High-angle annular dark-field imaging of self-assembled Ge islands on Si(0 0 1)

Low-resolution high-angle annular dark-field (HAADF) imaging is applied to the study of coherent Ge islands on a Si(0 0 1) substrate. Experimental HAADF images reveal a complicated pattern for a coherent Ge island under (0 0 1) zone axis conditions due to strain-induced interband scattering between different Bloch-wave branches. This complicated pattern varies with objective aperture size and defocus because of the effect from the depth of field. This suggests that the strain field of a coherent Ge island can be mapped out in 3 dimensions using HAADF imaging. When samples are tilted away from dynamical conditions, image contrast agrees with the predictions from atomic number variation (Z contrast). Therefore, quantitatively compositional analysis is feasible under kinematical imaging conditions when strain contrast is suppressed. Simulations using multi-beam Bloch-wave theory agree well with the experimental images on the complicated strain-induced and through-focus images.     

X射线成像波带片及制作

研究了X射线成像波带片的工作原理和制作工艺。从理论上分析了波带片的空间分辨率与最外环宽度的关系,以及波带片衍射效率与厚度和折射率的关系。利用国家同步辐射实验室发展的加工工艺,即电子束光刻技术和X射线光刻技术结合制作波带片。实验结果表明:波带片最外环宽度为150 nm,高宽比为4,基本满足高分辨X射线成像波带片的高空间分辨率、大高宽比、高精度等要求。     

Astigmatic intensity equation for electron microscopy based phase retrieval

Phase retrieval, in principle, can be performed in a transmission electron microscope (TEM) using arbitrary aberrations of electron waves; provided that the aberrations are well-characterised and known. For example, the transport of intensity equation (TIE) can be used to infer the phase from a through-focus series of images. In this work an "astigmatic intensity equation" (AIE) is considered, which relates phase gradients to intensity variations caused by TEM objective lens focus and astigmatism variations. Within the paraxial approximation, it is shown that an exact solution of the AIE for the phase can be obtained using efficient Fourier transform methods. Experimental requirements for using the AIE are the measurement of a through-focus derivative and another intensity derivative, which is taken with respect to objective lens astigmatism variation. Two quasi-experimental investigations are conducted to test the validity of the solution.     

Image deconvolution in spherical aberration-corrected high-resolution transmission electron microscopy

The method of image deconvolution developed previously for FEG high-resolution transmission electron microscope (HRTEM) without a spherical aberration (C(s)) corrector was for the first time applied to FEG HRTEM with a C(s)-corrector. The principle and the procedure of image deconvolution are briefly described. Four qualified [1 1 0] images of Si were selected from a through-focus series to perform image deconvolution. The projected potential is successfully derived from all the images, and the obtained "dumbbell" structure maps of Si [1 1 0] are in good agreement with the calculated potential map. The criterion of selecting qualified images for performing image deconvolution is indicated. The possibility of applying image deconvolution to defect study and to ab initio crystal structure determination is discussed.     

High-resolution wave field reconstruction using a through-focus series of images taken under limited electron dose

The three-dimensional Fourier filtering method and Schiske's Wiener filtering method are compared with the aim of high-resolution wave field reconstruction of an unstained deoxyribonucleic acid (DNA) molecular fiber using a through-focus series of images taken under a limited electron dose. There were some definite differences between the two reconstructed images, although the two kinds of processing are essentially equivalent except for the dimension and the filter used for processing. Through theoretical analyses together with computer simulations, the differences were proved to be primarily due to specimen drift during the experiment. Although the observed structure of the DNA molecular fiber was heavily damaged by electron beam irradiation, reconstructed images by the three-dimensional Fourier filtering method provided higher resolution information on the molecular structure even when relatively large specimen drift was included in the through-focus series. In contrast, in Schiske's Wiener filtering method, the detailed information of the structure was lost because of the drift, although the reconstructed image showed a higher signal-to-noise ratio. The three dimensional Fourier filtering method seems to be more applicable for observing radiation-sensitive materials under an extremely low electron dose, because specimen drift cannot be completely avoided.     

Caustic imaging of gallium droplets using mirror electron microscopy

We discuss a new interpretation of mirror electron microscopy (MEM) images, whereby electric field distortions caused by surface topography and/or potential variations are sufficiently large to create caustics in the image contrast. Using a ray-based trajectory method, we consider how a family of rays overlaps to create caustics in the vicinity of the imaging plane of the magnetic objective lens. Such image caustics contain useful information on the surface topography and/or potential, and can be directly related to surface features. Specifically we show how a through-focus series of MEM images can be used to extract the contact angle of a Ga droplet on a GaAs (001) surface.     

