Quantitative, in situ visualization of intracellular insulin vesicles in pancreatic beta cells.

Characterizing relationships between Zn2+, insulin, and insulin vesicles is of vital importance to the study of pancreatic beta cells. However, the precise content of Zn2+ and the specific location of insulin inside insulin vesicles are not clear, which hinders a thorough understanding of the insulin secretion process and diseases caused by blood sugar dysregulation. Here, we demonstrated the colocalization of Zn2+ and insulin in both single extracellular insulin vesicles and pancreatic beta cells by using an X-ray scanning coherent diffraction imaging (ptychography) technique. We also analyzed the elemental Zn2+ and Ca2+ contents of insulin vesicles using electron microscopy and energy dispersive spectroscopy (EDS) mapping. We found that the presence of Zn2+ is an important characteristic that can be used to distinguish insulin vesicles from other types of vesicles in pancreatic beta cells and that the content of Zn2+ is proportional to the size of insulin vesicles. By using dual-energy contrast X-ray microscopy and scanning transmission X-ray microscopy (STXM) image stacks, we observed that insulin accumulates in the off-center position of extracellular insulin vesicles. Furthermore, the spatial distribution of insulin vesicles and their colocalization with other organelles inside pancreatic beta cells were demonstrated using three-dimensional (3D) imaging by combining X-ray ptychography and an equally sloped tomography (EST) algorithm. This study describes a powerful method to univocally describe the location and quantitative analysis of intracellular insulin, which will be of great significance to the study of diabetes and other blood sugar diseases.

Design and first-round commissioning result of the SASE beamline at the Shanghai Soft X-ray FEL facility.

The Shanghai Soft X-ray Free-Electron Laser (SXFEL) is the first X-ray free-electron laser facility in China. The SASE beamline, which consists of a pink-beam branch and a mono-beam branch, is one of the two beamlines in the Phase-I construction. The pink-beam branch opened for users in 2023 after successful first-round beamline commissioning. In this paper, the design of the beamline is presented and the performance of the pink-beam branch is reported. The measured energy-resolving power of the online spectrometer is over 6000 @ 400 eV. The focusing spot size of the pink beam is less than 3 µm in both the horizontal and vertical at the endstation.

Fabrication of large-area photonic crystal-modified X-ray scintillator imager for optical coding imaging.

The limited pattern area of periodic nanostructures limits the development of practical devices. This study introduces an X-ray interference lithography (XIL) stitching technique to fabricate a large-area (1.5 cm × 1.5 cm) two-dimensional photonic crystal (PhC) on the YAG: Ce scintillator, which functions as an encoder in a high numerical aperture optical encoding imaging system to effectively capture high-frequency information. An X-ray imaging experiment revealed a substantial 7.64 dB improvement in the signal-to-noise ratio (SNR) across a large field of view (2.6 mm × 2.6 mm) and achieved comparable or superior image quality with half the exposure dose. These findings have significant implications for advancing practical applications of X-ray imaging.

Hydrogen Nanobubbles Generated In Situ from Nanoscale Zerovalent Iron with Water to Further Enhance Selenite Sequestration.

Gas nanobubbles used for water treatment and recovery give rise to great concern for their unique advantages of less byproducts, higher efficiency, and environmental friendliness. Nanoscale zerovalent iron (nZVI), which has also been widely explored in the field of environmental remediation, can generate gas hydrogen by direct reaction with water. Whether nanoscale hydrogen bubbles can be produced to enhance the pollution removal of the nZVI system is one significant concern involved. Herein, we report direct observations of in situ generation of hydrogen nanobubbles (HNBs) from nZVI in water. More importantly, the formed HNBs can enhance indeed the reduction of Se(IV) beyond the chemical reduction ascribed to Fe(0), especially in the anaerobic environment. The possible mechanism is that HNBs enhance the reducibility of the system and promote electron transport in the solution. This study demonstrates a unique function of HNBs combined with nZVI for the pollutant removal and a new approach for in situ HNB generation for potential applications in the fields of in situ remediation agriculture, biotechnology, medical treatment, health, etc.

Mutual optical intensity propagation through non-ideal two-dimensional mirrors.

