Focused Ion Beam Scanning Electron Microscopy Research Articles

Volume microscopic analysis of membrane contact sites in mouse kidney renal proximal tubule epithelial cells

Subcellular organelles communicate with one another and with the plasma membrane (PM). A wide variety of membrane contact sites (MCS) tether subcellular structures to one another and mediate inter-organelle communication and exchange. The nature, distribution and function of membrane contact sites within the segment-specific architecture of renal epithelial cells has only just begun to be elucidated. We have used advanced imaging techniques, including Focused Ion Beam Scanning Electron Microscopy (FIB-SEM) and machine learning-based segmentation, to create a comprehensive, high resolution, 3D and quantifiable volume microscopic atlas of the ultrastructure of proximal tubule epithelial cells of the mouse kidney. We applied this data set to identify and quantify MCS between the endoplasmic reticulum and mitochondria and the PM in these cells. FIB-SEM was performed on a 50 μm by 50 μm by 44 μm block of tissue prepared from the renal cortex of a perfusion fixed 12-week-old male mouse. OsO4-stained tissue was embedded in Durcupan and a 3D X-ray tomogram of the block was used to select a volume of interest that included extended lengths of multiple proximal tubule (PT). The volume was imaged by a customized enhanced FIB-SEM system. The X-Y scanning resolution was set to 4-nm, and 4-nm FIB milling was used to expose surfaces for successive imaging. Generation of the complete image stack required ~11,000 cycles of scanning/FIB ablation over ~4 weeks of uninterrupted imaging. Segmentation of the resultant image set was performed with Zeiss Arivis Pro technology. The boundary box for reconstruction had the dimensions of 29.2 μm x 24.8 μm x 8 μm with a volume of 5793.280 μm3 comprising of 2000 consecutive images at an isotropic voxel size of 4 nm. Deep learning segmentation teased apart 10 cells with distinct boundaries in the PT structure. The volume of the cells in our clipped volume ranged from 8.79 μm3 to 1438.53 μm3 with a median of 45.20 μm3. In one representative proximal tubule cell the volume of mitochondria makes up 84.91 μm3 (31.62%) of the cell volume and the volume of Endoplasmic Reticulum (ER) makes up 23.92 μm3 (8.91%) of the cell volume. The 3D reconstruction and quantitative analysis revealed that there is one dominant ER object accounting for most of the ER volume in the whole cell: i.e., the largest ER object takes up 22.37 μm3 or 93.5% of the ER volume in the cell. A post deep-learning processing algorithm used 3D object modifications to identify 11,109 3D MCS between mitochondria and ER in the 10 cells. The Mito-ER MCS ranged from 512 nm3 to 354,304 nm3 with a median volume of 1792 nm3. Visually we can see Mito-ER and Mito-PM MCS extending across a region of up to ~45 nm between the two organelles. Taken together, these data show that the basal lateral surfaces of PT cell PMs are generously invested with fenestrated smooth ER, as are mitochondria and peroxisomes. Extensive regions of direct contact between mitochondria and PM are also observed. The biochemical identities of these contacts and their physiological functions are being actively pursued. NIDDK DK072612 NIDDK DK120534. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

High Resolution Reconstruction of the Proximal Tubule Apical Endocytic Pathway

Kidney proximal tubule (PT) cells are specialized for effcient apical uptake of albumin and other proteins that escape the glomerular filtration barrier. The apical endocytic pathway is uniquely organized to maintain highly dynamic retrieval of these proteins via binding to the multiligand receptors megalin and cubilin. Ultrastructural studies following the fate of internalized markers have provided a temporal model for the delivery of receptor-bound ligands and fluid phase markers to lysosomes. In this model, internalization from the apical membrane occurs via budding of irregular clathrin-coated invaginations that form at the base of microvilli. After uncoating, budded vesicles fuse with apical early endosomes (AEEs) where acidification triggers dissociation of ligands from their receptors. AEEs further mature and expand into apical vacuoles (AVs) that ultimately deliver ligands and fluid to lysosomes. Receptor recycling to the apical membrane is thought to occur via dense apical tubules (DATs) that have a uniform diameter and membrane thickness and which appear to bud from both AEEs and AVs. We used Zeiss Arivis Pro technology to annotate and reconstruct apical membrane invaginations and endocytic compartments in a 3D volume of mouse kidney with isotropic voxels of 4 nm, acquired using focused ion-beam scanning electron microscopy (FIB-SEM). Strikingly, 3D reconstruction of DATs revealed a large and highly interconnected network of tubules in the subapical region of PT cells. Connections were observed between the DAT network and endosomal compartments, as well as between DATs and apical membrane invaginations. Our data suggest the possibility that DATs exist as a stable compartment that interacts transiently with the apical membrane to enable receptor recycling. Current studies are underway to better understand how PT cells modulate endocytic entry and recycling via the same portals. National Institute of Health: 5T32GM133353-03, R01 DK118726, R01 DK125049, U54 DK137329. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Synchrotron micro-computed tomography analysis of neutron-irradiated U-Mo fuel

