Optimized deconvolution for maximum axial resolution in three-dimensional aberration-corrected scanning transmission electron microscopy

Three-dimensional (3D) datasets were recorded of gold nanoparticles placed on both sides of silicon nitride membranes using focal series aberration-corrected scanning transmission electron microscopy (STEM). Deconvolution of the 3D datasets was applied to obtain the highest possible axial resolution. The deconvolution involved two different point spread functions, each calculated iteratively via blind deconvolution. Supporting membranes of different thicknesses were tested to study the effect of beam broadening on the deconvolution. It was found that several iterations of deconvolution was efficient in reducing the imaging noise. With an increasing number of iterations, the axial resolution was increased, and most of the structural information was preserved. Additional iterations improved the axial resolution by maximal a factor of 4 to 6, depending on the particular dataset, and up to 8 nm maximal, but also led to a reduction of the lateral size of the nanoparticles in the image. Thus, the deconvolution procedure optimized for the highest axial resolution is best suited for applications where one is interested in the 3D locations of nanoparticles only.     

Gold clusters showing pentagonal atomic arrays revealed by aberration-corrected scanning transmission electron microscopy

In this work we present the analysis by aberration corrected electron microscopy of the formation of gold clusters based on the proton irradiation of larger nanoparticles (NP). Pentagonal arrays have been observed and energetic calculations have been performed in order to prove the stability of these materials.     

Progress in aberration-corrected high-resolution transmission electron microscopy using hardware aberration correction.

The design and construction of a double-hexapole aberration corrector has made it possible to build the prototype of a spherical-aberration corrected transmission electron microscope dedicated to high-resolution imaging on the atomic scale. The corrected instrument, a Philips CM200 FEG ST, has an information limit of better than 0.13 nm, and the spherical aberration can be varied within wide limits, even to negative values. The aberration measurement and the corrector control provide instrument alignments stable enough for materials science investigations. Analysis of the contrast transfer with the possibility of tunable spherical aberration has revealed new imaging modes: high-resolution amplitude contrast, extension of the point resolution to the information limit, and enhanced image intensity modulation for negative phase contrast. In particular, through the combination of small negative spherical aberration and small overfocus, the latter mode provides the high-resolution imaging of weakly scattering atom columns, such as oxygen, in the vicinity of strongly scattering atom columns. This article reviews further lens aberration theory, the principle of aberration correction through multipole lenses, aspects for practical work, and materials science applications.     

Imaging of block copolymer vesicles in solvated state by wet scanning transmission electron microscopy

Morphology of polystyrene-block-poly(acrylic acid) vesicles was imaged by various modes of scanning electron microscopy (SEM), including recently developed wet scanning transmission electron microscopy (wet-STEM) by means of which we were able to follow the decrease in the thickness of a liquid solvent layer around the vesicles during controlled evaporation of water from the sample. Results show that wet-STEM allows for imaging of nanosized polymeric particles in the presence of the solvent.     

Electron microscopy methods for studying polymer blends—comparison of scanning electron microscopy and transmission electron microscopy

The possibility of using a scanning electron microscope (SEM) for studying the morphology of mechanical polymer blends was investigated. Compounds of SBS/EPDM, and both filled and unfilled NBR/EPDM were tested. OsO₄-stained thin-sections were also examined in a transmission electron microscope (TEM) and the results were compared.It seemed to be quite possible to use atomic number contrast detection in combination with OsO₄ staining for visualizing the morphology of the blends in SEM. Domains as small as 0·1 μm were clearly seen. This was done by means of a Robinson backscattered electron detector. Sample preparation was easy, 2 mm thick rubber plates were cut on dry ice to obtain a smooth surface. After staining, the samples were coated with a thin conductive carbon layer.The inner structures of SBS and the carbon black particles were not resolved in SEM but were easily seen in TEM.     

