Comparing electron tomography and HRTEM slicing methods as tools to measure the thickness of nanoparticles

Nanoparticles’ morphology is a key parameter in the understanding of their thermodynamical, optical, magnetic and catalytic properties. In general, nanoparticles, observed in transmission electron microscopy (TEM), are viewed in projection so that the determination of their thickness (along the projection direction) with respect to their projected lateral size is highly questionable. To date, the widely used methods to measure nanoparticles thickness in a transmission electron microscope are to use cross-section images or focal series in high-resolution transmission electron microscopy imaging (HRTEM “slicing”). In this paper, we compare the focal series method with the electron tomography method to show that both techniques yield similar particle thickness in a range of size from 1 to 5 nm, but the electron tomography method provides better statistics since more particles can be analyzed at one time. For this purpose, we have compared, on the same samples, the nanoparticles thickness measurements obtained from focal series with the ones determined from cross-section profiles of tomograms (tomogram slicing) perpendicular to the plane of the substrate supporting the nanoparticles. The methodology is finally applied to the comparison of CoPt nanoparticles annealed ex situ at two different temperatures to illustrate the accuracy of the techniques in detecting small particle thickness changes.     

Metrological challenges for reconstruction of 3‐D microstructures by focused ion beam tomography methods

The development of combined focused ion beam and scanning electron microscopes has enabled significant advances in the characterization of the 3‐D structure of materials. The repeated removal of thin layers or slices with an ion beam and imaging or mapping the chemical or crystallographic structure of each slice enables a 3‐D reconstruction from the images or maps. The accuracy of the reconstruction thus depends on the accuracy with which the slice thickness is measured and maintained throughout the process, and the alignment accuracy of the slices achieved during acquisition or by postacquisition corrections. A survey of papers published in this field suggests that the reconstruction accuracy is not often considered or reported. Using examples from examination of the 3‐D structure of hardmetals, issues affecting the accuracy of slice thicknesses and image realignments are examined and illustrated and potential errors quantified by the use of fiducial markers and the expected isotropy of the hardmetal structure itself.     

Non‐rigid alignment in electron tomography in materials science

Electron tomography is a key technique that enables the visualization of an object in three dimensions with a resolution of about a nanometre. High‐quality 3D reconstruction is possible thanks to the latest compressed sensing algorithms and/or better alignment and preprocessing of the 2D projections. Rigid alignment of 2D projections is routine in electron tomography. However, it cannot correct misalignments induced by (i) deformations of the sample due to radiation damage or (ii) drifting of the sample during the acquisition of an image in scanning transmission electron microscope mode. In both cases, those misalignments can give rise to artefacts in the reconstruction. We propose a simple‐to‐implement non‐rigid alignment technique to correct those artefacts. This technique is particularly suited for needle‐shaped samples in materials science. It is initiated by a rigid alignment of the projections and it is then followed by several rigid alignments of different parts of the projections. Piecewise linear deformations are applied to each projection to force them to simultaneously satisfy the rigid alignments of the different parts. The efficiency of this technique is demonstrated on three samples, an intermetallic sample with deformation misalignments due to a high electron dose typical to spectroscopic electron tomography, a porous silicon sample with an extremely thin end particularly sensitive to electron beam and another porous silicon sample that was drifting during image acquisitions.     

Morphology visualization of irregular shape bacteria by electron holography and tomography

In the current work, irregular morphology of Staphylococcus aureus bacteria has been visualized by phase retrieval employing off‐axis electron holography (EH) and 3D reconstruction electron tomography using high‐angle annular dark field scanning transmission electron microscopy (HAADF‐STEM). Bacteria interacting with gold nanoparticles (AuNP) acquired a shrunken or irregular shape due to air dehydration processing. STEM imaging shows the attachment of AuNP on the surface of cells and suggests an irregular 3D morphology of the specimen. The phase reconstruction demonstrates that off‐axis electron holography can reveal with a single hologram the morphology of the specimen and the distribution of the functionalized AuNPs. In addition, EH reduces significantly the acquisition time and the cumulative radiation damage (in three orders of magnitude) over biological samples in comparison with multiple tilted electron expositions intrinsic to electron tomography, as well as the processing time and the reconstruction artifacts that may arise during tomogram reconstruction.     

Evaluation of electron tomography reconstruction methods for interface roughness measurement

We evaluate the suitability of simultaneous iterative reconstruction technique (SIRT), filtered back projection, and simultaneous algebraic reconstruction technique methods for buried interface roughness measurements. We also investigate the effect of total electron dose distributed over the entire tilt series on measured roughness values. We investigate the applicability of the dose fractionation theorem by evaluating the effect of an increasing number of images, i.e., decreasing tilt increment size at fixed total electron irradiation dose on the quantitative measurement of buried interface roughness. The results indicate that SIRT is the most suitable method for reconstruction and a 3° to 5° angle is optimal for the roughness measurement.     