The three-dimensional (3D) microstructure of neutron-irradiated uranium-10 wt.% molybdenum (U-10Mo) fuel with a burn-up of 9.8 × 1021 fissions/cm3 was characterized using a novel, multi-modal synchrotron micro-computed tomography approach combining propagation-based phase-contrast enhanced and absorption contrast techniques. The porosity development, porosity interconnectedness, swelling, composition, local thickness of the zirconium (Zr) diffusion barrier, and the influence of the fuel–cladding interaction on the local composition and pore morphology, were uniquely determined in 3D. Two cuboids were produced using a focused ion beam-scanning electron microscope at the Zr diffusion barrier–fuel interface and in the bulk fuel. The bulk fuel sample swelled by 53.3 [+9.7/−3.1]%, while the fuel near the Zr–fuel interface swelled by 63.3 [+14.7/−7.2]%. The average local thickness of the Zr diffusion barrier decreased by 53 %, compared to the expected pre-irradiated thickness. Four pore morphology regions were identified initiating parallel to the fuel–Zr interaction region: (1) an interaction layer of suppressed porosity, (2) a layer of elongated and interconnected porosity, (3) a transition zone of low porosity, and (4) a layer of unoriented porosity representative of the bulk fuel behavior. The increase in porosity near the diffusion barrier corresponded to a higher U concentration compared to that in the bulk fuel. The interconnected porosity in the fuel near the diffusion barrier was extensive and oriented parallel to the diffusion barrier, while the bulk fuel had more compact and isolated pore networks. The interaction layer, despite having suppressed porosity, was nearly 100 wt.% U. Porosity suppression at the diffusion barrier corresponds to the expected reduction in radiation-driven diffusion of Xe at the interface despite the anticipated increase in fission product nucleation originating from a higher U concentration. The novel 3D insights of the porosity, swelling, and compositional variations characterized herein can improve the fidelity of fuel performance codes for proliferation-resistant fuels for research and test reactors.

Approaching Technological Limit for Wet‐Pulling Technique

Carbon nanotube fibers (CNTFs) are a promising economical replacement material for contemporary metallic wired conductors due to their lightweight and advanced electromechanical properties. The wet‐pulling technique for CNTF manufacturing is highly versatile, adaptable for both laboratory as well as industrial‐scale production, and can be optimized for the maximum enhancement of electrophysical properties. Herein, a mechanosolvent‐based postfabrication approach for maximizing densification and improving electrical properties of wet‐pulled CNTFs is examined. The experimental process results in fibers achieving 60% of the theoretically maximum density and conductivity of maximally densified metallic single‐walled carbon nanotube bundles. The technique allows a corresponding increase of ≈700% in fiber density (from 100 to 704 kg m−3), a simultaneous ≈530% increase in electrical conductivity (from 748 to 3990 S cm−1), and reduced volume defects from 18% to 2%. The approach was combined with a step‐wise microstructure monitoring using focused ion beam–scanning electron microscopy to determine the mechanisms behind the optimized structures. This work is the first to provide an experimental and theoretical base for the postfabrication optimization of wet‐pulled CNTFs and lays the foundation for further enhancement with techniques such as chemical doping, fiber compounding, and combined infiltration/densification mechanisms.