The mutual optical intensity (MOI) model is a partially coherent radiation propagation tool that can sequentially simulate beamline optics and provide beam intensity, local degree of coherence and phase distribution at any location along a beamline. This paper extends the MOI model to non-ideal two-dimensional (2D) optical systems, such as ellipsoidal and toroidal mirrors with 2D figure errors. Simulation results show that one can tune the trade-off between calculation efficiency and accuracy by varying the number of wavefront elements. The focal spot size of an ellipsoidal mirror calculated with 100 × 100 elements gives less than 0.4% deviation from that with 250 × 250 elements, and the computation speed is nearly two orders of magnitude faster. Effects of figure errors on 2D focusing are also demonstrated for a non-ideal ellipsoidal mirror and by comparing the toroidal and ellipsoidal mirrors. Finally, the MOI model is benchmarked against the multi-electron Synchrotron Radiation Workshop (SRW) code showing the model's high accuracy.

Partially coherent light propagation through a kinoform lens.

Combining wave optics propagation and geometric ray tracing, the mutual optical intensity (MOI) model is extended to quantitatively simulate the propagation of partially coherent light through a kinoform lens at high speed. The MOI model can provide both a high accuracy and a high efficiency simulation. The intensity and coherence degree distributions at the focal plane are calculated using the MOI model. It is beneficial to improve the focusing capability of the kinoform lens by reducing the coherence or increasing the number of lens steps. In addition, increasing the number of steps is also beneficial to increase the photon flux and reduce the depth of focus.

High-efficiency focusing and imaging by dielectric kinoform zone plate lenses with soft X-rays.

With fast advances in enhancing the focusing/imaging resolution of Fresnel zone plate lenses toward sub-10 nm, low diffraction efficiency in connection with their rectangular zone shape still remains a big issue in both soft and hard X-ray microscopy. In hard X-ray optics, encouraging progress has recently been reported in our earlier attempts of high focusing efficiency by 3D kinoform shaped metallic zone plates, formed by greyscale electron beam lithography. This paper addresses our efforts towards high focusing/imaging efficiency by developing a novel dielectric kinoform zone plate lens for soft X-rays. The effects of the zone materials and zone shapes on the focusing/imaging quality were first theoretically investigated by a modified thin-grating-approximation method, revealing superior efficiencies of dielectric kinoform zone plates over rectangular ones in metals. Optical characterizations of replicated dielectric kinoform zone plates by greyscale electron beam lithography demonstrate a focusing efficiency of 15.5% with a resolution of 110 nm in the water window of X-rays. Apart from high efficiency, the novel kinoform zone plate lenses developed in this work exhibit significant advantages over conventional zone plates, i.e. simplified process, low cost and no need for a beamstop.

A compound Kinoform/Fresnel zone plate lens with 15 nm resolution and high efficiency in soft x-ray.

X-ray microscope as an important nanoprobing tool plays a prevailing role in nano-inspections of materials. Despite the fast advances of high resolution focusing/imaging reported, the efficiency of existing high-resolution zone plates is mostly around 5% in soft x-ray and rapidly goes down to 1%-2% when the resolution approaches 10 nm. It is well known that the rectangular zone shape, beamstop, limited height/width ratios, material absorption of light and structural defects are likely responsible for the limited efficiency. Although zone plates with Kinoform profile are supposed to be efficient, progress for achieving both high resolution (<30 nm) and high efficiency (>5%) have hardly been addressed in soft x-ray. In this work, we propose a compound Kinoform/Fresnel zone plate (CKZP) by combing a dielectric Kinoform zone plate with a 15 nm resolution zone plate. Greyscale electron beam lithography was applied to form the 3D Kinoform zone plate and atomic layer deposition was carried out to form the binary zone plate. Optical characterizations demonstrated 15 nm resolution focusing/imaging with over 7.8% efficiency in soft x-ray. The origin of the efficiency improvement behind the proposed compound lens is theoretically analyzed and discussed.

Initial probe function construction in ptychography based on zone-plate optics.

X-ray ptychography is a popular variant of coherent diffraction imaging that offers ultrahigh resolution for extended samples. In x-ray ptychography instruments, the Fresnel zone-plate (FZP) is the most commonly used optical probe system for both soft x-ray and hard x-ray. In FZP-based ptychography with a highly curved defocus probe wavefront, the reconstructed image quality can be significantly impacted by the initial probe function form, necessitating the construction of a suitable initial probe for successful reconstruction. To investigate the effects of initial probe forms on FZP-based ptychography reconstruction, we constructed four single-mode initial probe models (IPMs) and three multi-mode IPMs in this study, and systematically compared their corresponding simulated and experimental reconstructions. The results show that the Fresnel IPM, spherical IPM, and Fresnel-based multi-mode IPMs can result in successful reconstructions for both near-focus and defocus cases, while random IPMs and constant IPMs work only for near-focus cases. Consequently, for FZP-based ptychography, the elaborately constructed IPMs that closely resemble real probes in wavefront phase form are more advantageous than natural IPMs such as the random or constant model. Furthermore, these IPMs with high phase similarity to the high-curvature large-sized probe adopted in experiments can help greatly improve ptychography experiment efficiency and decrease radiation damage to samples.

