Specimen Drift Research Articles

Automated feedback correction of the specimen drift at the atomic scale

Automated feedback correction of the specimen drift at the atomic scale

Software for automated acquisition of electron tomography tilt series.

For automated acquisition of tilt series for electron tomography, software needs to handle complications such as movements of the sample in x/y and z, increased projected thickness at high tilt, specimen drift, etc. In addition, many applications require special functionality such as low dose acquisition, automated sequential (batch) tomography, or montage tomography. After reviewing how these difficulties can be addressed and a closer look at what advanced acquisition strategies are employed in biosciences, this chapter introduces acquisition software both developed in academia as well as by hardware vendors. It covers the hardware requirements and compatibility, the functional principle and workflow implemented, as well as what advanced functions are supported by the individual programs.

Comparison of first moment STEM with conventional differential phase contrast and the dependence on electron dose

This study addresses the comparison of scanning transmission electron microscopy (STEM) measurements of momentum transfers using the first moment approach and the established method that uses segmented annular detectors. Using an ultrafast pixelated detector to acquire four-dimensional, momentum-resolved STEM signals, both the first moment calculation and the calculation of the differential phase contrast (DPC) signals are done for the same experimental data. In particular, we investigate the ability to correct the segment-based signal to yield a suitable approximation of the first moment for cases beyond the weak phase object approximation. It is found that the measurement of momentum transfers using segmented detectors can approach the first moment measurement as close as 0.13 h/nm in terms of a root mean square (rms) difference in 10 nm thick SrTiO3 for a detector with 16 segments. This amounts to 35% of the rms of the momentum transfers. In addition, we present a statistical analysis of the precision of first moment STEM as a function of dose. For typical experimental settings with recent hardware such as a Medipix3 Merlin camera attached to a probe-corrected STEM, we find that the precision of the measurement of momentum transfers stagnates above certain doses. This means that other instabilities such as specimen drift or scan noise have to be taken into account seriously for measurements that target, e.g., the detection of bonding effects in the charge density.

Temperature calibration of TEM specimen heating holders by isothermal sublimation of silver nanocubes

Micro electro mechanical systems (MEMS) based TEM specimen heating holders exhibit excellent thermal stability and minimal specimen drift, which allows thermally-activated processes to be studied dynamically at high spatial resolution. The advantages of MEMS-based devices arise from the very small thermal masses of the sample studied, but this poses particular challenges for the precise measurement of specimen temperature. Previously, it has been proposed that the size-dependent sublimation behavior of Ag nanoparticles could be used to measure the specimen temperature by applying the Kelvin equation, but the effects of the capping ligands used in the nanoparticle synthesis and of electron beam heating have limited the application of such approaches. Here it is shown that for an appropriate choice of experimental parameters (nanoparticle size, loading, intermediate holding temperature, and illumination conditions) the sublimation of Ag nano-cubes can be used to measure the specimen temperature to an accuracy of ±5 °C, over the range 700–850 °C. The measurements are reproducible from area to area on the same MEMS chip, and from chip to chip of the same type. The values of specimen temperature obtained are consistently lower than the calibrated MEMS heater plate temperatures, and it is shown that this cannot be explained on the basis of random errors in the experimental measurements or systematic errors in the materials parameters used for the Kelvin equation analysis. It is proposed that this is instead due to the low thermal conductivity of the electron-transparent amorphous silicon nitride support membrane on the chip. As further evidence for this, it is shown that for a thicker crystalline Si support with a higher thermal conductivity, the magnitude of the difference is smaller. This approach could be extended to other temperature ranges by using nanoparticles of other metals with different vapor pressures and sublimation temperatures.

Correcting the linear and nonlinear distortions for atomically resolved STEM spectrum and diffraction imaging.

Specimen and stage drift as well as scan distortions can lead to a mismatch between true and desired electron probe positions in scanning transmission electron microscopy (STEM) which can result in both linear and nonlinear distortions in the subsequent experimental images. This problem is intensified in STEM spectrum and diffraction imaging techniques owing to the extended dwell times (pixel exposure time) as compared to conventional STEM imaging. As a consequence, these image distortions become more severe in STEM spectrum/diffraction imaging. This becomes visible as expansion, compression and/or shearing of the crystal lattice, and can even prohibit atomic resolution and thus limits the interpretability of the results. Here, we report a software tool for post-correcting the linear and nonlinear image distortions of atomically resolved 3D spectrum imaging as well as 4D diffraction imaging. This tool improves the interpretability of distorted STEM spectrum/diffraction imaging data.