Correcting for 3D distortion when using backscattered electron detectors in a scanning electron microscope

A variable pressure scanning electron microscope (VPSEM) can produce a topographic surface relief of a physical object under examination, in addition to its two‐dimensional (2D) image. This topographic surface relief is especially helpful when dealing with porous rock because it may elucidate the pore‐space structure as well as grain shape and size. Whether the image accurately reproduces the physical object depends on the management of the hardware, acquisition, and postprocessing. Two problems become apparent during testing: (a) a topographic surface relief of a precision ball bearing is distorted and does not correspond to the physical dimensions of the actual sphere and (b) an image of a topographic surface relief of a Berea sandstone is geometrically tilted and topographically distorted even after standard corrections are applied. The procedure presented here is to ensure the veracity of the image, and includes: (a) adjusting the brightness and contrast levels originally provided by the manufacturer and (b) tuning the amplifiers of the backscatter detector plates to be equal to each other, and producing zero voltage when VPSEM is idle. This procedure is tested and verified on the said two physical samples. SCANNING 31: 59–64, 2009. © 2009 Wiley Periodicals, Inc.     

一种光学元件面形三维重建的算法研究

为了在线检测光学元件面形,介绍一种基于线结构光扫描测量和立体视觉测量相结合的三维检测方法,目前这种方法多用于检测高反射率的物体,因此将该方法运用于检测光学元件面形是一种新的尝试。实验的三维重建算法是通过MATLAB和VC++软件共同编写程序实现的,实验结果表明,将这种方法运用于检测光学元件是可以真实还原光学元件三维外貌特性的,所以它的研究具有可行性和研究价值。     

3D electromagnetic tomography using a single layer sensor array

Electromagnetic tomography (EMT) is a non-invasive technique to gain the induced voltages and to reconstruct the distribution of electromagnetic properties. A volume imaging using a single layer sensor array, namely, Single Layer 3D-EMT, has been developed in this paper. To solve the forward problem, three kinds of sensor arrays are established with different number of coils,—8, 12 and 16, to conduct simulations. Four typical conductivity distributions are used to verify the 3D-EMT imaging with single layer sensor array. The calculation of sensitivity maps in 3D views is presented, and the sensitivity maps of different layers, from z = 0 to z = 20, are analyzed. The conjugate gradient iterative method has been adopted in image reconstruction. It can be seen from the simulation results that the 3D reconstructed images using a single layer sensor array can display the distribution of the measured objects in position, shape, and size. By calculating the relative error, the quality of the reconstructed images is evaluated andthe result shows that the 12-coil sensor array has the best image quality among three sensor arrays. In addition, the single layer 3D experimental system has been established to evaluate the effectiveness of 3D imaging using a single layer sensor array. In order to save the hardware resources, the paper presents the author's research work about the possibility of Single Layer 3D-EMT imaging in simulations and experiments. This can also provide a guide to explore the potential application with electromagnetic testing technique.     

3-D reconstruction of the atomic positions in a simulated gold nanocrystal based on discrete tomography: prospects of atomic resolution electron tomography

A novel reconstruction procedure is proposed to achieve atomic resolution in electron tomography. The method exploits the fact that crystals are discrete assemblies of atoms (atomicity). This constraint enables us to obtain a three-dimensional (3-D) reconstruction of test structures from less than 10 projections even in the presence of noise and defects. Phase contrast transmission electron microscopy (TEM) images of a gold nanocrystal were simulated in six different zone axes. The discrete number of atoms in every column is determined by application of the channelling theory to reconstructed electron exit waves. The procedure is experimentally validated by experiments with gold samples. Our results show that discrete tomography recovers the shape of the particle as well as the position of its 309 atoms from only three projections.

Experiments on a nanocrystal that contains several missing atoms, both on the surface and in the core of the nanocrystal, while considering a high noise level in each simulated image were performed to prove the stability of the approach to reconstruct defects. The algorithm is well capable of handling structural defects in a highly noisy environment, even if this causes atom count “errors” in the projection data.     