Ultramicrohardness tester for use in a scanning electron microscope

Summary A novel microhardness-tester has been designed for loads between 10^–2 N and 5 x 10^–5 N. The apparatus can be operated in a scanning electron microscope without installation of additional adjusting shafts. The force is generated electromagnetically, transduced to the indenter (diamond cone or pyramid) by bending a double leaf spring, and measured by means of strain gauges fixed to the springs. The indentation cycle is fully programmable and is controlled by the strain gauge signal. The equipment can be tilted with respect to the electron beam thus making possible the observation of the tip during indentation.With 2 figures     

Atomic-scale imaging and spectroscopy for in situ liquid scanning transmission electron microscopy

Observation of growth, synthesis, dynamics, and electrochemical reactions in the liquid state is an important yet largely unstudied aspect of nanotechnology. The only techniques that can potentially provide the insights necessary to advance our understanding of these mechanisms is simultaneous atomic-scale imaging and quantitative chemical analysis (through spectroscopy) under environmental conditions in the transmission electron microscope. In this study we describe the experimental and technical conditions necessary to obtain electron energy loss (EEL) spectra from a nanoparticle in colloidal suspension using aberration-corrected scanning transmission electron microscopy (STEM) combined with the environmental liquid stage. At a fluid path length below 400 nm, atomic resolution images can be obtained and simultaneous compositional analysis can be achieved. We show that EEL spectroscopy can be used to quantify the total fluid path length around the nanoparticle and demonstrate that characteristic core-loss signals from the suspended nanoparticles can be resolved and analyzed to provide information on the local interfacial chemistry with the surrounding environment. The combined approach using aberration-corrected STEM and EEL spectra with the in situ fluid stage demonstrates a plenary platform for detailed investigations of solution-based catalysis.     

Spatial resolution and information transfer in scanning transmission electron microscopy

The relation between image resolution and information transfer is explored. It is shown that the existence of higher frequency transfer in the image is just a necessary but not sufficient condition for the achievement of higher resolution. Adopting a two-point resolution criterion, we suggest that a 10% contrast level between two features in an image should be used as a practical definition of resolution. In the context of scanning transmission electron microscopy, it is shown that the channeling effect does not have a direct connection with image resolution because sharp channeling peaks do not move with the scanning probe. Through a quantitative comparison between experimental image and simulation, a Fourier-space approach is proposed to estimate defocus and sample thickness. The effective atom size in Z-contrast imaging depends on the annular detector's inner angle. Therefore, an optimum angle exists for the highest resolution as a trade-off between reduced atom size and reduced signal with limited information transfer due to noise.     

Laser experiments in a scanning electron microscope

In order to increase the information content of electron microscopic investigations a scanning electron microscope was combined with a pulsed Nd-glass laser in both the free running and the Q-switch mode. The equipment allows an in situ observation of melting, sintering and evaporation processes as well as of crack generation and growth at target surfaces. This article describes evaporation experiments for laser PVD of HTSC layers, Langmuir probe measurements in the corresponding plasma and crack processes by thermal shock loading in the SEM.     

High-resolution confocal scanning light microscopy in biology

Confocal scanning light microscopy provides an appreciable improvement in resolving power compared with classical light microscopy; the amount of information that can be extracted from a specimen is increased by a factor of 3. When a laser serves as light source and u.v. wavelengths are used, resolutions of 140 nm are possible. Applications are the study of biological structures in the sub-μm region, for instance, the replicating nucleoid (DNA) morphology of live Escherichia coli and eukaryotic chromosomes.     

Protein-labeling effects in confocal laser scanning microscopy

Confocal laser scanning microscopy (CLSM) is being increasingly used for observing protein uptake in porous chromatography resins. Recent CLSM studies have revealed the possible existence of a nondiffusive protein transport mechanism. Observing protein uptake with CLSM requires labeling the protein with a fluorescent probe. This study examines the effect of the probe identity on the subsequent CLSM adsorption profiles. The adsorption of lysozyme conjugated with different fluorescent probes (Cy5, BODIPY FL, Atto 635, and Atto 520) on SP Sepharose Fast Flow was measured using CLSM and zonal chromatography experiments. Results from zonal chromatography show that the retention time of lysozyme-dye conjugates differ significantly from unlabeled lysozyme. The change in retention of lysozyme upon conjugation with a fluorescent probe is consistent with the difference in net charge between the lysozyme-dye conjugate and unlabeled lysozyme. The adsorption profiles measured by CLSM show significantly different behavior depending upon whether the lysozyme-dye conjugate is retained longer or shorter than the unlabeled lysozyme. These results strongly suggest that the lysozyme concentration overshoot observed in previous CLSM experiments is the result of displacement of weaker binding labeled lysozyme by stronger binding unlabeled lysozyme.     