Multiple parametric nanoscale measurements with high sensitivity based on through-focus scanning optical microscopy

High-throughput through-focus scanning optical microscopy (TSOM) involves defocusing along the optical axis and capturing a series of defocus images and is useful in optical nanoscale measurement. However, TSOM is usually affected by its optical and mechanical noises. In this study, the issue of sensitivity and application in three-dimensional (3D) multiple parameter measurement of TSOM is investigated. First, a TSOM system with objective scanning and its relative simulation algorithm are proposed. Second, based upon the system and algorithm, an experiment on an isolated Au line is performed and the corresponding matching library is established. Comparing the experimental TSOM image and simulated TSOM images of the library, 3D multiple parameter results of the Au line are extracted. Third, the precision of the system is analysed through a fidelity test particular for through-focus images. According to this study, the system is robust to the optical and mechanical noises and hence could be useful in 3D multiple parametric measurement and high-volume nanomanufacturing.     

Quantitative image correction and calibration for confocal fluorescence microscopy using thin reference layers and SIPchart-based calibration procedures

The fluorescence intensity image of an axially integrated through-focus series of a thin standardized uniform fluorescent layer can be used for image intensity correction and calibration in sectioning microscopy. This intensity image is in fact available from the earlier introduced Sectioned Imaging Property (SIP) charts ( Brakenhoff et al. , 2005 ). It is shown that the integrated intensity of a z -stack from a biological sample, imaged under identical conditions as the layer, can be calibrated in terms of fluorescence layer units of the calibration layer. The imaging after such calibration becomes, as a first approximation, independent of the microscope system and imaging conditions. This is demonstrated on axially integrated images of standard fluorescent beads and standard BPAE Fluorocells. Corrections on the microscope imaging conditions include shading effects, imaging with different magnifications and objectives, and using different microscope systems. It is also shown that with the present approach the actual underlying three-dimensional (3D) fluorescence data set itself can be corrected for variations in point spread function (PSF) imaging efficiency over the imaging data cube. Realizing such calibration between imaging conditions or systems requires basically only the 2D fluorescer molecule density of the reference layers and the section distances with which the layer data are collected.     

Manufacture and tests of objective zone plates for use in a scanning transmission X-ray microscope

A method of making high resolution zone plates for use as the focusing elements in soft X-ray microscopy is briefly described. Tests carried out on these zone plates indicate a first-order diffraction efficiency of ～0.3% rather than the calculated value of ～0.9%. This indicates that the zones are not positioned as accurately as expected, a conclusion also drawn from tests at optical wavelengths on electron micrographs of the zone plates. Modifications to the manufacturing method to enable zone plates with improved imaging properties to be made are described.     

The simulation of high resolution images of amorphous thin films

Computer-simulated high resolution electron microscopy (HREM) images of white-noise weak phase objects show features similar to those observed in images of very thin films (2 nm) of amorphous silicon. Local patches of regular fringes arise as a result of the spatial-frequency filtering effect of the aperture and need not indicate the presence of microcrystalline regions. It is shown how drastically the appearance of a simulated amorphous image is influenced by the transfer function of the objective lens and by the objective aperture or equivalent damping of high-frequency components due to partial temporal and spatial coherence of the electron beam. Simulated images of through-focus series also show the same general appearance as for an amorphous silicon film.     

Computed tomography of cryogenic biological specimens based on X-ray microscopic images

Soft X-ray microscopy employs the photoelectric absorption contrast between water and protein in the 2.34-4.38 nm wavelength region to visualize protein structures down to 30 nm size without any staining methods. Due to the large depth of focus of the Fresnel zone plates used as X-ray objectives, computed tomography based on the X-ray microscopic images can be used to reconstruct the local linear absorption coefficient inside the three-dimensional specimen volume. High-resolution X-ray images require a high specimen radiation dose, and a series of images taken at different viewing angles is needed for computed tomography. Therefore, cryo microscopy is necessary to preserve the structural integrity of hydrated biological specimens during image acquisition. The cryo transmission X-ray microscope at the electron storage ring BESSY I (Berlin) was used to obtain a tilt series of images of the frozen-hydrated green alga Chlamydomonas reinhardtii. The living specimens were inserted into borosilicate glass capillaries and, in this first experiment, rapidly cooled by plunging into liquid nitrogen. The capillary specimen holders allow image acquisition over the full angular range of 180 degrees. The reconstruction shows for the first time details down to 60 nm size inside a frozen-hydrated biological specimen and conveys a clear impression of the internal structures. This technique is expected to be applicable to a wide range of biological specimens, such as the cell nucleus. It offers the possibility of imaging the three-dimensional structure of hydrated biological specimens close to their natural living state.     