In Situ X-ray Absorption Spectroscopy of Metal/Nitrogen-doped Carbons in Oxygen Electrocatalysis.

Metal/nitrogen-doped carbons (M-N-C) are promising candidates as oxygen electrocatalysts due to their low cost, tunable catalytic activity and selectivity, and well-dispersed morphologies. To improve the electrocatalytic performance of such systems, it is critical to gain a detailed understanding of their structure and properties through advanced characterization. In situ X-ray absorption spectroscopy (XAS) serves as a powerful tool to probe both the active sites and structural evolution of catalytic materials under reaction conditions. In this review, we firstly provide an overview of the fundamental concepts of XAS and then comprehensively review the setup and application of in situ XAS, introducing electrochemical XAS cells, experimental methods, as well as primary functions on catalytic applications. The active sites and the structural evolution of M-N-C catalysts caused by the interplay with electric fields, electrolytes and reactants/intermediates during the oxygen evolution reaction and the oxygen reduction reaction are subsequently discussed in detail. Finally, major challenges and future opportunities in this exciting field are highlighted.

X-ray propagation through a kinoform lens.

Combining geometric ray tracing and wave optics propagation, a new simulation model named LWF is established to calculate the full coherent X-ray propagation through a kinoform lens. The LWF model is used to analyze the X-ray propagation through long and short kinoform lenses and calculate the intensity distribution at the focal plane. When the aperture is large, the focal spot for the long kinoform lens is smaller than that for the short kinoform lens. Due to the use of the geometric ray-tracing method to calculate the beam propagation inside the kinoform lens, the LWF model takes a low number of transversal wavefront segments, i.e. a short time, to achieve high accuracy. The simulation times for the one-dimensional and two-dimensional LWF models are 0.025 s and 5.3 s, respectively, with a calculation error of less than 0.5%. The high efficiency and high accuracy make the LWF model a strong tool in designing kinoform lenses.

Trapezoid-kinoform zone plate lens - a solution for efficient focusing in hard X-ray optics.

X-ray microscopes are powerful tools in the nano-inspection of materials owing to their ultra-high resolution at the molecular level. However, the focusing efficiency of binary zone plate lenses as key components in such probes is merely 5% in practice, hindering their application in advanced scientific research. Although kinoform zone plate lenses are in principle supposed to possess high efficiency beyond binary ones, little progress has been reported so far due to the shortage of both a theoretical calculation approach and greyscale lithography for generating fine three-dimensional (3D) kinoform zones of the lenses. This paper reports our theoretical work for a modified beam propagation method to compute the focusing performance and state-of-the-art 3D greyscale electron beam lithography for kinoform zone plate lenses. Three different zone shapes - binary, kinoform and top-flat kinoform (nicknamed the trapezoid-kinoform) - were compared both theoretically and experimentally. Theoretical calculations suggest, for the first time, that the trapezoid-kinoform zone plate gives rise to the highest focusing efficiency among the three lenses, which was proved by optical characterization of the fabricated lens with hard X-rays. As high as 40% of the focusing efficiency by Au trapezoid-kinoform lenses with resolution of 250 nm at 8 keV has been achieved, which is two times higher than that of binary zone plate lenses. The origin of the enhanced efficiency in the trapezoid-kinoform zone plate lens was explained by the joint contributions from both the refraction through the kinoform slope and the diffraction through the top flat part of the trapezoid-kinoform zone plate. Such a breakthrough in focusing efficiency sheds light on the further development of X-ray lenses with both high resolution and high efficiency.

Nanoscale Detection of Subcellular Nanoparticles by X-Ray Diffraction Imaging for Precise Quantitative Analysis of Whole Cancer Cells.

