Energy-filtered Electron Microscopy Research Articles

Femtosecond-resolved imaging of a single-particle phase transition in energy-filtered ultrafast electron microscopy

Using an energy filter in transmission electron microscopy has enabled elemental mapping at the atomic scale and improved the precision of structural determination by gating inelastic and elastic imaging electrons, respectively. Here, we use an energy filter in ultrafast electron microscopy to enhance the temporal resolution toward the domain of atomic motion. Visualizing transient structures with femtosecond temporal precision was achieved by selecting imaging electrons in a narrow energy distribution from dense chirped photoelectron packets with broad longitudinal momentum distributions and thus typically exhibiting picosecond durations. In this study, the heterogeneous ultrafast phase transitions of vanadium dioxide (VO2) nanoparticles, a representative strongly correlated system, were filmed and attributed to the emergence of a transient, low-symmetry metallic phase caused by different local strains. Our approach enables electron microscopy to access the time scale of elementary nuclear motion to visualize the onset of the structural dynamics of matter at the nanoscale.

Design of a novel correlative reflection electron microscope for in-situ real-time chemical analysis* *Project supported by the Shanghai Tech University and the National Natural Science Foundation of China (Grant No. 11774039).

A novel instrument that integrates reflection high energy electron diffraction (RHEED), electron energy loss spectroscopy (EELS), and imaging is designed and simulated. Since it can correlate the structural, elemental, and spatial information of the same surface region via the simultaneously acquired patterns of RHEED, EELS, and energy-filtered electron microscopy, it is named correlative reflection electron microscopy (c-REM). Our simulation demonstrates that the spatial resolution of this c-REM is lower than 50 nm, which meets the requirements for in-situ monitoring the structural and chemical evolution of surface in advanced material.

Graphene-diamond junction photoemission microscopy and electronic interactions

Polycrystalline graphene was transferred onto differently terminated epitaxial layers of boron-doped diamond deposited onto single crystal substrates. Chemical and electronic characterisation was performed using energy-filtered photoemission electron microscopy and angle-resolved photoemission spectroscopy. Electronic interaction between the diamond and graphene was observed, where doping of the graphene on the hydrogen and oxygen terminated diamond was n-doping of 250 meV and 0 meV respectively. We found that the wide window of achievable graphene doping is effectively determined by the diamond surface dipole, easily tuneable with a varying surface functionalisation. A Schottky junction using the graphene-diamond structure was clearly observed and shown to reduce downward band bending of the hydrogen terminated diamond, producing a Schottky barrier height of 330 meV.

Application of energy‐filtering electron microscopy (EFEM) for analysis of hydrogen peroxide and lignin in epidermal walls of cucumber leaves triggered by acibenzolar‐S‐methyl treatment prior to inoculation with Colletotrichum orbiculare

Hydrogen peroxide (H2O2) and lignin induced by acibenzolar‐S‐methyl (ASM) during the expression of systemic acquired resistance (SAR) of cucumber plants against the fungus Colletotrichum orbiculare was analysed with electron microscopy and energy‐filtering electron microscopy (EFEM). ASM pretreatment consistently led to the blockage of appressorial penetration pegs into leaves at early stages after inoculation, whereas the fungus was able to grow into the vascular tissue after distilled water pretreatment. The blocked pegs, seen frequently at epidermal pectin layers in leaves of ASM‐treated plants, were surrounded by abundant electron‐dense amorphous material and dots. These were identified as lignin, based on their appearance and high reactivity with KMnO4. Lignin first appeared as an amorphous material at pectin layers, and then as dots in some areas of the material. The results showed that ASM caused SAR in leaves via faster formation of lignin within 1 day after inoculation. After CeCl3 treatment to detect H2O2, H2O2‐reactive products (cerium perhydroxides) were seen near ASM‐induced lignin formation sites in pectin layers. EFEM analysis showed that Ce and O were located at the same sites as the lignin dots, suggesting that lignin was associated with sources of H2O2 generation. The results indicate that the faster H2O2 generation and lignification induced by ASM were the potential causes of SAR.