High-Precision 3D reconstruction and quantitative structure description: Linking microstructure to macroscopic heat transfer of aerogels

The intricate interplay between microstructure and macroscopic transfer processes in porous materials has long been recognized, yet achieving a definitive link between them remains a formidable challenge. Herein, we propose an effective approach to address this challenge by introducing a high-precision 3D reconstruction and structure descriptor database analysis method of nanoporous aerogels, which enables deeper understanding and accurate predictions of structure-directed properties. By employing focused ion beam-scanning electron microscope (FIB-SEM) technique supplemented with image preprocessing and image segmentation, we successfully built a 3D reconstruction model that exhibits high consistency with experimental results. Then, a structure descriptor database from the view of original porous structure, structure connectivity, and topological information is established, and 10 descriptors are preliminarily filtered relevance with thermal conductivity. Consequently, the quantitative relationship between descriptor and thermal conductivity is evaluated via a back propagation (BP) neural network, and three categories structure descriptors which have the most significant impact are determined as (i) the description of gaseous heat transfer pathways, (ii) the description of three-dimensional connectivity, and (iii) the description of interface complexity. With the profound understanding of structure descriptor, the present work offers a novel, structure-focused strategy that closely link the microstructure to macroscopic properties of porous materials.

Experimental determination of Fe–Mg interdiffusion in orthopyroxene as a function of Fe content

We have measured the dependence of the Fe–Mg interdiffusion coefficient, DFe-Mg, on the ferrosilite component in orthopyroxene, which so far has not been experimentally calibrated. Diffusion couples, consisting of approximately 1 µm thin-films were deposited by pulsed laser ablation on orthopyroxene crystals of En91Fs9 composition. Diffusion experiments were carried out at atmospheric pressure in vertical gas mixing furnaces (CO-CO2) at temperatures between 950 and 1100 °C at constant fO2 = 10–7 Pa. Using a focused ion beam-scanning electron microscope (FIB-SEM), FIB-foils were cut from diffusion couples before and after annealing. Diffusion profiles were extracted by using combined backscattered electron (BSE) imaging and energy dispersive X-ray (EDXS) mapping on FIB-foils which allowed to resolve concentration gradients within 20 nm. The microstructure of the diffusion experiments was investigated using transmission electron microscopy (TEM). Using this method, we obtained the first experimentally determined data on the dependence of DFe-Mg on the ferrosilite content in orthopyroxene at different temperatures, appearing to increase with increasing a temperature. For the temperature range (950 – 1100 °C), log fO2 = -7 Pa, along [001] in the composition Fs9, the log DFe-Mg yields the following Arrhenius equation:DFe-Mg[m2/s]=3.8·10-9exp[-261.07±24[kJ/mol]/(R/T[K])]\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$D_{Fe - Mg} [m^2 /s] = 3.8 \\cdot 10^{ - 9} \\exp [ - 261.07 \\pm 24[kJ/mol]/(R/T[K])]$$\\end{document}The effect of XFe on DFe-Mg, given by D(XFe)=D(XFe=0.09)·10m(XFe-0.09),\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$D(X_{Fe} ) = D(X_{Fe} = 0.09) \\cdot {10}^{m (X_{Fe} - 0.09)},$$\\end{document} can be calculated by the following parameterization where m follows a linear regression of m on temperature: m=-2.711·104/T(K)+23.5408\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$m = - 2.711 \\cdot 10^4 /T(K) + 23.5408$$\\end{document}By considering the DFe-Mg dependence on XFe, the timescales of natural processes obtained from modelling the compositional zoning of natural crystal may considerably differ from previously estimated timescales.

Integrated Physical and Digital Chalk Relative Permeability Evaluation: A Case Study

