Large-scale electron microscopy (EM) has enabled the reconstruction of brain connectomes at the synaptic level by serially scanning over massive areas of sample sections. The acquired big EM data sets raise the great challenge of image mosaicking at high accuracy. Currently, it simply follows the conventional algorithms designed for natural images, which are usually composed of only a few tiles, using a single type of keypoint feature that would sacrifice speed for stronger performance. Even so, in the process of stitching hundreds of thousands of tiles for large EM data, errors are still inevitable and diverse. Moreover, there has not yet been an appropriate metric to quantitatively evaluate the stitching of biomedical EM images. Here we propose a two-stage error detection method to improve the EM image mosaicking. It firstly uses point-based error detection in combination with a hybrid feature framework to expedite the stitching computation while maintaining high accuracy. Following is the second detection of unresolved errors with a newly designed metric of EM stitched image quality assessment (EMSIQA). The novel detection-based mosaicking pipeline is tested on large EM data sets and proven to be more effective and as accurate when compared with existing methods.

Structural studies require the production of target proteins in large quantities and with a high degree of purity. For membrane proteins, the bottleneck in determining their structure is the extraction of the target protein from the cell membranes. A detergent that improperly mimics the hydrophobic environment of the protein of interest can also significantly alter its structure. Recently, using lipodiscs with styrene-maleic acid (SMA), copolymers became a promising strategy for the purification of membrane proteins. Here, we describe in detail the one-step affinity purification of potassium ion channels solubilized in SMA and sample preparation for future structural studies.

OBJECTIVE: Podocyte injury plays an essential role in the progression of diabetic nephropathy (DN). The associations between the ultrastructural changes of podocyte with proteinuria and the pathological classification of DN proposed by Renal Pathology Society (RPS) have not been clarified in patients with type 2 diabetic nephropathy (T2DN).
METHODS: We collected 110 patients with kidney biopsy-confirmed T2DN at Peking University First Hospital from 2017 to 2022. The morphometric analysis on the podocyte foot process width (FPW) and podocyte detachment (PD) as markers of podocyte injury was performed, and the correlations between the ultrastructural changes of podocytes with severity of proteinuria and the RPS pathological classification of DN were analyzed.
RESULTS: Mean FPW was significantly broader in the group of T2DN patients with nephrotic proteinuria (565.1 nm) than those with microalbuminuria (437.4 nm) or overt proteinuria (494.6 nm). The cut-off value of FPW (> 506 nm) could differentiate nephrotic proteinuria from non-nephrotic proteinuria with a sensitivity of 75.3% and a specificity of 75.8%. Percentage of PD was significantly higher in group of nephrotic proteinuria (3.2%) than that in microalbuminuria (0%) or overt proteinuria (0.2%). FPW and PD significantly correlated with proteinuria in T2DN (r = 0.473, p < 0.001 and r = 0.656, P < 0.001). FPW and PD correlated with RPS pathological classification of T2DN (r = 0.179, P = 0.014 and r = 0.250, P = 0.001). FPW value was increased significantly with more severe DN classification (P for trend =0.007). The percentage of PD tended to increase with more severe DN classification (P for trend = 0.017).
CONCLUSIONS: Podocyte injury, characterized by FPW broadening and PD, was associated with the severity of proteinuria and the pathological classification of DN.