Charged nanoparticle dynamics in water induced by scanning transmission electron microscopy

Using scanning transmission electron microscopy we image ~4 nm platinum nanoparticles deposited on an insulating membrane, where the membrane is one of two electron-transparent windows separating an aqueous environment from the microscope's high vacuum. Upon receiving a relatively moderate dose of ~10(4) e/nm(2), initially immobile nanoparticles begin to move along trajectories that are directed radially outward from the center of the field of view. With larger dose rates the particle motion becomes increasingly dramatic. These observations demonstrate that, even under mild imaging conditions, the in situ electron microscopy of aqueous environments can produce electrophoretic charging effects that dominate the dynamics of nanoparticles under observation.     

A coincidence transmission electron microscope with digital waveform measuring system for analytical microscopy

We have been developing a new analytical transmission electron microscope (TEM), called a coincidence TEM, which in principle enables elemental mapping images to be observed at a high signal‐to‐noise (S/N) ratio under very low dose radiation conditions. In this paper, we report the development of a coincidence TEM with a digital waveform measuring system for obtaining a coincidence elemental mapping image. In this system, analog signals detected by a Si(Li) detector and a multianode, position‐sensitive photomultiplier (PSPM) are continuously converted into 12‐bit digital waveform data at a rate of 100 MHz, and transferred to a PC. From the transferred digital waveform data, information on X‐ray photon energy, electron incident position, and detection times of both X rays and electrons are calculated by digital waveform measurement, which lead to the observation of a successful coincidence elemental mapping image. Copyright © 2005 John Wiley & Sons, Ltd.     

Electron holography with a Cs-corrected transmission electron microscope.

Cs correctors have revolutionized transmission electron microscopy (TEM) in that they substantially improve point resolution and information limit. The object information is found sharply localized within 0.1 nm, and the intensity image can therefore be interpreted reliably on an atomic scale. However, for a conventional intensity image, the object exit wave can still not be detected completely in that the phase, and hence indispensable object information is missing. Therefore, for example, atomic electric-field distributions or magnetic domain structures cannot be accessed. Off-axis electron holography offers unique possibilities to recover completely the aberration-corrected object wave with uncorrected microscopes and hence we would not need a Cs-corrected microscope for improved lateral resolution. However, the performance of holography is affected by aberrations of the recording TEM in that the signal/noise properties ("phase detection limit") of the reconstructed wave are degraded. Therefore, we have realized off-axis electron holography with a Cs-corrected TEM. The phase detection limit improves by a factor of four. A further advantage is the possibility of fine-tuning the residual aberrations by a posteriori correction. Therefore, a combination of both methods, that is, Cs correction and off-axis electron holography, opens new perspectives for complete TEM analysis on an atomic scale.     

Amyloidosis of Alzheimer's Abeta peptides: solid-state nuclear magnetic resonance, electron paramagnetic resonance, transmission electron microscopy, scanning transmission electron microscopy and atomic force microscopy studies

Aggregation cascade for Alzheimer's amyloid-beta peptides, its relevance to neurotoxicity in the course of Alzheimer's disease and experimental methods useful for these studies are discussed. Details of the solid-phase peptide synthesis and sample preparation procedures for Alzheimer's beta-amyloid fibrils are given. Recent progress in obtaining structural constraints on Abeta-fibrils from solid-state NMR and scanning transmission electron microscopy (STEM) data is discussed. Polymorphism of amyloid fibrils and oligomers of the 'Arctic' mutant of Abeta(1-40) was studied by (1)H,(13)C solid-state NMR, transmission electron microscopy (TEM) and atomic force microscopy (AFM), and a real-time aggregation of different polymorphs of the peptide was observed with the aid of in situ AFM. Recent results on binding of Cu(II) ions and Al-citrate and Al-ATP complexes to amyloid fibrils, as studied by electron paramagnetic resonance (EPR) and solid-state (27)Al NMR techniques, are also presented.     

