Abstract:Microscopy with extreme ultraviolet (EUV) light can provide many advantages over optical, hard x-ray or electron-based techniques. However, traditional EUV sources and optics have large disadvantages of scale and cost. Here, we demonstrate the use of a laboratory-scale, coherent EUV source to image biological samples—mouse hippocampal neurons—providing quantitative phase and amplitude transmission information with a lateral resolution of 80 nm and an axial sensitivity of ~1 nm. A comparison with fluorescence i… Show more
“…It allows the reconstruction of both the phase and the amplitude of an object by reverse calculating it from the reciprocal space data. The basic principle of this technique was proposed more than 50 years ago (Hoppe, 1969) and it has been widely used in X-ray (Pfeiffer, 2018) and extreme ultraviolet (Seaberg et al, 2014;Odstrcil et al, 2016;Baksh et al, 2020) studies in the past decades (Rodenburg & Maiden, 2019). Only recently, catalyzed by the introduction of fast direct electron counting pixelated detectors (MacLaren et al, 2020, Schayck et al, 2020, has cryogenic electron ptychography started to receive more attention.…”
Imaging of biomolecules by ionizing radiation, such as electrons, causes radiation damage which introduces structural and compositional changes of the specimen. The total number of high-energy electrons per surface area that can be used for imaging in cryogenic electron microscopy (cryo-EM) is severely restricted due to radiation damage, resulting in low signal-to-noise ratios (SNR). High resolution details are dampened by the transfer function of the microscope and detector, and are the first to be lost as radiation damage alters the individual molecules which are presumed to be identical during averaging. As a consequence, radiation damage puts a limit on the particle size and sample heterogeneity with which electron microscopy (EM) can deal. Since a transmission EM (TEM) image is formed from the scattering process of the electron by the specimen interaction potential, radiation damage is inevitable. However, we can aim to maximize the information transfer for a given dose and increase the SNR by finding alternatives to the conventional phase-contrast cryo-EM techniques. Here some alternative transmission electron microscopy techniques are reviewed, including phase plate, multi-pass transmission electron microscopy, off-axis holography, ptychography and a quantum sorter. Their prospects for providing more or complementary structural information within the limited lifetime of the sample are discussed.
“…It allows the reconstruction of both the phase and the amplitude of an object by reverse calculating it from the reciprocal space data. The basic principle of this technique was proposed more than 50 years ago (Hoppe, 1969) and it has been widely used in X-ray (Pfeiffer, 2018) and extreme ultraviolet (Seaberg et al, 2014;Odstrcil et al, 2016;Baksh et al, 2020) studies in the past decades (Rodenburg & Maiden, 2019). Only recently, catalyzed by the introduction of fast direct electron counting pixelated detectors (MacLaren et al, 2020, Schayck et al, 2020, has cryogenic electron ptychography started to receive more attention.…”
Imaging of biomolecules by ionizing radiation, such as electrons, causes radiation damage which introduces structural and compositional changes of the specimen. The total number of high-energy electrons per surface area that can be used for imaging in cryogenic electron microscopy (cryo-EM) is severely restricted due to radiation damage, resulting in low signal-to-noise ratios (SNR). High resolution details are dampened by the transfer function of the microscope and detector, and are the first to be lost as radiation damage alters the individual molecules which are presumed to be identical during averaging. As a consequence, radiation damage puts a limit on the particle size and sample heterogeneity with which electron microscopy (EM) can deal. Since a transmission EM (TEM) image is formed from the scattering process of the electron by the specimen interaction potential, radiation damage is inevitable. However, we can aim to maximize the information transfer for a given dose and increase the SNR by finding alternatives to the conventional phase-contrast cryo-EM techniques. Here some alternative transmission electron microscopy techniques are reviewed, including phase plate, multi-pass transmission electron microscopy, off-axis holography, ptychography and a quantum sorter. Their prospects for providing more or complementary structural information within the limited lifetime of the sample are discussed.
“…Ptychographical iterative engine (PIE) has attracted a great deal of attention in various research fields, such as x-ray imaging [1], electron imaging [2], bio-and semiconductorimaging [3][4][5], and terahertz imaging [6]. In the PIE's configuration, the spot on the object surface overlaps substantially with its neighboring spots, which can increase the robustness and remove the need for a prior knowledge of the object and probe [7,8].…”
Ptychographical iterative engine (PIE) is an attractive modality of phase retrieval that can provide the quantitative phase of the sample and extend the field of view. For a large sample, a great deal of images with a large dataset are recorded, which will result in high requirements for the computing power and increase the calculation burden. Here, we propose a pixel binning strategy to improve the computational efficiency and reduce the calculation time of PIE. In this method, the recorded image chosen as the amplitude constraint in reconstruction algorithm is compressed by merging the amplitude values of the adjacent positions into one value by the linear superposition, and the compressed pattern is set as a new amplitude constraint to reconstruct the compressed object and probe with low pixel resolution. Then, the compressed values are extended by the interpolation method. The improvement in the computational efficiency at the point lies in the fact that it takes less time to do the free-space diffraction propagation calculation for small-size images. Experiments demonstrate that the proposed method behaves good performance with high computational efficiency. The proposed approach would be helpful for large-scale imaging with high computational efficiency.
“…Nevertheless, ptychography with table-top HHG sources 23 is arguably in its infancy. Despite recent highlights in EUV ptychography, such as sub-wavelength resolution on periodic samples 24 , bioimaging of hippocampal neurons 25 as well as phase-sensitive reflectometry 26 , additional element-specificity is needed to meet the demands from the semiconductor industry and harness the prominent contrast mechanisms in silicon-based environments. Fig.…”
Microscopy with extreme ultraviolet (EUV) radiation holds promise for high-resolution imaging with excellent material contrast, due to the short wavelength and numerous element-specific absorption edges available in this spectral range. At the same time, EUV radiation has significantly larger penetration depths than electrons. It thus enables a nano-scale view into complex three-dimensional structures that are important for material science, semiconductor metrology, and next-generation nano-devices. Here, we present high-resolution and material-specific microscopy at 13.5 nm wavelength. We combine a highly stable, high photon-flux, table-top EUV source with an interferometrically stabilized ptychography setup. By utilizing structured EUV illumination, we overcome the limitations of conventional EUV focusing optics and demonstrate high-resolution microscopy at a half-pitch lateral resolution of 16 nm. Moreover, we propose mixed-state orthogonal probe relaxation ptychography, enabling robust phase-contrast imaging over wide fields of view and long acquisition times. In this way, the complex transmission of an integrated circuit is precisely reconstructed, allowing for the classification of the material composition of mesoscopic semiconductor systems.
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