Soot particles emitted from aircraft engines constitute a major anthropogenic source of pollution in the vicinity of airports and at cruising altitudes. This emission poses a significant threat to human health and may alter the global climate. Understanding the characteristics of soot particles, particularly those generated from Twin Annular Premixing Swirler (TAPS) combustors, a mainstream combustor in civil aviation engines, is crucial for aviation environmental protection. In this study, a comprehensive characterization of soot particles emitted from TAPS combustors was conducted using scanning electron microscopy (SEM), high-resolution transmission electron microscopy (HRTEM), and Raman spectroscopy. The morphology and nanostructure of soot particles were examined across three distinct fuel stage ratios (FSR), at 10%, 15%, and 20%. The SEM analysis of soot particle morphology revealed that coated particles constitute over 90% of the total particle sample, with coating content increasing proportionally to the fuel stage ratio. The results obtained from HRTEM indicated that average primary particle sizes increase with the fuel stage ratio. The results of HRTEM and Raman spectroscopy suggest that the nanostructure of soot particles becomes more ordered and graphitized with an increasing fuel stage ratio, resulting in lower oxidation activity. Specifically, soot fringe length increased with the fuel stage ratio, while soot fringe tortuosity and separation distance decreased. In addition, there is a prevalent occurrence of defects in the graphitic lattice structure of soot particles, suggesting a high degree of elemental carbon disorder.

In animal cells, the Golgi apparatus serves as the central hub of the endomembrane secretory pathway. It is responsible for the processing, modification, and sorting of proteins and lipids. The unique stacking and ribbon-like architecture of the Golgi apparatus forms the foundation for its precise functionality. Under cellular stress or pathological conditions, the structure of the Golgi and its important glycosylation modification function may change. It is crucial to employ suitable methodologies to study the structure and function of the Golgi apparatus, particularly when assessing the involvement of a target protein in Golgi regulation. This article provides a comprehensive overview of the diverse microscopy techniques used to determine the specific location of the target protein within the Golgi apparatus. Additionally, it outlines methods for assessing changes in the Golgi structure and its glycosylation modification function following the knockout of the target gene.

This study discusses the micro-level structural details of Cichorieae pollen sources elucidated by scanning electron microscopy (SEM) and explains their symmetry and morphometry. The in-depth knowledge from the electron ultrastructure of Asteraceae pollen has provided insights into enhanced pollen morphology, and the antimicrobial significance of species under study presents novel avenues for their natural defense mechanisms in the development of antimicrobial agents. In this research, both quantitative and qualitative features of pollen were examined. The pollen grains are prolate-spheroidal and oblate-spheroidal in shape, characterized by a maximum polar diameter of 55.6-61.0 μm and a maximum equatorial distance of 68.3-74.4 μm. SEM reveals various configurations such as echinate perforate-tectate, psilate, and echino-lophate perforate. The Cichorieae species have significant antimicrobial efficacy and are promising sources for the development of novel antimicrobial drugs with potential implications in pharmaceutical and healthcare industries. SEM analysis of Cichorieae pollens has provided remarkable insights into their unique structures, revealing diverse shapes and surface ornamentations, which can be used for accurate Asteraceae species identification. RESEARCH HIGHLIGHTS: SEM provides unique pollen surface structures and patterns of Chicory pollen grains. Chemical composition of Chicory botanical sources provides valuable information on their potential as antimicrobial agents. SEM imaging reveals specialized fenestrate grain structures of taxonomic importance.

This review presents an overview of the application of electrochemical liquid-phase transmission electron microscopy (ELP-TEM) in visualizing rechargeable battery reactions. The technique provides atomic-scale spatial resolution and real-time temporal resolution, enabling direct observation and analysis of battery materials and processes under realistic working conditions. The review highlights key findings and insights obtained by ELP-TEM on the electrochemical reaction mechanisms and discusses the current limitations and future prospects of ELP-TEM, including improvements in spatial and temporal resolution and the expansion of the scope of materials and systems that can be studied. Furthermore, the review underscores the critical role of ELP-TEM in understanding and optimizing the design and fabrication of high-performance, long-lasting rechargeable batteries.