Robust image alignment for cryogenic transmission electron microscopy

Cryo-electron microscopy recently experienced great improvements in structure resolution due to direct electron detectors with improved contrast and fast read-out leading to single electron counting. High frames rates enabled dose fractionation, where a long exposure is broken into a movie, permitting specimen drift to be registered and corrected. The typical approach for image registration, with high shot noise and low contrast, is multi-reference (MR) cross-correlation. Here we present the software package Zorro, which provides robust drift correction for dose fractionation by use of an intensity-normalized cross-correlation and logistic noise model to weight each cross-correlation in the MR model and filter each cross-correlation optimally. Frames are reliably registered by Zorro with low dose and defocus. Methods to evaluate performance are presented, by use of independently-evaluated even- and odd-frame stacks by trajectory comparison and Fourier ring correlation. Alignment of tiled sub-frames is also introduced, and demonstrated on an example dataset. Zorro source code is available at github.com/CINA/zorro.

Detection of picometer-order atomic displacements in drift-compensated HAADF-STEM images of gold nanorods.

Cs-corrected atomic resolution scanning transmission electron microscopy under a drift-compensated operation enabled us to acquire high-angle annular dark-field (HAADF) images of entire gold nanorods without distortion induced by specimen drift. The precision in locating the atomic columns was evaluated to be ±5 pm in the images thus obtained, which is comparable to the image pixel size. A high-precision HAADF image of a single-crystalline gold nanorod revealed that the tip portions at both ends tended to undergo outward displacements along the rod axis and inward contraction along the perpendicular direction. A single nanosecond pulse shot of laser light with a wavelength of 1064 nm and an average intensity of 7.3 kJ/m2 pulse deformed the nanorods into spherical shapes. Simultaneously, the particle interior was completely changed into a multiple twin structure. Substantial displacements of atomic columns on the order of several tens of picometers were confirmed to be localized in the corners of domains at multiple twin junctions. Both the magnitude and direction of displacements were linearly relaxed with increasing distance from a multiple junction.

Improving Signal-to-Noise Ratio in Scanning Transmission Electron Microscopy Energy-Dispersive X-Ray (STEM-EDX) Spectrum Images Using Single-Atomic-Column Cross-Correlation Averaging.

Acquiring an atomic-resolution compositional map of crystalline specimens has become routine practice, thus opening possibilities for extracting subatomic information from such maps. A key challenge for achieving subatomic precision is the improvement of signal-to-noise ratio (SNR) of compositional maps. Here, we report a simple and reliable solution for achieving high-SNR energy-dispersive X-ray (EDX) spectroscopy spectrum images for individual atomic columns. The method is based on standard cross-correlation aided by averaging of single-column EDX maps with modifications in the reference image. It produces EDX maps with minimal specimen drift, beam drift, and scan distortions. Step-by-step procedures to determine a self-consistent reference map with a discussion on the reliability, stability, and limitations of the method are presented here.

The Use of Regularized Least Squares Minimization for the Deconvolution of SEM Images

Major advances are currently taking place in image processing algorithm development. When combined with today’s high speed computers utilizing multicore processing and very large memories the door is opened to the practical application of regularization techniques to the restoration of SEM images with improved spatial resolution. It is well recognized that an observed SEM image can be thought of as the convolution of a point spread function (psf) arising from measurement broadening and the true structure imaged. However, even with the abovementioned advances, a variety of additional limitations arise that are preventing the practical implementation of deconvolution to SEM image restoration. These factors include noise as well as a lack of detailed knowledge of the psf since no experimental technique is currently available to directly measure it with level of spatial resolution required. Furthermore, the problem is complicated by other factors such as specimen drift, contamination, the three dimensionality of specimen, signal excitation volume considerations, non-linearity in the scanning system and also nonlinear behavior in the detection chain [1].

Studies of local structural distortions in strained ultrathin BaTiO3 films using scanning transmission electron microscopy.

Ultrathin ferroelectric heterostructures (SrTiO3/BaTiO3/BaRuO3/SrRuO3) were studied by scanning transmission electron microscopy (STEM) in terms of structural distortions and atomic displacements. The TiO2-termination at the top interface of the BaTiO3 layer was changed into a BaO-termination by adding an additional BaRuO3 layer. High-angle annular dark-field (HAADF) imaging by aberration-corrected STEM revealed that an artificially introduced BaO-termination can be achieved by this interface engineering. By using fast sequential imaging and frame-by-frame drift correction, the effect of the specimen drift was significantly reduced and the signal-to-noise ratio of the HAADF images was improved. Thus, a quantitative analysis of the HAADF images was feasible, and an in-plane and out-of-plane lattice spacing of the BaTiO3 layer of 3.90 and 4.22 Å were determined. A 25 pm shift of the Ti columns from the center of the unit cell of BaTiO3 along the c-axis was observed. By spatially resolved electron energy-loss spectroscopy studies, a reduction of the crystal field splitting (CFS, ΔL3=1.93 eV) and an asymmetric broadening of the eg peak were observed in the BaTiO3 film. These results verify the presence of a ferroelectric polarization in the ultrathin BaTiO3 film.

