Journal of microscopy最新文献

{"title":"Annotation and automated segmentation of single-molecule localisation microscopy data","authors":"Oliver Umney,&nbsp;Joanna Leng,&nbsp;Gianluca Canettieri,&nbsp;Natalia A. Riobo-Del Galdo,&nbsp;Hayley Slaney,&nbsp;Philip Quirke,&nbsp;Michelle Peckham,&nbsp;Alistair Curd","doi":"10.1111/jmi.13349","DOIUrl":"10.1111/jmi.13349","url":null,"abstract":"<p>Single Molecule Localisation Microscopy (SMLM) is becoming a widely used technique in cell biology. After processing the images, the molecular localisations are typically stored in a table as <i>xy</i> (or <i>xyz</i>) coordinates, with additional information, such as number of photons, etc. This set of coordinates can be used to generate an image to visualise the molecular distribution, for example, a 2D or 3D histogram of localisations. Many different methods have been devised to analyse SMLM data, among which cluster analysis of the localisations is popular. However, it can be useful to first segment the data, to extract the localisations in a specific region of a cell or in individual cells, prior to downstream analysis. Here we describe a pipeline for annotating localisations in an SMLM dataset in which we compared membrane segmentation approaches, including Otsu thresholding and machine learning models, and subsequent cell segmentation. We used an SMLM dataset derived from dSTORM images of sectioned cell pellets, stained for the membrane proteins EGFR (epidermal growth factor receptor) and EREG (epiregulin) as a test dataset. We found that a Cellpose model retrained on our data performed the best in the membrane segmentation task, allowing us to perform downstream cluster analysis of membrane versus cell interior localisations. We anticipate this will be generally useful for SMLM analysis.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"296 3","pages":"214-226"},"PeriodicalIF":1.5,"publicationDate":"2024-08-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.13349","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141875098","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":"Surpassing light inhomogeneities in structured-illumination microscopy with FlexSIM","authors":"Emmanuel Soubies,&nbsp;Alejandro Nogueron,&nbsp;Florence Pelletier,&nbsp;Thomas Mangeat,&nbsp;Christophe Leterrier,&nbsp;Michael Unser,&nbsp;Daniel Sage","doi":"10.1111/jmi.13344","DOIUrl":"10.1111/jmi.13344","url":null,"abstract":"<p>Super-resolution structured-illumination microscopy (SIM) is a powerful technique that allows one to surpass the diffraction limit by up to a factor two. Yet, its practical use is hampered by its sensitivity to imaging conditions which makes it prone to reconstruction artefacts. In this work, we present FlexSIM, a <i>flexible</i> SIM reconstruction method capable to handle highly challenging data. Specifically, we demonstrate the ability of FlexSIM to deal with the distortion of patterns, the high level of noise encountered in live imaging, as well as out-of-focus fluorescence. Moreover, we show that FlexSIM achieves state-of-the-art performance over a variety of open SIM datasets.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"296 1","pages":"94-106"},"PeriodicalIF":1.5,"publicationDate":"2024-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.13344","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141620144","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.13201","DOIUrl":"10.1111/jmi.13201","url":null,"abstract":"","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"295 2","pages":"83-84"},"PeriodicalIF":1.5,"publicationDate":"2024-07-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.13201","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141608517","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":"Deep learning for quantifying spatial patterning and formation process of early differentiated human-induced pluripotent stem cells with micropattern images","authors":"Slo-Li Chu,&nbsp;Kuniya Abe,&nbsp;Hideo Yokota,&nbsp;Dooseon Cho,&nbsp;Yohei Hayashi,&nbsp;Ming-Dar Tsai","doi":"10.1111/jmi.13346","DOIUrl":"10.1111/jmi.13346","url":null,"abstract":"<p>Micropatterning is reliable method for quantifying pluripotency of human-induced pluripotent stem cells (hiPSCs) that differentiate to form a spatial pattern of sorted, ordered and nonoverlapped three germ layers on the micropattern. In this study, we propose a deep learning method to quantify spatial patterning of the germ layers in the early differentiation stage of hiPSCs using micropattern images. We propose decoding and encoding U-net structures learning labelled Hoechst (DNA-stained) hiPSC regions with corresponding Hoechst and bright-field micropattern images to segment hiPSCs on Hoechst or bright-field images. We also propose a U-net structure to extract extraembryonic regions on a micropattern, and an algorithm to compares intensities of the fluorescence images staining respective germ-layer cells and extract their regions. The proposed method thus can quantify the pluripotency of a hiPSC line with spatial patterning including cell numbers, areas and distributions of germ-layer and extraembryonic cells on a micropattern, and reveal the formation process of hiPSCs and germ layers in the early differentiation stage by segmenting live-cell bright-field images. In our assay, the cell-number accuracy achieved 86% and 85%, and the cell region accuracy 89% and 81% for segmenting Hoechst and bright-field micropattern images, respectively. Applications to micropattern images of multiple hiPSC lines, micropattern sizes, groups of markers, living and fixed cells show the proposed method can be expected to be a useful protocol and tool to quantify pluripotency of a new hiPSC line before providing it to the scientific community.