Germline cells are critical for transmitting genetic information to subsequent generations in biological organisms. While their differentiation from somatic cells during embryonic development is well-documented in most animals, the regulatory mechanisms initiating plant germline cells are not well understood. To thoroughly investigate the complex morphological transformations of their ultrastructure over developmental time, nanoscale 3D reconstruction of entire plant tissues is necessary, achievable exclusively through electron microscopy imaging. This paper presents a full-process framework designed for reconstructing large-volume plant tissue from serial electron microscopy images. The framework ensures end-to-end direct output of reconstruction results, including topological networks and morphological analysis. The proposed 3D cell alignment, denoise, and instance segmentation pipeline (3DCADS) leverages deep learning to provide a cell instance segmentation workflow for electron microscopy image series, ensuring accurate and robust 3D cell reconstructions with high computational efficiency. The pipeline involves five stages: the registration of electron microscopy serial images; image enhancement and denoising; semantic segmentation using a Transformer-based neural network; instance segmentation through a supervoxel-based clustering algorithm; and an automated analysis and statistical assessment of the reconstruction results, with the mapping of topological connections. The 3DCADS model\'s precision was validated on a plant tissue ground-truth dataset, outperforming traditional baseline models and deep learning baselines in overall accuracy. The framework was applied to the reconstruction of early meiosis stages in the anthers of Arabidopsis thaliana, resulting in a topological connectivity network and analysis of morphological parameters and characteristics of cell distribution. The experiment underscores the 3DCADS model\'s potential for biological tissue identification and its significance in quantitative analysis of plant cell development, crucial for examining samples across different genetic phenotypes and mutations in plant development. Additionally, the paper discusses the regulatory mechanisms of Arabidopsis thaliana\'s germline cells and the development of stamen cells before meiosis, offering new insights into the transition from somatic to germline cell fate in plants.

P2X receptors are extracellular ATP-gated ion channels that form homo- or heterotrimers and consist of seven subtypes. They are expressed in various tissues, including neuronal and nonneuronal cells, and play critical roles in physiological processes such as neurotransmission, inflammation, pain, and cancer. As a result, P2X receptors have attracted considerable interest as drug targets, and various competitive inhibitors have been developed. However, although several P2X receptor structures from different subtypes have been reported, the limited structural information of P2X receptors in complex with competitive antagonists hampers the understanding of orthosteric inhibition, hindering the further design and optimization of those antagonists for drug discovery. We determined the cryogenic electron microscopy (cryo-EM) structures of the mammalian P2X7 receptor in complex with two classical competitive antagonists of pyridoxal-5\'-phosphate derivatives, pyridoxal-5\'-phosphate-6-(2\'-naphthylazo-6\'-nitro-4\',8\'-disulfonate) (PPNDS) and pyridoxal phosphate-6-azophenyl-2\',5\'-disulfonic acid (PPADS), and performed structure-based mutational analysis by patch-clamp recording as well as molecular dynamics (MD) simulations. Our structures revealed the orthosteric site for PPADS/PPNDS, and structural comparison with the previously reported apo- and ATP-bound structures showed how PPADS/PPNDS binding inhibits the conformational changes associated with channel activation. In addition, structure-based mutational analysis identified key residues involved in the PPNDS sensitivity of P2X1 and P2X3, which are known to have higher affinity for PPADS/PPNDS than other P2X subtypes.

Herein, we present an atomic in-situ investigation of Cu oxidation along different orientations stimulated by high-energy electron beams (E-Beam) in transmission electron microscopy (TEM). By following the microstructural evolution of the Cu substrate in real time, high-resolution TEM (HRTEM) images reveal an orientation-dependent oxidation mechanism, whereby Cu along [110] zone axis migrates onto the surface and be oxidized while Cu along [100] zone axis is oxidized completely both in bulk and at the surface. The different oxidation mechanisms can be attributed to the differing diffusion rates of oxygen in Cu structures along directions. Moreover, the growth of Cu oxides is found to follow a layer-by-layer mechanism, where Cu mostly migrates onto nanocrystal {110} planes. This behavior would lead to the oxides wider in geometric shape and therefore promote the aggregation of adjacent oxides. These findings have important implications for the practical use of copper-based materials in oxidizing environments.