The effect of mechanical vibration and drift on the reconstruction of exit waves from a through focus series of HREM images

Experimental HREM images can show a limited resolution as a result of mechanical vibration and drift. In this paper the effect of such mechanical vibrations on the accuracy of the through focus exit wave reconstruction method is investigated for different thicknesses of a test structure of La₃Ni₂B₂N₃. A through-focus series of HREM images for this structure is simulated for different kinds of mechanical vibration corresponding to an information limit g of about 7 nm⁻¹: (1) no mechanical vibration, (2) isotropic mechanical vibration, and (3) several anisotropic mechanical vibrations. From these through-focus series the reconstructed exit wave is calculated (Ultramicroscopy 64 (1996) 109). The above isotropic and anisotropic mechanical vibrations have a large effect on the reconstructed exit waves when compared with the reconstructed exit wave without mechanical vibration, i.e. the range of amplitudes and phases in a reconstructed exit wave decreases and the background intensity increases. The initial thickness and orientation can be obtained using a least-squares refinement procedure (Acta Crystallogr. A 54 (1998) 91) when there is no mechanical vibration present. In the case of isotropic or anisotropic vibration, the refined thickness and orientation are likely to give wrong results depending on the size of the vibrations and on the number of significant reflections (which is related to the size of the unit cell, the thickness and the misorientation).     

Wave field restoration using three-dimensional Fourier filtering method.

A wave field restoration method in transmission electron microscopy (TEM) was mathematically derived based on a three-dimensional (3D) image formation theory. Wave field restoration using this method together with spherical aberration correction was experimentally confirmed in through-focus images of amorphous tungsten thin film, and the resolution of the reconstructed phase image was successfully improved from the Scherzer resolution limit to the information limit. In an application of this method to a crystalline sample, the surface structure of Au(110) was observed in a profile-imaging mode. The processed phase image showed quantitatively the atomic relaxation of the topmost layer.     

大高宽比硬X射线波带片制作及聚焦测试

为得到同步辐射光源硬X射线波段(2keV)需要的高宽比高分辨率波带片,本文利用高加速电压(100kV)电子束光刻配合Si3N4镂空薄膜直写来减少背散射的方法,对硬X射线波带片制作技术进行了蒙特卡洛模拟和电子束光刻实验。模拟结果显示:Si_3N_4镂空薄膜衬底可以有效降低电子在抗蚀剂中传播时的背散射,进而改善高密度大高宽比容易引起的结构倒塌和粘连问题。通过调整电子束的曝光剂量,在500nm厚的镂空Si_3N_4薄膜衬底上制备出最外环宽度为150nm、金吸收体的厚度为1.6μm,高宽比大于10的硬X射线波带片。同时,引入随机支撑点结构,实现了波带片结构自支撑,提高了大高宽比波带片的稳定性。将利用该工艺制作的波带片在北京同步辐射装置X射线成像4W1A束线8keV能量下进行了聚焦测试,得到清晰的聚焦结果。     

Evaluating sharpness functions for automated scanning electron microscopy

Fast and reliable autofocus techniques are an important topic for automated scanning electron microscopy. In this paper, different autofocus techniques are discussed and applied to a variety of experimental through-focus series of scanning electron microscopy images with different geometries. The procedure of quality evaluation is described, and for a variety of scanning electron microscope samples it is demonstrated that techniques based on image derivatives and Fourier transforms are in general better than statistical, intensity and histogram-based techniques. Further, it is shown that varying of an extra parameter can dramatically increase quality of an autofocus technique.     