Nanoparticles show great potential for drug delivery systems in cancer treatment and diagnosis, which mainly rely on the interaction between nanoparticles and living cells. However, there is still a lack of accurate and large field-of-view imaging techniques to reveal the aggregation and distribution behavior of nanoparticles in whole cancer cells without being destroyed. Here, we demonstrated quantitative imaging of unstained and intact mouse breast cancer cells (4T1) containing 50 nm gold nanoparticles (Au@citrate NPs) using an X-ray scanning coherent diffraction imaging (ptychography) technique in a large field-of-view. A two-dimensional spatial resolution of 17 nm was achieved on the 4T1 cell. We combine X-ray ptychography and equally sloped tomography (EST) to perform three-dimensional structural mapping, distribution, and aggregation behavior of Au@citrate NPs in cancer cells. By taking full advantage of the large field-of-view, high-resolution, and quantitative imaging technique, the single intracellular Au@citrate NPs are observed and the amount of Au@citrate NPs in aggregations can be accurately quantified. In addition, the morphological changes of lysosomes containing Au@citrate NPs can be observed in the high-contrast mass density images. This study provides an approach for exploring quantitative analysis and physiological delivery of nanomaterials in intact cancer cells at nanoscale resolution, which may greatly benefit the interdisciplinary research of material science, nanomedicine, and nanotoxicology.

A bidirectional scanning method for scanning transmission X-ray microscopy.

Scanning mode is a key factor for the comprehensive performance, including imaging efficiency, of scanning transmission X-ray microscopy (STXM). Herein is presented a bidirectional scanning method designed for STXM with an S-shaped moving track. In this method, artificially designed ramp waves are generated by a piezo-stage controller to control the two-dimensional scanning of the sample. The sample position information is measured using laser interferometric sensors and sent to a field-programmable gate array (FPGA) board which also acquires the X-ray signals simultaneously from the detector. Since the data recorded by the FPGA contain the real position of each scanned point, the influence of the backlash caused by the back-turning movement on the STXM image can be eliminated. By employing an adapted post-processing program, a re-meshed high-resolution STXM image can be obtained. This S-track bidirectional scanning method in fly-scan mode has been implemented on the STXM endstation at the Shanghai Synchrotron Radiation Facility (SSRF), and successfully resolved the ∼30 nm interval between the innermost strips of a Siemens star. This work removes the limitation on bidirectional scanning caused by motor backlash and vibration, and significantly improves the efficiency of STXM experiments.

Three-dimensional fast elemental mapping by soft X-ray dual-energy focal stacks imaging.

The three-dimensional (3D) dual-energy focal stacks (FS) imaging method has been developed to quickly obtain the spatial distribution of an element of interest in a sample; it is a combination of the 3D FS imaging method and two-dimensional (2D) dual-energy contrast imaging based on scanning transmission soft X-ray microscopy (STXM). A simulation was firstly performed to verify the feasibility of the 3D elemental reconstruction method. Then, a sample of composite nanofibers, polystyrene doped with ferric acetylacetonate [Fe(acac)3], was further investigated to quickly reveal the spatial distribution of Fe(acac)3 in the sample. Furthermore, the data acquisition time was less than that for STXM nanotomography under similar resolution conditions and did not require any complicated sample preparation. The novel approach of 3D dual-energy FS imaging, which allows fast 3D elemental mapping, is expected to provide invaluable information for biomedicine and materials science.

Analysis of partially coherent light propagation through the soft X-ray interference lithography beamline at SSRF.

The mutual optical intensity (MOI) model is extended to the simulation of the interference pattern produced by extreme ultraviolet lithography with partially coherent light. The partially coherent X-ray propagation through the BL08U1B beamline at Shanghai Synchrotron Radiation Facility is analysed using the MOI model and SRW (Synchrotron Radiation Workshop) method. The fringe intensity at the exposure area is not uniform but has similar envelope lines to Fresnel diffraction, which is explained by the diffraction from the finite grating modelled as a single aperture. By balancing the slit size and photon stop size, the fringe visibility, photon flux and intensity slope can be optimized. Further analysis shows that the effect of pink light on the aerial images is negligible, whereas the third-harmonic light should be considered to obtain a balance between high fringe visibility and high flux. Two grating interference exposure experiments were performed in the BL08U1B beamline. The aerial image depth showed that the polymethyl methacrylate photoresist depth was determined by the X-ray coherence properties.

Virtual depth-scan multi-slice ptychography for improved three-dimensional imaging.

Multi-slice ptychography (MSP) is a fast three-dimensional ptychography technology developed on the basis of conventional ptychography. With this method, three-dimensional imaging can be achieved without rotating the sample. The prototype multi-slice algorithm can only reconstruct three-dimensional samples with a limited number of slices, which greatly limits the depth range and resolution of sample imaging. Here we reported a virtual depth-scan scheme of MSP in which a thick sample is scanned virtually in the depth direction across its whole thickness range within the reconstruction process, thereby eliminating the restriction on slice number and potentially improving the depth resolution of MSP. This new approach also improves the flexibility of multi-slice ptychography. Both the simulation and experimental results validate the feasibility of our new approach.