Surface Proximity Effect, Imprint Memory of Ferroelectric Twins, and Tweed in the Paraelectric Phase of BaTiO3

We have used energy-filtered photoemission electron microscopy (PEEM) at the photoemission threshold to carry out a microscopic scale characterization of the surface charge and domain structure of the (001) surface in BaTiO3. Signatures of ferroelectric and ferroelastic domains, and tweed, dominate the surface structure of BaTiO3 at room temperature. The surface ferroic signatures are maintained on heating to temperature (~550 K), well above the transition temperature (393 K). This surface proximity effect provides the mechanism for memory of the bulk ferroelectric domain arrangement up to 150 K above TC and thus can be considered as a robust fingerprint of the ferroelectric state near the surface. Self-reversal of polarization is observed for the tweed below TC and for the surface domains above TC. Annealing at higher temperature triggers the dynamic tweed which in turn allows a full reorganization of the ferroic domain configuration.

Sub-nanometre resolution imaging of polymer-fullerene photovoltaic blends using energy-filtered scanning electron microscopy.

The resolution capability of the scanning electron microscope has increased immensely in recent years, and is now within the sub-nanometre range, at least for inorganic materials. An equivalent advance has not yet been achieved for imaging the morphologies of nanostructured organic materials, such as organic photovoltaic blends. Here we show that energy-selective secondary electron detection can be used to obtain high-contrast, material-specific images of an organic photovoltaic blend. We also find that we can differentiate mixed phases from pure material phases in our data. The lateral resolution demonstrated is twice that previously reported from secondary electron imaging. Our results suggest that our energy-filtered scanning electron microscopy approach will be able to make major inroads into the understanding of complex, nano-structured organic materials.

Contrast in atomically resolved EF-SCEM imaging

Energy-filtered scanning confocal electron microscopy (EF-SCEM) is a technique that uses the reduced depth of field of an aberration-corrected transmission electron microscope to provide three-dimensional (3D) compositional information. Using a silicon sample in the <110> orientation, we show that EF-SCEM image data can be recorded that shows lattice resolution in the plane perpendicular to the incident beam direction. The confocal effect is demonstrated through the reduction of the mean intensity as the confocal plane is displaced from the sample mid-plane, unlike optical sectioning in high-angle annular dark-field scanning transmission electron microscopy (STEM). Simulations of the EF-SCEM data show agreement with the experimental data, and allow the interpretability of the data to be explored. The effects of channelling, absorption and delocalisation complicate the quantitative and qualitative interpretation of the data, highlighting the need for matching to simulations. Finally the effects of the finite detector pin-hole aperture size are explored, and we show that the EF-SCEM contrast in the plane perpendicular to the beam direction starts to resemble that of a STEM spectrum imaging experiment as the aperture size increases.

A hydro/organo/hybrid gelator: A peptide lipid with turning aspartame head groups

This work presents a novel bola-type peptide lipid which can gelate water, organic solvents, and water/organic-solvent mixtures. In its molecular structure, an amphiphilic dipeptide aspartame (l-α-aspartyl-l-phenylalanine methyl ester) is connected at both ends of an alkylene linker. The different morphologies in the hydrogel (helical nanotapes) and the organogel (tape-like nanostructures) were visualized by energy-filtering transmission electron microscopy (EF-TEM) and energy-filtering scanning electron microscopy (FE-SEM), and the molecular arrangement was examined using X-ray diffraction (XRD), infrared (IR) spectroscopy, and circular dichroism (CD) spectroscopy. Possessing a hydrophilic aspartic acid group and a (relatively) hydrophobic phenylalanine methyl ester group, the dipeptide head group can turn about in response to solvent polarity. As a consequence, the solvent condition changed the molecular packing of the gelator and affected the overall supramolecular structure of the gel. It is noted that the peptide lipid gelated mixed solvents of water and organic solvents such as dichloromethane, liquid-paraffin, olive-oil, silicone-oils, and so on. The present hybrid gel can simultaneously hold hydrophilic and hydrophobic functional materials.