Study of multi-electrode excitation mode for 3D electrical resistance tomography

Multi-electrode excitation is an effective way to improve the performance of 3D electrical tomography (ET) system compared with single electrode excitation. This paper systematically discusses various sensing strategies for multi-electrode excitation with different combinations in radial and axial directions. A typical 3 × 8 3D ERT sensor is adopted to analyze the influence of different excitation modes on performance indexes, such as number of independent measurements, dynamic range of measurements, sensitivity distribution and correlation coefficient of image reconstruction. On the basis of radial rotation angle between the excitation electrodes mapped to the same plane and whether the excitation electrodes are located in the same axial layer, five typical electrode combination modes are designed under dual-electrode excitation and single-electrode measurement protocol. The results show that the variation trend of different indexes influenced by excitation mode is inconsistent. The entropy-based method is applied to comprehensively evaluate the different excitation modes with the selected multiple performance indexes. Among all above modes, the mode in which excitation electrodes are vertically aligned is better universal one. The experimental results show that the position characteristics of the preferable mode can obtain better image reconstruction quality in different distribution models. Furthermore, the better excitation mode will provide guidance for design of other multi-electrode excitation in 3D ET systems with different structures.     

Reduction of the scanning time by total variation minimization reconstruction for X‐ray tomography in a SEM

Total variation minimization is applied to the particular case of X‐ray tomography in a scanning electron microscope. To prove the efficiency of this reconstruction method, noise‐free and noisy data based on the Shepp & Logan phantom have been simulated. These simulations confirm that Total variation minimization‐reconstruction algorithm better manages data containing low number of projections with respect to simultaneous iterative reconstruction technique or filtered backprojection, even in the presence of noise. The algorithm has been applied to real data sets, with a low angular sampling and a high level of noise. Two samples containing micro‐interconnections have been analyzed and 3D reconstructions show that Total variation minimization‐based algorithm performs well even with 60 projections in order to properly recover a 500 nm diameter void inside a copper interconnection.     

Characterization and reconstruction of 3D stochastic microstructures via supervised learning

The need for computational characterization and reconstruction of volumetric maps of stochastic microstructures for understanding the role of material structure in the processing–structure–property chain has been highlighted in the literature. Recently, a promising characterization and reconstruction approach has been developed where the essential idea is to convert the digitized microstructure image into an appropriate training dataset to learn the stochastic nature of the morphology by fitting a supervised learning model to the dataset. This compact model can subsequently be used to efficiently reconstruct as many statistically equivalent microstructure samples as desired. The goal of this paper is to build upon the developed approach in three major directions by: (1) extending the approach to characterize 3D stochastic microstructures and efficiently reconstruct 3D samples, (2) improving the performance of the approach by incorporating user‐defined predictors into the supervised learning model, and (3) addressing potential computational issues by introducing a reduced model which can perform as effectively as the full model. We test the extended approach on three examples and show that the spatial dependencies, as evaluated via various measures, are well preserved in the reconstructed samples.     

Method for 3D fibre reconstruction on a microrobotic platform

Automated handling of a natural fibrous object requires a method for acquiring the three‐dimensional geometry of the object, because its dimensions cannot be known beforehand. This paper presents a method for calculating the three‐dimensional reconstruction of a paper fibre on a microrobotic platform that contains two microscope cameras. The method is based on detecting curvature changes in the fibre centreline, and using them as the corresponding points between the different views of the images. We test the developed method with four fibre samples and compare the results with the references measured with an X‐ray microtomography device. We rotate the samples through 16 different orientations on the platform and calculate the three‐dimensional reconstruction to test the repeatability of the algorithm and its sensitivity to the orientation of the sample. We also test the noise sensitivity of the algorithm, and record the mismatch rate of the correspondences provided. We use the iterative closest point algorithm to align the measured three‐dimensional reconstructions with the references. The average point‐to‐point distances between the reconstructed fibre centrelines and the references are 20–30 μm, and the mismatch rate is low. Given the manipulation tolerance, this shows that the method is well suited to automated fibre grasping. This has also been demonstrated with actual grasping experiments.     

A vision-based, 3D reconstruction technique for scanning electron microscopy: direct comparison with atomic force microscopy

High-resolution, detailed 3D reconstructions of biological specimens obtained from scanning electron microscopy stereo-micrographs and proprietary software were compared with Tapping-Mode AFM datasets of the same fields. The reconstruction software implements several original solutions including a neural adaptive point-matching technique, the ability to build an irregular triangulated mesh rather than a regular orthogonal grid, and the ability to re-map one of the original images exactly onto the reconstructed surface. The technique was applied to human nerve tissue to obtain 1,424 x 968-pixel, texture-mapped datasets, which were subsequently compared against 512 x 512-pixel AFM datasets from the same viewfields. Accounting for the inherent differences of the two techniques, direct comparison revealed an excellent visual match. The correspondence was also quantified by calculating the cross-correlation coefficient between corresponding altimetric profiles in SEM and AFM data, which consistently exceeded a figure of 0.9, with a rate of point mismatch in the order of 0.01%. Research is still underway to improve the robustness of the technique when applied to arbitrary images     