Hybrid application of laser-focused atomic deposition and extreme ultraviolet interference lithography methods for manufacturing of self-traceable nanogratings.

A novel hybrid method that combines the laser-focused atomic deposition (LFAD) and extreme ultraviolet (EUV) interference lithography has been introduced. The Cr grating manufactured by LFAD has advantages of excellent uniformity, low line edge roughness and its pitch value determined directly by nature constants (i.e. self-traceable). To further enhance the density of the Cr grating, the EUV interference lithography with 13.4 nm wavelength was employed, which replicated the master Cr grating onto a Si wafer with its pitch reduced to half. In order to verify the performance of the gratings manufactured by this novel method, both mask grating (Cr grating) and replicated grating (silicon grating) were calibrated by the metrological large range scanning probe microscope (Met.LR-SPM) at Physikalisch-Technische Bundesanstalt (PTB). The calibrated results show that both gratings have excellent short-term and long-term uniformity: (i) the calibrated position deviation (i.e. nonlinearity) of the grating is below ±1 nm; (ii) the deviation of mean pitch values of 6 randomly selected measurement locations is below 0.003 nm. In addition, the mean pitch value of the Cr grating is calibrated as 212.781 ± 0.008 nm (k = 2). It well agrees with its theoretical value of 212.7787 ± 0.0049 nm, confirming the self-traceability of the manufactured grating by the LFAD. The mean pitch value of the Si grating is calibrated as 106.460 ± 0.012 nm (k = 2). It corresponds to the shrinking factor of 0.500 33 of the applied EUV interference lithographic technique. This factor is very close to its theoretical value of 0.5. The uniform, self-traceable gratings fabricated using this novel approach can be well applied as reference materials in calibrating, e.g. the magnification and uniformity of almost all kinds of high resolution microscopes for nanotechnology.

Ultrahigh Density of Gas Molecules Confined in Surface Nanobubbles in Ambient Water.

To understand the unexpected and puzzling long-term stability of nanoscale gas bubbles, it is crucial to probe their nature and intrinsic properties. We report herein synchrotron-based scanning transmission X-ray microscopy (STXM) evidence of highly condensed oxygen gas molecules trapped as surface nanobubbles. Remarkably, the analysis of absorption spectra of a single nanobubble revealed that the oxygen density inside was 1-2 orders of magnitude higher than that in atmospheric pressure, and these bubbles were found in a highly saturated liquid environment with the estimated oxygen concentration to be hundreds of times higher than the known oxygen solubility in equilibrium. Molecular dynamics simulations were performed to investigate the stability of surface nanobubbles on a heterogeneous substrate in gas-oversaturated water. These results indicated that gas molecules within confinement such as the nanobubbles could maintain a dense state instead of the ideal gas state, as long as their surrounding liquid is supersaturated. Our findings should help explain the surprisingly long lifetime of the nanobubbles and shed light on nanoscale gas aggregation behaviors.

In Situ Formation of Hierarchical Bismuth Nanodots/Graphene Nanoarchitectures for Ultrahigh-Rate and Durable Potassium-Ion Storage.

Metallic bismuth (Bi) has been widely explored as remarkable anode material in alkali-ion batteries due to its high gravimetric/volumetric capacity. However, the huge volume expansion up to ≈406% from Bi to full potassiation phase K3 Bi, inducing the slow kinetics and poor cycling stability, hinders its implementation in potassium-ion batteries (PIBs). Here, facile strategy is developed to synthesize hierarchical bismuth nanodots/graphene (BiND/G) composites with ultrahigh-rate and durable potassium ion storage derived from an in situ spontaneous reduction of sodium bismuthate/graphene composites. The in situ formed ultrafine BiND (≈3 nm) confined in graphene layers can not only effectively accommodate the volume change during the alloying/dealloying process but can also provide high-speed channels for ionic transport to the highly active BiND. The BiND/G electrode provides a superior rate capability of 200 mA h g-1 at 10 A g-1 and an impressive reversible capacity of 213 mA h g-1 at 5 A g-1 after 500 cycles with almost no capacity decay. An operando synchrotron radiation-based X-ray diffraction reveals distinctively sharp multiphase transitions, suggesting its underlying operation mechanisms and superiority in potassium ion storage application.