The Valhall chalk field has produced more than 1 billion barrels of oil equivalents over the last 40 years, primarily from the homogeneous Tor Formation. The underlying Hod Formation is more heterogeneous and is less maturely developed. The extent of heterogeneity poses a challenge in the evaluation of multiphase fluid flow properties. The objective of the work was to use digital core analysis to generate early relative permeability data to leverage and compare with conventional physical steady-state relative permeability data. Accurate digital and physical description of capillary pressure and relative permeability in the high-porosity chalk is complicated by both low permeabilities and heterogeneity. The main challenge with chalk is that flow occurs in a nano-environment. Physically, the nano-environment translates to low permeability, difficult rock preparation, and extensive experimental time, especiallyfor steady-state flow experiments. Representative three-dimensional (3D) digital rocks were generated using a combination of X-ray computed tomography (CT) and focused ion beam-scanning electron microscopy (FIB-SEM) methods. The digital rocks were used to simulate two-phase flow and generate relative permeabilities and sensitivity to wettability. Physical steady-state and digital relative permeabilities on several core plugs and subsets are compared in this study, which discusses the advantages of performing both as part of the formation evaluation process. The physical and digital results compare reasonably well. The physical results provide a relative permeability anchor, and the digital results provide the leverage of early results, parametric sensitivities, and quality assurance. Hence, integration between digital and physical core analysis yields a robust understanding and input for uncertainty modeling.

Indirect Correlative Light and Electron Microscopy (iCLEM): A Novel Pipeline for Multiscale Quantification of Structure From Molecules to Organs.

Correlative light and electron microscopy (CLEM) methods are powerful methods that combine molecular organization (from light microscopy) with ultrastructure (from electron microscopy). However, CLEM methods pose high cost/difficulty barriers to entry and have very low experimental throughput. Therefore, we have developed an indirect correlative light and electron microscopy (iCLEM) pipeline to sidestep the rate-limiting steps of CLEM (i.e., preparing and imaging the same samples on multiple microscopes) and correlate multiscale structural data gleaned from separate samples imaged using different modalities by exploiting biological structures identifiable by both light and electron microscopy as intrinsic fiducials. We demonstrate here an application of iCLEM, where we utilized gap junctions and mechanical junctions between muscle cells in the heart as intrinsic fiducials to correlate ultrastructural measurements from transmission electron microscopy (TEM), and focused ion beam scanning electron microscopy (FIB-SEM) with molecular organization from confocal microscopy and single molecule localization microscopy (SMLM). We further demonstrate how iCLEM can be integrated with computational modeling to discover structure-function relationships. Thus, we present iCLEM as a novel approach that complements existing CLEM methods and provides a generalizable framework that can be applied to any set of imaging modalities, provided suitable intrinsic fiducials can be identified.

Melanin granules morphology and distribution in human black hair investigated by focused ion beam scanning electron microscopy: Differences between Asian and Caucasian hair

Melanin granules (melanosomes) in Asian and Caucasian black hairs were investigated by focused ion beam scanning electron microscopy (FIB-SEM). This technique facilitates a direct evaluation of the three-dimensional distribution and morphology of melanin granules without requiring their isolation from hair. Three-dimensional reconstructed images of melanin granule distribution in hair samples were obtained using serial SEM images observed by FIB-SEM. Melanin granules in black hair tended to be three-dimensionally dense in the outer periphery of the cortex. The morphometric parameters of melanin granules were calculated using the reconstructed three-dimensional images. The results confirmed that melanin granules in Caucasian black hair were much smaller those in Asian black hair. Moreover, it was indicated that the relative frequency distribution of the volume of melanin granules was significantly different between Asians and Caucasians.

Abstract 2117: Elucidation of the fates of CDK2 inhibited aneuploid and residual lung cancers

