Journal of microscopy最新文献

{"title":"Ptychography: A brief introduction","authors":"John Rodenburg","doi":"10.1111/jmi.70025","DOIUrl":"10.1111/jmi.70025","url":null,"abstract":"<p>For anyone new to ptychography, the first obstacle to overcome is how to pronounce its name. The author has heard many tortured attempts trying to simultaneously incorporate the ‘p’ with the ‘t’—an impossible task. The answer is very simple: forget the ‘p’—in English it is silent, just as in ‘psychology’. Pronounce it as ‘tykography’.</p><p>Ptychography overcomes the two most enduring historical weaknesses of conventional transmission (and reflection) microscopy. It can in principle obtain wavelength limited resolution, unaffected by lens aberration or the maximum scattering angle imposed by the numerical aperture of the lens. This is especially important for X-ray and electron imaging where, for various intractable reasons, the useable numerical aperture of the available lenses is so small. It can also record the image phase near perfectly, meaning that otherwise transparent objects can be imaged with very high contrast.</p><p>Unlike conventional microscopy with lenses, ptychography does not provide a real or virtual image that can be seen directly. Instead, it uses a computer to process a very large quantity of data that bear no relationship to the final image that it ‘reconstructs’. Ordinary microscopists—that is, those who simply want to see a magnified image of their specimen and do not want to understand exactly how the image is computed—can find this circuitous process all rather alienating. First results from the author's group in the early 1990s were widely dismissed by the community. A leading microscopist at the time asserted that he would never believe in an image that came out of a computer. A further problem was that the pictures we could obtain in those days were so small and totally unconvincing. Ptychography had to wait for Moore's Law to catch up with its greedy data requirements.</p><p>However, in the last 10–15 years, ptychography has become the technique of choice for very high-resolution X-ray imaging and tomography. In the last 5 years or so, some extraordinary electron ptychography results have been reported, far surpassing the resolution limit that for so many years had seemed insurmountable using magnetic lenses and aberration correction. Optical microscopy is already wavelength limited, but the very sensitive phase image that ptychography supplies has removed the need for staining or labelling, thus allowing live imaging of biological cells.</p><p>The experimental method is deceptively simple. We have a source of radiation which shines upon the specimen. The wavefield at the exit surface of this specimen is then allowed to propagate some distance downstream of the object where the pattern of scattered intensity is recorded on a two-dimensional detector. It is important to understand that this detector can be as large as we like. It can capture scattering up to large angles, where high-resolution information is expressed. Electron and X-ray lenses can only capture and focus reliably small angles of scatter, which seve","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"300 2","pages":"153-155"},"PeriodicalIF":1.9,"publicationDate":"2025-08-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.70025","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144957862","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"TOC - Issue Information","authors":"","doi":"10.1111/jmi.13330","DOIUrl":"https://doi.org/10.1111/jmi.13330","url":null,"abstract":"","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"299 3","pages":""},"PeriodicalIF":1.9,"publicationDate":"2025-08-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.13330","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144833137","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Application of STEM tomography to investigate smooth ER morphology under stress conditions","authors":"V. Heinz,&nbsp;R. Rachel,&nbsp;C. Ziegler","doi":"10.1111/jmi.70020","DOIUrl":"https://doi.org/10.1111/jmi.70020","url":null,"abstract":"<p>The endoplasmic reticulum (ER) is a highly dynamic organelle that undergoes significant morphological alterations in response to cellular stress. While conventional transmission electron microscopy (TEM) has provided valuable insights into these changes, such as the formation of crystalloid-ER and ER whorls, obtaining comprehensive three-dimensional (3D) information on these large structures within their cellular context has remained a challenge. To overcome these limitations, this study introduces an innovative application of dual-axis scanning transmission electron microscopy (STEM) tomography to investigate ER morphology under stress conditions in human embryonic kidney (HEK) cells overexpressing the cation