Improving Signal to Noise in Labeled Biological Specimens Using Energy-Filtered TEM of Sections with a Drift Correction Strategy and a Direct Detection Device

Energy filtered transmission electron microscopy techniques are regularly used to build elemental maps of spatially distributed nanoparticles in materials and biological specimens. When working with thick biological sections, electron energy loss spectroscopy techniques involving core-loss electrons often require exposures exceeding several minutes to provide sufficient signal to noise. Image quality with these long exposures is often compromised by specimen drift, which results in blurring and reduced resolution. To mitigate drift artifacts, a series of short exposure images can be acquired, aligned, and merged to form a single image. For samples where the target elements have extremely low signal yields, the use of charge coupled device (CCD)-based detectors for this purpose can be problematic. At short acquisition times, the images produced by CCDs can be noisy and may contain fixed pattern artifacts that impact subsequent correlative alignment. Here we report on the use of direct electron detection devices (DDD's) to increase the signal to noise as compared with CCD's. A 3× improvement in signal is reported with a DDD versus a comparably formatted CCD, with equivalent dose on each detector. With the fast rolling-readout design of the DDD, the duty cycle provides a major benefit, as there is no dead time between successive frames.

Averaging scheme for atomic resolution off-axis electron holograms

All micrographs are limited by shot-noise, which is intrinsic to the detection process of electrons. For beam insensitive specimen this limitation can in principle easily be circumvented by prolonged exposure times. However, in the high-resolution regime several instrumental instabilities limit the applicable exposure time. Particularly in the case of off-axis holography the holograms are highly sensitive to the position and voltage of the electron-optical biprism. We present a novel reconstruction algorithm to average series of off-axis holograms while compensating for specimen drift, biprism drift, drift of biprism voltage, and drift of defocus, which all might cause problematic changes from exposure to exposure. We show an application of the algorithm utilizing also the possibilities of double biprism holography, which results in a high quality exit-wave reconstruction with 75pm resolution at a very high signal-to-noise ratio.

Special issue on multidimensional signal processing applications

This special issue onmultidimensional signal processing applications covers a broad range of methodologies and algorithms for spatially extended configurations of sensors and emitters for various purposes and applications. The spectrum of the typical wavelengths for these applications ranges from very short ones in electron transmission microsopy to visibile light, up to radio frequencies and further to sound waves in the audio range suitable for human perception. In spite of the diversity of the underlying physical principles there are similarities in the arising problems. Consequently also some of the approaches to the processing of the resulting sensor or emitter signals are shared between these applications. One common problem is the detection of drift and translation between images acquired by electron transmission microsopy and conventional cameras. Also the detection of motion in video sequences falls into this category. Another set of problems is concerned with the detection of the direction of a source of either electromagnetic or sound waves using suitable receiver arrays. But the situation may also be turned around by forming directed wave fronts from loudspeaker arrays to produce sound fields with certain spatial requirements. This set of seven papers shows thatmodernmultidimensional signal processing overcomes the traditional separation of image processing and audio processing. The presented methods and algorithms prescind from plain processing of available sensor or emitter signals. Instead the focus lies on physical processes andmathematical foundations underlying all applications. The paper A comparison between minimum variance control and other online compensation methods for specimen drift in transmission electron microscopy by A. Tejada and A.J. den Dekker presents an online method to reduce image blurring by specimen drift dur-

A comparison between minimum variance control and other online compensation methods for specimen drift in transmission electron microscopy

Transmission electron microscopes (TEMs) are the tools of choice in materials science, semiconductor, and biological research and it is expected that they will be increasingly used to autonomously perform high-volume, repetitive, nano-measurements in the near future. Thus, there is a clear need to develop automation strategies for these microscopes. In particular, an important feature in need of automation is specimen drift compensation, which is a common cause of image blurring in long-exposure TEM images, especially at high magnifications. In this paper, a systematic online approach to specimen drift compensation, called adaptive minimum variance control, is discussed in detail. The method makes use of an identified drift model, continuously updated from online drift measurements, to predict and ameliorate future drift values, significantly reducing their variance. The method's performance, measured in terms of drift variance reduction, is illustrated using both experimental and simulated data, and it is then compared with the performance of two pragmatic model-free methods: last data point prediction and linear extrapolation prediction.