Abstract Genomic instability is a hallmark of cancer. Unlike normal diploid cells, cancer cells exhibit abnormal centrosome numbers and chromosomal instability. Mitosis with supernumerary centrosomes yields progeny with aberrant chromosome segregation and aneuploidy. While most progeny cells with abnormal chromosomes are eliminated through apoptosis, some have distinct fates with a growth advantage. These cancer cells can acquire resistance to antineoplastic agents. This confers an aggressive tumor biology and unfavorable clinical outcomes. Cyclin-dependent kinase 2 (CDK2) regulates cell cycle progression and the centrosome cycle. We previously reported that CDK2 inhibition prevents supernumerary centrosome clustering, causing multipolar mitosis and anaphase catastrophe in lung and other aneuploid cancers. This was associated with residual transplanted tumors in mice. To elucidate cell fates of progeny with multipolar mitosis after CDK2/9 inhibition with CYC065 (0.2 µM) treatment, a panel of murine and human lung cancer cells underwent time-lapse fluorescent microscopy. This elucidated multipolar mitotic events within the progeny. Intriguingly, distinct outcomes occurred after mitosis including formation of multipolar and multinucleated cells. Cell death of progeny is the predominant fate, but some progeny survive despite continuous CDK2 inhibition. Surprisingly, some of these cells fuse together. Those multinucleated cells can undergo multiple cell cycles without successful cytokinesis. To confirm these effects were through CDK2 inhibition, the selective CDK2 inhibitor Tagtociclib (PF-07104091, at the 2 µM dosage) was used. This treatment statistically-significantly increased multipolar lung cancer cells and increased apoptotic death during time-lapse fluorescent microscopy. Findings were independently validated by CDK2 shRNA knockdown. Focused ion beam scanning electron microscopy (FIB-SEM) and immunofluorescent staining revealed the ultrastructure of multinucleated cells. Insights into tumor biology came from intravital imaging of transplanted lung cancer cells in mice. Multipolar and multinuclear cells followed CDK2 antagonism. Incucyte® Live-Cell Analysis System and Artificial Intelligence-based imaging determined distinct growth responses in aneuploid as compared to non-aneuploid lung cancer cells after CDK2 inhibitor treatments. In summary, these findings are translationally relevant. CDK2 inhibition of aneuploid lung cancers yields distinct cell fates. This is linked to resistance to CDK2 antagonism and to formation of residual in vivo tumors. Citation Format: Liliya Tyutyunyk, Zibo Chen, Xiuxia Liu, Yeap Ng, Aayush Bhatawadekar, Kedar Narayan, Roberto Weigert, Xi Liu, Ethan Dmitrovsky. Elucidation of the fates of CDK2 inhibited aneuploid and residual lung cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2117.

Microscopic seepage simulation of gas and water in shale pores and slits based on VOF

Abstract The microscopic pore-fracture structure and wettability have a significant influence on the two-phase seepage of shale gas and water. Due to the limitation of experimental conditions, the seepage patterns of gas and water in shale pores and slits under different wetting conditions have not been clarified yet. In this study, the three-dimensional digital rock models of shale inorganic pores, organic pores, and microfractures are established by focused ion beam-scanning electron microscopy scanning, and gas-driven water seepage simulation in shale microscopic pore-fracture structure under different wetting conditions is carried out based on volume of fluid method. The simulation results show that the gas–water relative permeability curves of microfractures are up-concave, and the gas–water relative permeability curves of inorganic and organic pores are up-convex; the gas–water two-phase percolation in microfractures is least affected by the change of wettability, the gas–water two-phase percolation in inorganic pores is most affected by the change of wettability, and the organic pores are in between; the gas–water two-phase percolation zone of microfractures is the largest, and the isotonic saturation is the highest; under the water-wet conditions, the critical gas saturation of microfractures, inorganic pores, and organic pores are 0.13, 0.315, and 0.34, respectively, and the critical gas saturation of organic pores under non-water-wet conditions is 0.525, indicating that under water-bearing conditions, the shale gas flow capacity in water-wet microfractures is the strongest, followed by water-wet inorganic pores, water-wet organic pores, and hydrophobic organic pores, respectively.

Direct Visualization of Chemical Transport in Solid-State Chemical Reactions by Time-of-Flight Secondary Ion Mass Spectrometry.

Systematic control and design of solid-state chemical reactions are required for modifying materials properties and in novel synthesis. Understanding chemical dynamics at the nanoscale is therefore essential to revealing the key reactive pathways. Herein, we combine focused ion beam-scanning electron microscopy (FIB-SEM) and time-of-flight secondary ion mass spectrometry (TOF-SIMS) to track the migration of sodium from a borate coating to the oxide scale during in situ hot corrosion testing. We map the changing distribution of chemical elements and compounds from 50 to 850 °C to reveal how sodium diffusion induces corrosion. The results are validated by in situ X-ray diffraction and post-mortem TOF-SIMS. We additionally retrieve the through-solid sodium diffusion rate by fitting measurements to a Fickian diffusion model. This study presents a step change in analyzing microscopic diffusion mechanics with high chemical sensitivity and selectivity, a widespread analytical challenge that underpins the defining rates and mechanisms of solid-state reactions.