Atom Probe Tomography of Apatites and Bone-Type Mineralized Tissues

Nanocrystalline biological apatites constitute the mineral phase of vertebrate bone and teeth. Beyond their central importance to the mechanical function of our skeleton, their extraordinarily large surface acts as the most important ion exchanger for essential and toxic ions in our body. However, the nanoscale structural and chemical complexity of apatite-based mineralized tissues is a formidable challenge to quantitative imaging. For example, even energy-filtered electron microscopy is not suitable for detection of small quantities of low atomic number elements typical for biological materials. Herein we show that laser-pulsed atom probe tomography, a technique that combines subnanometer spatial resolution with unbiased chemical sensitivity, is uniquely suited to the task. Common apatite end members share a number of features, but can clearly be distinguished by their spectrometric fingerprint. This fingerprint and the formation of molecular ions during field evaporation can be explained based on the chemistry of the apatite channel ion. Using end members for reference, we are able to interpret the spectra of bone and dentin samples, and generate the first three-dimensional reconstruction of 1.2 × 10(7) atoms in a dentin sample. The fibrous nature of the collagenous organic matrix in dentin is clearly recognizable in the reconstruction. Surprisingly, some fibers show selectivity in binding for sodium ions over magnesium ions, implying that an additional, chemical level of hierarchy is necessary to describe dentin structure. Furthermore, segregation of inorganic ions or small organic molecules to homophase interfaces (grain boundaries) is not apparent. This has implications for the platelet model for apatite biominerals.

Direct measurements of exciton diffusion length limitations on organic solar cell performance

Through a combination of X-ray scattering and energy-filtered electron microscopy, we have quantitatively examined the relationship between the mesostructure of the photoactive layer and device performance in PBTTT/PC(71)BM solar cells. We can predict device performance from X-ray structural data through a simple morphological model which includes the exciton diffusion length.

Nano-sized CuO, TiO2 and ZnO affect Xenopus laevis development

The teratogenic potential of commercially available copper oxide (CuO), titanium dioxide (TiO2) and zinc oxide (ZnO) nanoparticles (NPs) was evaluated using the standardized FETAX test. After characterization of NP suspensions by TEM, DLS and AAS, histopathological screening and advanced confocal and energy-filtered electron microscopy techniques were used to characterize the induced lesions and to track NPs in tissues. Except for nCuO, which was found to be weakly embryolethal only at the highest concentration tested, the NPs did not cause mortality at concentrations up to 500 mg/L. However, they induced significant malformation rates, and the gut was observed to be the main target organ. CuO NPs exhibited the highest teratogenic potential, although no specific terata were observed. ZnO NPs caused the most severe lesions to the intestinal barrier, allowing NPs to reach the underlying tissues. TiO2 NPs showed mild embryotoxicity, and it is possible that this substance could be associated with hidden biological effects. Ions from dissolved nCuO contributed greatly to the observed embryotoxic effects, but those from nZnO did not, suggesting that their mechanisms of action may be different.

Quantitative chemical analysis of ocular melanosomes in stained and non-stained tissues

Energy-filtered Analytical Electron Microscopy (AEM) was used to image the ultrastructure and determine quantitatively the chemical composition of rat melanosomes of the choroid and the Retinal Pigment Epithelium (RPE). For the first time, the effect of staining in elemental analysis of melanosomes was investigated. Detection limits and accuracies of the applied methods were determined.Compared to previous work applying only quantitative Energy Dispersive X-ray microanalysis (EDX) in the TEM (Eibl, O., et al., 2006. Micron 37, 262), here we present a combined quantitative EDX and Electron Energy Loss Spectroscopy (EELS) analysis, including N. This yields the fraction of eumelanin and pheomelanin in melanosomes by the S/N mole fraction ratio. Melanosomes of the sepia ink sac, used as eumelanin standard, showed an S/N mole fraction ratio of <0.004. Thus, they consist primarily of eumelanin as reported by degradation analysis. In contrast, melanosomes of the rats contained mixed melanin with significant amounts of pheomelanin (S/N 0.02) in the RPE and the choroid. Consistent with the previous publication, it was shown that oxygen mole fractions are especially large in melanosomes (7–10at.%) compared to other cell compartments, e.g. 2–4at.% oxygen in the cytoplasm. In the melanosomes of non-stained tissue, the oxygen mole fraction clearly correlated with the Ca mole fraction.EDX spectra used for quantitative analysis had about 15,000 net counts under the oxygen peak, which is necessary to obtain (i) a small statistical error for oxygen and (ii) optimum minimum detectable mole fractions for S, Ca and transition metals. The precise determination of the oxygen mole fraction in melanosomes is important for understanding metabolism. Therefore, a detailed analysis was carried out on the possible errors affecting quantification.While O, S, and N mole fractions yielded similar results in stained and non-stained ocular melanosomes of rats, transition metals can only be determined reliably in non-stained tissues. High-precision EDX analysis of melanosomes yielded minimum detectable mole fractions of less than 0.04at.% for Cu and Zn, these elements were present in melanosomes with mole fractions of about 0.3at.% and 0.1at.%, respectively. Zn is of great importance for metabolism and for age related macular degeneration. Its mole fraction in melanosomes of rats is large enough to be detected and to be quantitatively analyzed by EDX spectroscopy. Ultrastructural information can now be correlated to the elemental composition. This is important to better understand the physical and chemical properties of melanosomal metabolism and turnover.