Atomic imaging in aberration-corrected high-resolution transmission electron microscopy

In recent years, the successful implementation of a spherical-aberration corrector in a Philips CM 200 FEG ST microscope achieved by Haider et al. has attracted a great deal of attention. However, thus far extensive applications of this novel high-resolution transmission electron microscope (HRTEM) to materials research have been hampered by the problems concerning optimum imaging conditions and image interpretation. In this paper, we present our points of view concerning atomic imaging in an aberration-corrected HRTEM. Since atomic resolution images can also be obtained with other techniques such as through-focus exit-wave function reconstruction (TF-EWR), we have to emphasis that the strength of the aberration-corrected HRTEM particularly lies on its ability to resolve the atomic structure in real time. However, for this purpose it is mandatory that the image contrast be related in a one-to-one function with the projected structure of the object. We analyzed the atomic imaging conditions in much detail and we come to the following conclusion: this novel facility is no doubt a powerful and advanced HRTEM instrument in achieving atomic images with its highest resolution (information limit). We furthermore demonstrate that the combination of the new microscope and TF-EWR will yield optimal results.     

Imaging and analysis of nanowires

We used vapor-liquid-solid (VLS) methods to synthesize discrete single-element semiconductor nanowires and multicomposition nanowire heterostructures, and then characterized their structure and composition using high-resolution electron microscopy (HRTEM) and analytical electron microscopy techniques. Imaging nanowires requires the modification of the established HRTEM imaging procedures for bulk material to take into consideration the effects of finite nanowire width and thickness. We show that high-resolution atomic structure images of nanowires less than 6 nm in thickness have lattice "streaking" due to the finite crystal lattice in two dimensions of the nanowire structure. Diffraction pattern analysis of nanowires must also consider the effects of a finite structure producing a large reciprocal space function, and we demonstrate that the classically forbidden 1/3 [422] reflections are present in the [111] zone axis orientation of silicon nanowires due to the finite thickness and lattice plane edge effects that allow incomplete diffracted beam cancellation. If the operating conditions are not carefully considered, we found that HRTEM image delocalization becomes apparent when employing a field emission transmission electron microscope (TEM) to image nanowires and such effects have been shown to produce images of the silicon lattice structure outside of the nanowire itself. We show that pseudo low-dose imaging methods are effective in reducing nanowire structure degradation caused by electron beam irradiation. We also show that scanning TEM (STEM) with energy dispersive X-ray microanalysis (EDS) is critical in the examination of multicomponent nanowire heterostructures.     

Quantitative elemental X-ray imaging of frozen-hydrated biological samples

It is shown that quantitative X-ray imaging of planed, frozen-hydrated, biological bulk samples that have not been etched is possible. X-ray imaging represents a better alternative to static beam (selected area) analysis of fractured frozen-hydrated samples. This procedure avoids the undesirable necessity of etching planed frozen-hydrated samples to provide an interpretable electron image. Qualitative oxygen and carbon X-ray images, which can be acquired in a short time, can be used for distinguishing morphological features and remove the requirement for electron images. In test samples of frozen-hydrated albumin, containing salts, analyses by X-ray images compared well with static beam (selected area) analyses from the same samples. An example of an analysis of frozen-hydrated insect Malpighian tubules is given in which the response to ouabain treatment was analysed. In this example X-ray imaging showed that ouabain resulted in a significant increase in cytoplasmic and luminal Na and a significant decrease in cytoplasmic and luminal K. X-ray imaging also showed that there was a significant increase in cellular water content. The presence of a potassium gradient in soybean root nodules was also demonstrated. The use of standard deviation images for processing low count images increases analytical precision but results in underestimates of the true concentrations.     

基于稀疏处理的多能X射线分离成像

利用独立成分分析(Independent Component Analysis,ICA)并结合多能X射线图像的丰富信息可以将二维X射线图像中重叠目标分离成像,但是海量的图像数量,以及高像素数的要求均会使内存占有量和计算速度面临挑战,因此本研究将压缩感知(Compressed Sensing,CS)与ICA相结合进行分离成像,以提高计算速度和分离成像性能。研究过程中,首先根据被拍摄物体的物质组成确定拍摄多能X射线图像数量,并选取CS技术中K均值奇异值分解(K-means SingularValue Decomposition,K-SVD)稀疏基将多能X射线图像进行稀疏表示,然后利用ICA将此稀疏表示进行盲源分离得到独立源,最后采用正交匹配追踪算法(Orthogonal Matching Pursuit,OMP)将独立源进行重构实现分离成像。研究结果表明:采用ICACS技术比仅采用ICA进行目标分离成像的运行时间减少了46.14s(23.3%)、内存占有率降低了21%、重构图像峰值信噪比(Peak Signal to Noise Ratio,PSNR)提高了2.665dB、边缘梯度提高了0.001、信息熵提高了0.09。