Key role of gradient-nanostructure and extremely thin amorphous passive film on tribocorrosion behavior of a novel Cr+N alloyed high-Mn austenitic steel

High manganese steel is influenced by the combined effects of corrosion and wear during service under specific conditions, resulting in components with shortened service life. Under harsh service environments, newly developed Cr+N alloyed austenitic high manganese steel (Mn18Cr7C0.6N0.2 steel) as a new type of railway steel has shown promising results. Herein, the tribocorrosion behaviors of Mn18Cr7C0.6N0.2 and Mn13C1.1 steels in artificial acid rain were studied using electrochemical methods, field emission scanning electron microscope (FE-SEM), focused ion beam scanning electron microscopy (FIB-SEM), scanning transmission electron microscopemicroscopy energy dispersive X-ray spectroscopy (STEM-EDS), and high resolution transmission electron microscopy (HR-TEM). The results showed the formation of passive film and gradient-nanostructure on surfaces of both test steels during tribocorrosion. A dense and tightly adhering amorphous passive film was formed on Mn18Cr7C0.6N0.2 steel surface with a superior blocking effect toward invasive ions. The high strain resistance and plasticity of the Mn18Cr7C0.6N0.2 steel resulted in an intact gradient-nanostructure, suitable for obtaining high surface hardness and maintaining the toughness of the matrix, contribute to providing high wear resistance. The oxide film on the Mn13C1.1 steel surface comprised crystal α-FeOOH. Vortex cracks formed in its gradient structure due to strain localization. Meanwhile, the relatively poor corrosion protection ability and vortex cracks of its gradient-nanostructure prevented it from maintaining interface integrity of the gradient nanostructure, resulting in inferior tribocorrosion resistance. In artificial acid rain, the new type of Cr+N alloyed Mn18Cr7C0.6N0.2 steel exhibited higher tribocorrosion resistance than traditional high manganese steel, promising for use as an excellent wear-resistant material under corrosive environments.

A Multiscale Approach for Assessing Shale Oil Availability: Digital Core, Molecular Simulation, and Machine Learning Analysis

Summary This study presents a novel multiscale approach for assessing the accessibility of shale oil in cores by use of focused ion beam-scanning electron microscopy (FIB-SEM) equipment to build digital core, watershed, and maximum ball methods to extract pore size and shape factor. Then, molecular simulation is used to study the availability of shale oil in individual pores with different shapes and radii. Finally, combining the results of the above two scales, machine learning is used to predict shale oil availability across the entire core. On the core scale, the watershed and maximum ball methods are used to extract the core pore network model, and it is found that square pores occupy the highest proportion among the three pore types, and most of the radii are distributed in the range of 2–3 nm. The molecular-scale dynamic simulation results show that the adsorption forms of shale oil are different in different pores, and the adsorption of shale oil in circular pores is less than that in flat pores. The proportion of shale oil adsorption in square pores is the highest, followed by triangular pores, and the proportion of shale oil adsorption in circular pores is the lowest. The random forest machine learning algorithm is used to predict the availability of shale oil with different pore shapes and obtain the shale oil availability ratio of the whole core. The results show that the pores with a more obvious angular structure show a lower shale oil availability ratio. In general, the impact of pore shapes on shale oil availability is not significant, and the difference between different pore shapes is only 10%. The multiscale evaluation method for shale oil availability proposed in this study is helpful to better understand the availability of shale oil in reservoirs and to optimize recovery strategies.

Synthesis of submicron-sized LiNi0.5Co0.2Mn0.3O2 cathode particles via reverse microemulsion method: Tailoring size, interfacial properties, and enhanced electrochemical performance for lithium-ion battery