channel polycystin-2 (PC-2). Benefitting from high-resolution, increased depth-of-focus, and reduced aberrations, STEM tomography enabled the detailed 3D reconstruction of large cellular subvolumes, providing unprecedented views of stress-induced ER structures. Our findings reveal distinct ultrastructural details of both crystalloid-ER and ER whorls. Crystalloid-ER exhibited a tubular architecture with potential interconnectedness, while ER whorls displayed a lamellar organisation and distinct membrane curvature. We observed the co-occurrence of these distinct smooth ER (sER) morphotypes within the same cell, yet they remained spatially separated, suggesting potential functional specialisation. Furthermore, we identified direct membrane contacts in mixed morphotypes, hinting at a shared origin or dynamic relationship between these structures. The study also elucidated the interactions of these organised smooth ER (OSER) structures with other organelles, such as mitochondria (MAM sites) and vesicles. In summary, the presented ultra-structural insights have a significant impact on our understanding of stress-related ER morphology changes. The ability to visualise the intricate 3D architecture and spatial relationships of these structures provides novel perspectives on the ER's adaptive responses to stress, including potential roles in lipid and protein biosynthesis and intracellular communication. These findings underscore the power of dual-axis STEM tomography for elucidating complex organellar organisation and dynamics in their native cellular context.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"299 3","pages":"228-241"},"PeriodicalIF":1.9,"publicationDate":"2025-08-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.70020","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144832891","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Introduction to the Festschrift for Paul Walther","authors":"Clarissa Read,&nbsp;Ulla Neumann","doi":"10.1111/jmi.70023","DOIUrl":"10.1111/jmi.70023","url":null,"abstract":"","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"299 3","pages":"179-185"},"PeriodicalIF":1.9,"publicationDate":"2025-08-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144816904","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A quantitative irradiation microscopy platform for in situ observation of living cells with subcellular resolution under radiation conditions.","authors":"Zhao Chen, Liang Li, Jianli Liu, Yufang Zhao, Chenguang Liu, Jian Liu","doi":"10.1111/jmi.70024","DOIUrl":"https://doi.org/10.1111/jmi.70024","url":null,"abstract":"<p><p>To address the critical need for investigating proton radiation effects on living cells in space environments and deciphering biological mechanisms underlying low-dose cumulative radiation effects, this study developed a microbeam irradiation microscopy platform. The system integrates a 10 MeV proton accelerator with a vertical microbeam line design. An ultrafast single-proton counting and radiation synchronisation control module-employing proton-photon-electron conversion and high-speed photoelectric circuitry achieve deterministic irradiation control with an end-to-end operational delay of 273.5 ns. Coupled with wide-field and confocal fluorescence microscopy, the platform enables real-time in situ observation during quantitative cellular irradiation, facilitating mechanistic studies of radiation-induced damage patterns and signal transduction in low-dose scenarios. Experimental validation using human embryonic kidney 293T cells demonstrated successful simulation of space radiation environments: dose-dependent DNA double-strand breaks (visualised via γ-H2AX foci) and radiation-induced bystander effects triggering damage propagation. These results establish the platform as an indispensable tool for space radiation health risk assessment while providing foundational insights into microscale energy deposition dynamics for proton therapy research.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":" ","pages":""},"PeriodicalIF":1.9,"publicationDate":"2025-08-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144799440","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"PRIAMOS: A technique for mixing embedding media for freely adjusting pH value and refractive index (RI) for optical clearing (OC) of whole tissue samples.","authors":"Ulrich Leischner, Martin Reifarth, Monika S Brill, Florian Schmitt, Stephanie Hoeppener, David Unnersjö Jess, Hjalmar Brismar, Ulrich S Schubert, Rainer Heintzmann","doi":"10.1111/jmi.70022","DOIUrl":"https://doi.org/10.1111/jmi.70022","url":null,"abstract":"<p><p>Investigations of biological samples often require sample transparency, which is achieved by embedding the sample in a high-refractive index (RI) medium to obtain a homogenous RI distribution in the sample, referred to as optical