Cornea and its adaptation to environment and accommodation function in veiled chameleon (Chamaeleo calyptratus): ultrastructure and 3D transmission electron tomography

We investigate the ultrastructural features and 3D electron tomography of chameleon (Chamaeleon calyptratus) which is a native of desert environments of Saudi Arabia. The corneas of the chameleon were fixed in 2.5% glutaraldehyde containing cuprolinic blue in sodium acetate buffer for electron microscopy and tomography, and observed under a JEOL 1400 transmission electron microscope. The thin cornea (21.92 μm) contained 28–30 collagen fibril lamellae. The middle stromal lamellae (from 13 to 19) contained keratocytes with a long cell process and filled with granular material. The CF diameter increased from lamella 1 (30.44 ± 1.03) to Lamella 5 (52.83 ± 2.00) then decreased towards the posterior stoma. The percentage of large CF diameters (55–65 nm) was very high in the lamellae L14 (38.8%) and L15 (85.7%). The mean PGs area of the posterior stroma (448.21 ± 24.84 nm²) was significantly larger than the mean PGs area of the anterior, (309.86 ± 8.2 nm²) and middle stroma 245.94 ± 8.28 nm²). 3D electron tomography showed the distribution of PGs around and over the CF. Variable diameters of CFs in the anterior stroma may provide compact lamellae which may restrict the low wavelength of light. Variable diameters of CFs in the anterior stroma may provide compact lamellae which may restrict the low wavelength of light. This accommodation function is achieved by bending of the cornea. During bending the anterior stroma was stretched and the posterior stroma was compressed due to the presence of small CFs. The middle stroma remained stiff due to the presence of large CFs. Large proteoglycans not only maintain hydration for a longer period of time, but also act as a lubricant to neutralise the shear forces in the anterior and posterior stroma during bending.     

3D reconstruction of prior β grains in friction stir–processed Ti–6Al–4V

The prior β grain structure and orientations in the central stir zone of friction stir–processed Ti–6Al–4V were reconstructed from measured α phase orientations obtained by three‐dimensional serial sectioning in a dual‐beam focused ion beam scanning electron microscope. The data were processed to obtain the α colony and β grain size distributions in the volume. Several β grains were individually analysed to determine the total number of unique α variants and the respective volume fractions of each. The analysis revealed that some grains experienced overwhelming variant selection (i.e. one variant dominated) whereas other β grains contained a more evenly distributed mixture of all 12 variants.     

Towards automated electron holographic tomography for 3D mapping of electrostatic potentials

Electron-holographic tomography (EHT), that is, the combination of off-axis electron holography with electron tomography, was successfully applied for the quantitative 3D mapping of electrostatic potentials at the nanoscale. Here we present the first software package (THOMAS) for semi-automated acquisition of holographic tilt series, a prerequisite for efficient data collection. Using THOMAS, the acquisition time for a holographic tilt series, consisting of object and reference holograms, is reduced by a factor of five on average, compared to the previous, completely manual approaches. Moreover, the existing software packages for retrieving amplitude and phase information from electron holograms have been extended, now including a one-step procedure for holographic tilt series reconstruction. Furthermore, a modified SIRT algorithm (WSIRT) was implemented for the quantitative 3D reconstruction of the electrostatic potential from the aligned phase tilt series. Finally, the application of EHT to a polystyrene latex sphere test-specimen and a pn-doped Ge ‘needle’-shaped specimen are presented, illustrating the quantitative character of EHT. For both specimens the mean inner potential (MIP) values were accurately determined from the reconstructed 3D potential. For the Ge specimen, additionally the ‘built-in’ voltage across the pn junction of 0.5 V was obtained.     

X-ray microfocus computed tomography: a powerful tool for structural and functional characterisation of 3D printed dosage forms

One of the most promising advances in modern pharmaceutical technology is the introduction of three-dimensional (3D) printing technology for the fabrication of drug products. 3D printed dosage forms have the potential to revolutionise pharmacotherapy as streamlined production of structurally complex formulations with optimal drug releasing properties is now made possible. 3D printed formulations are derived as part of a process where a ‘print-head’ deposits, or sinters material under computer control to produce a drug carrier. However, this manufacturing route inherently generates objects that deviate from the ideal designed template for reasons specific to the 3D printing method used. This short opinion article discusses the potential of high-resolution nondestructive 3D (volume) imaging by means of X-ray microfocus Computed Tomography (μCT) as a Process Analytical Technology for the structural and functional characterisation of 3D printed dosage forms.