This study explores the impact of submicron-sized cathode materials on lithium-ion battery performance, an area with limited prior research due to synthetic challenges. Employing the reverse microemulsion method, LiNi0.5Co0.2Mn0.3O2 (NCM) cathode particles with tunable sizes (180 nm to 340 nm) were synthesized. Notably, NCM6.67-4.5, which prepared under the volume ratio of the oil phase to the water phase (i.e. 6.67) and applying 4.5 mL NH4OH as the complexing agent, obtained the largest particle size (340 nm). Further investigation revealed that NCM6.67-4.5 exhibited superior discharge specific capacity (177.54 mAh·g−1) and a high coulomb efficiency (99.5 %). The larger particle size in the submicron range facilitated heightened lithium-ion diffusion rates and reduced polarization. Focused Ion Beam-Scanning Electron Microscopy (FIB-SEM) confirmed that increased particle size diminishes specific surface area, mitigating interfacial side reactions during battery cycling. This research underscores the importance of controlling cathode particle size for optimizing electrochemical performance in lithium-ion batteries.

Silencing NlFAR7 destroyed the pore canals and related structures of the brown planthopper.

Fatty acyl-CoA reductase (FAR) is one of the key enzymes, which catalyses the conversion of fatty acyl-CoA to the corresponding alcohols. Among the FAR family members in the brown planthopper (Nilaparvata lugens), NlFAR7 plays a pivotal role in both the synthesis of cuticular hydrocarbons and the waterproofing of the cuticle. However, the precise mechanism by which NlFAR7 influences the formation of the cuticle structure in N. lugens remains unclear. Therefore, this paper aims to investigate the impact of NlFAR7 through RNA interference, transmission electron microscope, focused ion beam scanning electron microscopy (FIB-SEM) and lipidomics analysis. FIB-SEM is employed to reconstruct the three-dimensional (3D) architecture of the pore canals and related cuticle structures in N. lugens subjected to dsNlFAR7 and dsGFP treatments, enabling a comprehensive assessment of changes in the cuticle structures. The results reveal a reduction in the thickness of the cuticle and disruptions in the spiral structure of pore canals, accompanied by widened base and middle diameters. Furthermore, the lipidomics comparison analysis between dsNlFAR7- and dsGFP-treated N. lugens demonstrated that there were 25 metabolites involved in cuticular lipid layer synthesis, including 7 triacylglycerols (TGs), 5 phosphatidylcholines (PCs), 3 phosphatidylethanolamines (PEs) and 2 diacylglycerols (DGs) decreased, and 4 triacylglycerols (TGs) and 4 PEs increased. In conclusion, silencing NlFAR7 disrupts the synthesis of overall lipids and destroys the cuticular pore canals and related structures, thereby disrupting the secretion of cuticular lipids, thus affecting the cuticular waterproofing of N. lugens. These findings give significant attention with reference to further biochemical researches on the substrate specificity of FAR protein, and the molecular regulation mechanisms during N. lugens life cycle.

Pore-space characterisation of Tamusu mudstone based on FIB-SEM

This study focuses on characterising the pore space in Tamusu mudstone, a preferred nuclear waste disposal host medium in China, using focused ion beam/scanning electron microscopy (FIB-SEM). The investigation is conducted at the mesoscale, with a sample volume of 7·96 × 9·16 × 10·00 μm3 and voxel sizes of 5 × 5 × 10 nm3. The findings reveal that the pores in Tamusu mudstone primarily exist at the nanoscale, typically within its mineral matrix or at interfaces with non-porous minerals. At the FIB/SEM-imaged scale, the pore space in the Tamusu mudstone is mainly unconnected, with an analysed porosity of approximately 3·5%. Pore characteristics within the specimen exhibit relative consistency within a box size of 6–8 μm. Furthermore, the pore paths in Tamusu mudstone exhibit a preferred orientation within the bedding plane, suggesting anisotropy in pore space, possibly attributed to changes in pore path orientation and tortuosity.

The application and development of electron microscopy for three-dimensional reconstruction in life science: a review.

