstractAn enzyme‐mediated approach for the assembly of singly modified RNA constructs in which specific G residues are replaced with thG, an emissive isomorphic G surrogate, is reported. Transcription in the presence of thG and native nucleoside triphosphates enforces initiation with the unnatural analogue, yielding 5′‐end modified transcripts that can be mono‐phosphorylated and ligated to provide longer site‐specifically modified RNA constructs. The scope of this unprecedented enzymatic approach to non‐canonical purine‐containing RNAs is explored via the assembly of several altered hammerhead (HH) ribozymes and a singly modified HH substrate. By strategically modifying key positions, a mechanistic insight into the ribozyme‐mediated cleavage is gained. Additionally, the emissive features of the modified nucleoside and its responsiveness to environmental changes can be used to monitor cleavage in real time by steady state fluorescence spectroscopy.