Experimental setup for energy-filtered scanning confocal electron microscopy (EFSCEM) in a double aberration-corrected transmission electron microscope

Scanning confocal electron microscopy (SCEM) is a new imaging mode in electron microscopy. Spherical aberration corrected electron microscope instruments fitted with two aberration correctors can be used in this mode which provides improved depth resolution and selectivity compared to optical sectioning in a conventional scanning transmission geometry. In this article, we consider a confocal optical configuration for SCEM using inelastically scattered electrons. We lay out the necessary steps for achieving this new operational mode in a double aberration-corrected instrument with uncorrected chromatic aberration and present preliminary experimental results in such mode.

Titanium uptake, induction of RANK‐L expression, and enhanced proliferation of human T‐lymphocytes

There is increasing evidence that titanium ions are released from orthopedic implants by biocorrosion. The aim of this study was to investigate titanium uptake by human T-lymphocytes and its effects on phenotype and proliferation. Freshly isolated human nonadherent peripheral blood mononuclear cells (NA-PBMC), were exposed to TiCl4 [Ti(IV)]. Bioavailability and distribution of Ti(IV) in T-lymphocytes was determined by energy-filtered electron microscopy (EFTEM). The effects of Ti(IV) challenge on nonactivated and PHA-activated cells were assessed by flow cytometric analysis of surface markers, RANK-L production, and proliferation assays. EFTEM colocalized Ti(IV) with phosphorus in the nucleus, ribosomes, cytoplasmic membranes, and the surface membrane of T-lymphocytes. Ti(IV) increased significantly the expression of CD69, CCR4, and RANK-L in a concentration-dependent manner. Titanium enters T-lymphocytes through a currently unknown mechanism and binds to phosphorus-rich cell structures. Titanium influences phenotype and function of T-lymphocytes, resulting in activation of a CD69+ and CCR4+ T-lymphocyte population and secretion of RANK-L. These results strongly suggest the involvement of titanium ions challenged T-lymphocytes in the complex pathophysiological mechanisms of aseptic loosening of orthopedic implants.

Identifying Spinel Phases in Nearly Monodisperse Iron Oxide Colloidal Nanocrystal

Nearly monodisperse iron oxide colloidal nanocrystals prepared by nonhydrolytic high-temperature solution method were obtained with two different sizes and degrees of oxidation. The characterization of the structural features of the nanocrystals was performed by a multitechnique approach including transmission electron microscopy, X-ray diffraction and X-ray absorption spectroscopy, energy filtered electron microscopy imaging, and SQUID magnetometry. The different techniques provided complementary information on the local oxidation state of iron in the iron oxide nanoparticles, the stability of the phases, the exact crystal structure, and the compositional homogeneity. X-ray diffraction, transmission electron microscopy, and extended X-ray absorption spectroscopy show that the addition of oxidizer to the iron precursor gives rise to monodisperse polycrystalline nanoparticles made out of FeO plus a spinel phase. X-ray absorption near-edge structure, which is very sensitive to the oxidation state and local environment of iron in the different iron oxides, was used to distinguish among isostructural spinel phases of iron (II,III) oxide (magnetite) and iron(III) oxide (maghemite). Single-crystalline spinel nanoparticles are obtained upon sequential oxidation: in smaller nanoparticles a mixture of mainly Fe3O4 and γ-Fe2O3 is present, whereas the larger nanoparticles are made out of γ-Fe2O3, as also supported by SQUID magnetization measurements. The importance of a multitechnique approach for the elucidation of the compositional and structural details in addition to geometrical parameters in the characterization of nanocrystalline iron oxides is pointed out.