Imaging technologies have played a pivotal role in advancing biological research by enabling visualization of biological structures and processes. While traditional electron microscopy (EM) produces two-dimensional images, emerging techniques now allow high-resolution three-dimensional (3D) characterization of specimens in situ, meeting growing needs in molecular and cellular biology. Combining transmission electron microscopy (TEM) with serial sectioning inaugurated 3D imaging, attracting biologists seeking to explore cell ultrastructure and driving advancement of 3D EM reconstruction. By comprehensively and precisely rendering internal structure and distribution, 3D TEM reconstruction provides unparalleled ultrastructural insights into cells and molecules, holding tremendous value for elucidating structure-function relationships and broadly propelling structural biology. Here, we first introduce the principle of 3D reconstruction of cells and tissues by classical approaches in TEM and then discuss modern technologies utilizing TEM and on new SEM-based as well as cryo-electron microscope (cryo-EM) techniques. 3D reconstruction techniques from serial sections, electron tomography (ET), and the recent single-particle analysis (SPA) are examined; the focused ion beam scanning electron microscopy (FIB-SEM), the serial block-face scanning electron microscopy (SBF-SEM), and automatic tape-collecting lathe ultramicrotome (ATUM-SEM) for 3D reconstruction of large volumes are discussed. Finally, we review the challenges and development prospects of these technologies in life science. It aims to provide an informative reference for biological researchers.

Robust Preparation of Sub-20-nm-Thin Lamellae for Aberration-Corrected Electron Microscopy.

Aberration-corrected scanning transmission electron microscopy (STEM) has been advancing resolution, sensitivity, and microanalysis due to the intense demands of atomic-level microstructural investigations. Recent STEM technologies require preparing a thin lamella whose thickness is ideally below 20 nm. Although focused-ion-beam/scanning-electron-microscopy (FIB/SEM) is an established method to prepare a high-quality lamella, nanometer-level controllability of lamella thickness remains a fundamental problem. Here, the robust preparation of a sub-20-nm-thin lamella is demonstrated by FIB/SEM with real-time feedback from thickness quantification. The lamella thickness is quantified by back-scattered-electron SEM imaging in a thickness range between 0 and 100 nm without any reference to numerical simulation. Using real-time feedback from the thickness quantification, the FIB/SEM terminates thinning a lamella at a targeted thickness. The real-time feedback system eventually provides 1-nm-level controllability of the lamella thickness. As a proof-of-concept, a near-10-nm-thin lamella is prepared from a SrTiO3 crystal by ourmethodology. Moreover, the lamella thickness is controllable at a target heterointerface. Thus, a sub-20-nm-thin lamella is prepared from a LaAlO3 /SrTiO3 heterointerface. The methodology offers a robust and operator-independent platform to prepare a sub-20-nm-thin lamella from various materials. This platform will broadly impact aberration-corrected STEM studies in materials science and the semiconductorindustry.

Development and application of a 3D image analysis strategy for focused ion beam - Scanning electron microscopy tomography of porous soft materials.

In recent years, the potential of porous soft materials in various device technologies has increased in importance due to applications in fields, such as wearable electronics, medicine, and transient devices. However, understanding the 3-dimensional architecture of porous soft materials at the microscale remains a challenge. Herein, we present a method to structurally analyze soft materials using Focused Ion Beam - Scanning Electron Microscopy (FIB-SEM) tomography. Two materials, polymethyl methacrylate (PMMA) membrane and pine wood veneer were chosen as test-cases. FIB-SEM was successfully used to reconstruct the true topography of these materials in 3D. Structural and physical properties were subsequently deduced from the rendered 3D models. The methodology used segmentation, coupled with optimized thresholding, image processing, and reconstruction protocols. The 3D models generated pore size distribution, pore inter-connectivity, tortuosity, thickness, and curvature data. It was shown that FIB-SEM tomography provides both an informative and visual depiction of structure. To evaluate and validate the FIB-SEM reconstructions, porous properties were generated from the physical property analysis techniques, gas adsorption analysis using Brunauer-Emmett-Teller (BET) surface area analysis and mercury intrusion porosimetry (MIP) analysis. In general, the data obtained from the FIB-SEM reconstructions was well-matched with the physical data. RESEARCH HIGHLIGHTS: Porous specimens of both synthetic and biological nature, a poly(methyl methacrylate) membrane and a pine veneer respectively, are reconstructed via FIB-SEM tomography without resin-embedding. Different thresholding and reconstruction methods are explored whereby shadowing artifacts are present with the aid of free open-source software. Reconstruction data is compared to physical data: MIP, gas adsorption isotherms which are analyzed via BET and Barrett-Joyner-Halenda (BJH) analysis to yield a full picture of the materials.