Noise models and cryo-EM drift correction with a direct-electron camera

Blurring due to specimen-holder drift is a common occurrence in cryo-EM images. Cameras employing active-pixel sensors are capable of high frame rates such that a single low-dose exposure can be acquired as a series of frames. In this paper we consider the possibility of tracking and compensating for overall drift in typical single-particle specimens through the analysis of frame sequences. A problem that arises in tracking through cross-correlation of frames obtained with the DE-12 camera from Direct Electron LLC is the presence of "hot-pixel noise". This random pattern of bright pixels is highly correlated among frames. We show how a model of this noise can be employed to greatly reduce its effects. A filter function is derived that optimizes the tracking of image shifts by cross-correlation, and we demonstrate the tracking of specimen drift in typical cryo-EM specimens.

Computer simulation of electric field variations due to movements of electric charges

Simulations were carried out for the orbit of electron-induced secondary electrons around a charged microfibril of a sciatic nerve tissue. In order to set the parameters for the simulation, the shape of the microfibril was determined from a transmission electron microscopy image, while the electric potential on the surface of the charged microfibril was evaluated from a reconstructed phase image obtained with electron holography. On the other hand, the passing point and the angle of secondary electrons at the microfibril surface were determined from a reconstructed amplitude image. Eventually, simulation of orbits of secondary electrons was carried out by changing the kinetic energy of the secondary electrons. Under the given conditions, the orbit of secondary electrons with a kinetic energy of 29.6eV fits the observations. If there are thin layers of electrons, the secondary electrons do not reach the surface but they go over it due to the repulsive Coulomb force resulting in successive revolving motion around the charged microfibril. Furthermore, the electric field variation due to the movement of the electric charges resulting from the specimen drift is also discussed briefly comparing it with electron holography data.

Investigation of effect of near-fault motions on substandard bridge structures

The objective of this study was to investigate the effects of near-fault ground motions on substandard bridge columns and piers. To accomplish these goals, several large scale reinforced concrete models were constructed and tested on a shake table using near- and far-field ground motion records. Because the input earthquakes for the test models had different characteristics, three different measures were used to evaluate the effect of the input earthquake. These measures are peak shake table acceleration, spectral acceleration at the fundamental period of the test specimens, and the specimen drift ratios. For each measure, force-displacement relationships, strains, curvatures, drift ratios, and visual damage were evaluated. Results showed that regardless of the measure of input or response, the near-fault record generally led to larger strains, curvatures, and drift ratios. Furthermore, residual displacements were small compared to those for columns meeting current seismic code requirements.

Noise limit on practical electron ptychography

Ptychography is a wavelength-limited phase retrieval method that promises to provide sub-0.5Å resolution in the electron microscope, even if the lens employed (which is required only to form an illumination spot) can itself only achieve rather modest resolution. The fidelity of a practical specimen reconstruction using ptychography hinges on various experimental factors, the major one of which is the source brightness. Achieving a high degree of coherence of an electron beam requires source demagnification which reduces the electron counts reaching the detector for a given exposure time. Specimen drift, damage and contamination also limits the practical exposure time. In this paper we investigate the effect of the counting statistics required for a good quality ptychographic reconstruction

A method for correcting the effect of specimen drift on coherent diffractive imaging

Coherent diffractive imaging involves the inversion of a diffraction pattern to find the wave function at the exit-surface plane of the specimen. It is a promising technique for imaging, for example, nanoparticles with electrons and biological molecules with X-rays. If the illumination is not a plane wave of infinite extent, then a relative drift between the illumination and the object introduces errors into the diffraction pattern; an issue which is often overlooked. This may be of particular importance for applications with electron microscopes which use nanoscale probes. Here we show that beams which are uniform over a sufficiently large region can be used to pose a phase retrieval problem that is immune from specimen drift, provided suitable analysis of the diffraction data is undertaken. The method only applies to objects contained within a support that is smaller than a uniform region of the beam.

Size-effects in time-dependent mechanics in metallic MEMS

Reliability of microelectromechanical systems (MEMS) depends a.o. on time-dependent deformation such as creep and fatigue [1]. It is known from literature that this behavior is affected by size-effects: the interaction between microstructural length scales and dimensional length scales [2,3]. Not much research has focused on characterizing size-effects in time-dependent material behavior, specifically for free-standing thin films [3]. This study investigates size-effects caused by grain statistics in timedependent deformation in µm-sized free-standing aluminum cantilever beams. A numeric-experimental method is used to determine material parameters. The experiment entails applying a constant deflection to the micro-beams for a prolonged period. The deflection is achieved with 50 nm resolution via a micro-clamp. The beams are then released. Immediately the deformation evolution is recorded by acquiring surface height profiles with a confocal optical profiler. Image correlation of the full-field beam profiles is applied to correct for specimen drift and tilt. The experiment yields the tip deflection as function of time with ~3 nm precision. In the numerical part, this data is combined with a finite element model based on a standard-solid material model. In this way material parameters describing time-dependent behavior are extracted. The time constant for the deflection evolution is determined within 20%, as verified by predicting a different experiment. Figure 1 shows the model and the numeric prediction of an experiment.