clearing (OC). Here, we introduce a method for designing embedding media with an increased RI by increasing molecular orbitals, which is achieved by replacing elements in molecules commonly used for OC with elements possessing a more pronounced polarisability. Briefly, we took the established embedding medium Glycerol and exchanged the OH-groups by Thiol-groups, resulting in an embedding medium with very similar properties, but with a higher refractive index. We describe a procedure-abbreviated PRIAMOS-to render biological samples transparent using an RI-matching liquid, which we refer to as pH-value and Refractive Index Adjustment by Mixing highly polarisable molecular Orbital Substances. We focus on optical clearing in three-dimensional tissue samples whilst preserving fluorescence of fluorescent labels. The clearing procedure requires 2-3 days, yielding highly transparent samples, preserving the fluorescence of fluorescent proteins like the yellow fluorescent protein (YFP). This is demonstrated on mouse brain samples, imaged with standard confocal microscopy down to 1.6 mm depth, as well as on kidney samples. Mixtures of monothioglycerol, dithioglycerol and tributylamine achieve RI values between 1.52 and 1.57, and an acidity equivalent to pH values between 5 and 8. Our PRIAMOS approach can serve as a guideline for optimising optical clearing protocols.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":" ","pages":""},"PeriodicalIF":1.9,"publicationDate":"2025-08-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144799441","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Comparison of two Monte Carlo approaches for homogeneous bulk samples","authors":"Dawei Gao,&nbsp;Yu Yuan,&nbsp;Nicolas Brodusch,&nbsp;Raynald Gauvin","doi":"10.1111/jmi.70018","DOIUrl":"10.1111/jmi.70018","url":null,"abstract":"<p>This manuscript presents a comparative analysis of two software packages, MC X-ray and PENELOPE, focusing on their accuracy and efficiency in simulating k-ratios for binary compounds and comparing their spectra with experimental data for pure elements and compounds. Based on the Pouchou database, MC X-ray slightly outperforms PENELOPE in k-ratio calculations, achieving a root mean square error (RMSE) of 2.71% with a standard deviation of 0.027, compared to 2.87% with a standard deviation of 0.053. Discrepancies between the two programs emerge at lower beam energies (3 and 5 keV) when comparing simulated spectra with experimental data; however, at higher energies (20 and 30 keV), both software packages exhibit consistent and reliable performance across a range of atomic numbers. While both tools are effective for analysing homogeneous bulk samples, MC X-ray offers significant advantages in processing speed and user-friendliness. This study underscores the strengths and limitations of each package, providing valuable insights for researchers engaged in X-ray simulation and microanalysis.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"300 1","pages":"134-145"},"PeriodicalIF":1.9,"publicationDate":"2025-08-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.70018","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144789380","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A novel experimental platform to monitor solid/water interfaces under freeze-thaw cycles.","authors":"Chiara Recalcati, Rossella Yivlialin, Lamberto Duò, Alberto Guadagnini, Gianlorenzo Bussetti","doi":"10.1111/jmi.70017","DOIUrl":"https://doi.org/10.1111/jmi.70017","url":null,"abstract":"<p><p>We design and implement an original experimental platform resting on Atomic Force Microscopy (AFM) to capture nanoscale insights into key characteristics of solid/water interfaces subject to freeze-thaw conditions. The work is motivated by the observation that freezing and thawing underpin a variety of processes in the context of, e.g., climate and material sciences or cryobiology. Despite their key role, fundamental processes driving freezing and thawing are still elusive and their direct documentation is still challenging. This primarily stems from operational difficulties in replicating these processes under laboratory conditions, as well as constraints of current technology in matching temporal and spatial scales at which these phenomena take place. Here, we propose an experimental strategy to control freezing at solid/water interfaces while maintaining the bulk water as liquid. Our platform favors operational simplicity and can be integrated with any tip-scanning AFM. The strength of our set-up is assessed upon experiments performed on Highly Oriented Pyrolytic Graphite (HOPG) as a model substrate.