4D scanning transmission ultrafast electron microscopy: Single-particle imaging and spectroscopy

We report the development of 4D scanning transmission ultrafast electron microscopy (ST-UEM). The method was demonstrated in the imaging of silver nanowires and gold nanoparticles. For the wire, the mechanical motion and shape morphological dynamics were imaged, and from the images we obtained the resonance frequency and the dephasing time of the motion. Moreover, we demonstrate here the simultaneous acquisition of dark-field images and electron energy loss spectra from a single gold nanoparticle, which is not possible with conventional methods. The local probing capabilities of ST-UEM open new avenues for probing dynamic processes, from single isolated to embedded nanostructures, without being affected by the heterogeneous processes of ensemble-averaged dynamics. Such methodology promises to have wide-ranging applications in materials science and in single-particle biological imaging.     

球差校正扫描透射电子显微镜(ac-STEM)探测单原子催化中的活性中心

近年来,"单原子催化"逐渐得到人们的认可与关注,成为催化领域新的前沿与热点之一.高分辨扫描透射电子显微镜(STEM),特别是带有球差校正(ac)的高角环形暗场扫描透射电子显微镜(ac-STEM-HAADF)是唯一能够直接"看到"催化剂中单个原子的工具.在单原子催化的发现、单原子催化剂的开发、催化剂制备的优化、以及单原子催化机理的理解方面具有举足轻重的、甚至是不可替代的作用.本文首先简述了早期的高分辨透射电子显微镜(HRTEM)逐步发展成目前的ac-STEM-HAADF的过程以及过程中各种电镜在单原子观察中的功能.然后综述了ac-STEM在单原子成像、检测单原子分散度及改进催化剂制备参数、检测单原子催化剂中单原子的配位及催化活性中心中的应用及其机遇与挑战.1990年代后期在透镜相差校正领域技术的突破使STEM的分辨率显著提高,因此STEM的单原子成像灵敏度得到明显改善,因而能够对实用催化剂中的单个重原子进行具有很好相衬度的常规检测.2000年代末期刘景月课题组开始进行系统的单原子催化剂合成与表征,表明ac-STEM-HAADF毫无疑问地能够进行实用催化剂中的单个金属原子的常规检测.随后与张涛课题组合作启动了一项采用简便可放大的共沉淀法制备单原子催化剂的项目并取得成功.相比于光谱表征提供样品平均与整体信息,电子显微镜能够提供具有空间分辨的局部信息.对于含有从微米到纳米不同级别不均一性的多相催化剂而言,该技术尤其具有价值.除了能够提供负载金属单原子的空间分布与分散度信息外,如果载体是晶体,ac-STEM-HAADF还能够提供每个金属原子相对于载体表面的空间配位信息.而对于非晶载体例如活性炭、无定型氧化硅等,则以上信息全部丢失,只能提供负载金属原子的空间分布与分散度信息.电子显微镜面临的最主要挑战在于电子束辐射效应:高能电子与样品作用会导致表面金属原子的迁移、表面官能团的消失或改变以及损坏样品等;还会引起环境电镜中气体的离子化.对于大多数催化剂而言,采用低能量电子束能够有效减少电子束辐射影响.由于电子束影响,目前还难以对单原子催化剂进行X射线能量色散谱和电子能量损失谱的常规检测,需要开发更加高效的检测器.此外,电子显微镜只提供三维材料的二维投影,因此在成像过程中不但三维信息丢失,而且对二维投影的解释也具有挑战,因此需要开发具有原子分辨率的三维成像技术.为了更好理解催化剂的制备和催化反应过程,也亟需发展具有单原子分辨率的环境电镜.     

X-Ray fluorescence analysis in the scanning electron microscope

Summary X-ray fluorescence analysis can be performed in a scanning electron microscope equipped with an energy dispersive X-ray spectrometer by means of a special attachment. For the production of primary X-rays a target is positioned in the electron beam. With this technique an improvement of the detection limits is obtained because of the small fraction of bremsstrahlung in the X-ray spectrum. As a disadvantage a loss of lateral resolution occurs. The performance features of different types of construction (transmission and massive targets) are be compared for experimental results and theoretical models.The paper has been presented at 12th Colloquium on Materials Analysis, Institute for Analytical Chemistry, Technical University in Vienna, May 13–15, 1985.