Au@Pd Core−Shell Nanoparticles through Digestive Ripening

Colloids of gold and palladium nanoparticles have been prepared by the Solvated Metal Atom Dispersion (SMAD) method. Digestive ripening, a process involving mixing of the as-prepared gold and palladium colloids consisting of polydisperse nanoparticles and refluxing the mixture in the presence of a surfactant resulted in highly monodisperse Au@Pd core−shell nanoparticles with an average diameter of 6.6 ± 0.5 nm. The synthesis has been successfully carried out reproducibly on gram scale. The core−shell structure was established by using UV−visible spectroscopy, high-resolution electron microscopy, energy filtered electron microscopy, energy dispersive X-ray analysis, high angle annular dark field imaging, and powder X-ray diffraction methods.

Synthesis of Cu@ZnO Core−Shell Nanocomposite through Digestive Ripening of Cu and Zn Nanoparticles

The synthesis of colloids of copper and zinc nanoparticles by solvated metal atom dispersion (SMAD) is described. The as-prepared colloids with a large size distribution of the particles are transformed into colloidal nanoparticles of a narrow size distribution by the digestive ripening process which involves refluxing the colloid at or near the boiling point of the solvent in the presence of a passivating ligand. The copper nanoparticles of 2.1 ± 0.3 nm and zinc nanoparticles of 3.9 ± 0.3 nm diameters have thus been obtained. Digestive ripening of the as-prepared copper and zinc colloids together in the presence of a passivating agent gave Cu@ZnO core−shell nanoparticles, with an average diameter of 3.0 ± 0.7 nm. Particles synthesized in this manner were characterized by UV−visible spectroscopy, high-resolution electron microscopy, energy-filtered electron microscopy, and powder X-ray diffraction methods which confirm the core−shell structure.

Polyphosphate at the Streptomyces lividans cytoplasmic membrane is enhanced in the presence of the potassium channel KcsA

The distribution of polyphosphate (polyP) within the cytoplasmic membrane of Streptomyces lividans hyphae or protoplasts has been determined at high spatial resolution by elemental mapping using energy-filtered electron microscopy (EFTEM). The results revealed that polyP was best traceable after its interaction with lead ions followed by their precipitation as lead sulphide. Concomitant studies of the S.lividans wildtype (WT) strain and its co-embedded mutant DeltaK (lacking a functional kcsA gene) were conducted by labelling as the surface matrix of either one was labelled by cationic colloidal thorium dioxide. Within the WT strain, additional polyP was found to accumulate distinctly at the inner face of the cytoplasmic membrane. After removal of the cell wall (within protoplasts), the polyP-derived lead-sulphide (PbS) precipitate formed clusters of fibrillar material extending up to 50 nm into the cytoplasm. This feature was absent in the DeltaK mutant strain. Together the results revealed that the presence of the KcsA channel and the structured polyP coincide.

Energy-filtered electron microscopy for imaging core–shell nanostructures

CuAg core–shell nanoparticles are synthesized by ultra-high vacuum thermal evaporation. We show on this system how the Energy-Filtered Transmission Electron Microscopy (EFTEM) technique allows one to improve the characterization by precisely pointing out the formation of core–shell arrangements in bimetallic nanoparticle assemblies. A criterion to measure the shell thickness from EFTEM images on unique core–shell nanoparticles is defined, that can be used for core–shell nanoparticles of any sizes, with shell thicknesses over 1 nm. It is based on the intensity variation along a line drawn across a core–shell nanoparticle on a EFTEM image. This criterion has been validated by a close comparison of the shell thickness measurements performed in this work and the ones obtained by acoustic micro-Raman spectroscopy. Using this criterion, we report a strong correlation between the size of the Cu cores and the formation of the core–shell arrangements in the nanoparticle assembly studied in this work. The influence of the Cu core shape is also evidenced. The characterisation of such systems using High Resolution TEM (HRTEM) is also discussed.

Energy-filtered X-ray photoemission electron microscopy and its applications to surface and organic materials

Energy-filtered X-ray photoemission electron microscopy (EXPEEM) is a new surface chemical imaging method that combines X-ray photoelectron spectroscopy (XPS) and photoemission electron microscopy (PEEM). We have developed a collinear type EXPEEM system using a Wien-filter-type electron energy analyzer. The collinear arrangement has the advantage of carrying out an easy alignment of the electron optical axis. We have measured EXPEEM images, μ-X-ray absorption near edge structure (μ-XANES) and μ-XPS of Au on Ta and Ag(DM) 2. We discuss the advantage of EXPEEM and future applications to organic devices.