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":" ","pages":""},"PeriodicalIF":1.9,"publicationDate":"2025-08-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144784477","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Ultrastructural expansion microscopy reveals unexpected levels of glycosome heterogeneity in African trypanosomes.","authors":"Heidi Anderson, Rhonda Reigers Powell, Meredith Teilhet Morris","doi":"10.1111/jmi.70019","DOIUrl":"10.1111/jmi.70019","url":null,"abstract":"<p><p>Kinetoplastid parasites include several species. Trypanosoma brucei causes African sleeping sickness in humans and a wasting disease nagana in livestock. Trypanosoma cruzi is the causative agent of Chagas disease and Leishmania species cause leishmaniasis, which can present with visceral, cutaneous, or mucocutaneous symptoms. All kinetoplastids harbour specialised peroxisomes called glycosomes, so named because most of the glycolytic pathway that is cytosolic in other eukaryotes is localised to these organelles. Glycosomes lack DNA and are essential for parasite viability. Despite their name, glycosomes also house enzymes involved in diverse pathways, including the pentose phosphate pathway, ether lipid biosynthesis, purine salvage, and sugar nucleotide biosynthesis. The degree to which these biochemical pathways localise together within the same organelle or to different glycosome populations is unclear. Biochemical fractionations and imaging data strongly suggest that glycosomes are heterogeneous in composition and that even within a single parasite, there are different glycosome populations. Until recently, we lacked the technology to systematically characterise glycosome populations within parasites. Glycosome morphology, composition, and localisation have historically been studied using widefield fluorescence and electron microscopy (EM). While EM can resolve individual organelles, it is extremely low throughput and requires specialised expertise and equipment. Widefield fluorescence imaging is higher throughput and more accessible. However, the small size of T. brucei cells, which are ∼20 µM in length and 3-5 µM in width, and glycosomes (100 nm in diameter) place these organelles below the resolution limits of standard microscopy and require super-resolution techniques to be resolved. These resolution issues are compounded by the cytoplasm's crowded nature, making it hard to discern individual organelles from each other. To overcome this, we leveraged recent advances in super-resolution microscopy, including a method called Ultrastructure Expansion Microscopy (U-ExM) combined with confocal imaging and LIGHTNING™ deconvolution to optimise the resolution of individual glycosomes. We found that antibodies against two different glycosome marker proteins (aldolase and GAPDH) exhibit discrete staining patterns. This high-resolution approach also revealed that glycosome morphology varies between monomorphic parasites that cannot complete the lifecycle and pleomorphic parasites that can, and is dynamically influenced by extracellular conditions, such as glucose availability, underscoring the adaptability of T. brucei's compartmentalisation to environmental changes.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":" ","pages":""},"PeriodicalIF":1.9,"publicationDate":"2025-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144753627","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Analytical solution of the classical Rayleigh length definition, including truncation at arbitrary values","authors":"Aufried Lenferink,&nbsp;Cees Otto","doi":"10.1111/jmi.70016","DOIUrl":"10.1111/jmi.70016","url":null,"abstract":"<p>We present the analytical solution to the diffraction integral that describes the Rayleigh length for a focused Gaussian beam with any value of a spherical truncating aperture. This exact solution is in precise agreement with numerical calculations for the light distribution in the near focal area. The solution arises under assumption of the paraxial approximation, which also provides the basis for the classical Rayleigh length definition. It will be shown that the non-paraxial regime can be included by adding an empirical term (<i>C</i>_np) to the solution of the diffraction integral. This extends the validity of the expression to high numerical apertures (<i>NA</i>) up to <i>n</i> times 0.95, with <i>n</i> being the refractive index of the immersion medium. Thus, the entire practical range of NA, encountered in optical microscopy, is covered with a calculated error of less than 0.4% in the non-paraxial limit. This theoretical result is important in the design of optical instrumentation, where overall light efficiency in excitation and detection and spatial resolution must be optimised together.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"300 1","pages":"124-133"},"PeriodicalIF":1.9,"publicationDate":"2025-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.70016","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144753626","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}