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"296 1","pages":"79-93"},"PeriodicalIF":1.5,"publicationDate":"2024-07-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.13346","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141590537","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":"The wavenumber linearisation without calibration device for spectral-domain optical coherence tomography","authors":"Xiupin Wu,&nbsp;Wanrong Gao,&nbsp;Zhiyuan Qiu,&nbsp;Chunyou Wang","doi":"10.1111/jmi.13345","DOIUrl":"10.1111/jmi.13345","url":null,"abstract":"<p>The wavenumber nonlinearity leads to blurred reconstructed images in spectral-domain optical coherence tomography (SDOCT). In this work, a wavenumber-linearisation method without calibration devices is presented, based on the fact that the difference between the phases of adjacent peak and valley points is equal to <span></span><math>\u0000 <semantics>\u0000 <mi>π</mi>\u0000 <annotation>$pi $</annotation>\u0000 </semantics></math>. The theoretical model is derived, and the efficacy of the method was proven by acquiring SDOCT data from TiO₂ phantom and zebrafish. The results exhibit the superior performance of our method. Compared with the linear phase-based method, the resolution could be improved at least a factor of 2. Compared with the polynomial fitting method, the resolution could also be improved by nearly half.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"296 1","pages":"70-78"},"PeriodicalIF":1.5,"publicationDate":"2024-07-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141563562","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":"Unveiling the limits of precision in iterative MINFLUX","authors":"Carlas Smith,&nbsp;Dylan Kalisvaart,&nbsp;Kirti Prakash","doi":"10.1111/jmi.13338","DOIUrl":"10.1111/jmi.13338","url":null,"abstract":"<p>In single-molecule microscopy, a big question is how precisely we can estimate the location of a single molecule. Our research shows that by using iterative localisation microscopy and factoring in the prior information, we can boost precision and reduce the number of photons needed. Leveraging the Van Trees inequality aids in determining the optimal precision achievable. Our approach holds promise for wider application in discerning the optimal precision across diverse imaging scenarios, encompassing various illumination strategies, point spread functions and overarching control methodologies.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"296 2","pages":"129-132"},"PeriodicalIF":1.5,"publicationDate":"2024-07-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.13338","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141563563","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":"Quantifying superimposed protein flow dynamics in live cells using spatial filtering and spatiotemporal image correlation spectroscopy.","authors":"Rodrigo A Migueles-Ramírez, Alessandra Cambi, Arnold Hayer, Paul W Wiseman, Koen van den Dries","doi":"10.1111/jmi.13342","DOIUrl":"https://doi.org/10.1111/jmi.13342","url":null,"abstract":"<p><p>Flow or collective movement is a frequently observed phenomenon for many cellular components including the cytoskeletal proteins actin and myosin. To study protein flow in living cells, we and others have previously used spatiotemporal image correlation spectroscopy (STICS) analysis on fluorescence microscopy image time series. Yet, in cells, multiple protein flows often occur simultaneously on different scales resulting in superimposed fluorescence intensity fluctuations that are challenging to separate using STICS. Here, we exploited the characteristic that distinct protein flows often occur at different spatial scales present in the image series to disentangle superimposed protein flow dynamics. We employed a newly developed and an established spatial filtering algorithm to alternatively accentuate or attenuate local image intensity heterogeneity across different spatial scales. Subsequently, we analysed the spatially filtered time series with STICS, allowing the quantification of two distinct superimposed flows within the image time series. As a proof of principle of our analysis approach, we used simulated fluorescence intensity fluctuations as well as time series of nonmuscle myosin II in endothelial cells and actin-based podosomes in dendritic cells and revealed simultaneously occurring contiguous and noncontiguous flow dynamics in each of these systems. Altogether, this work extends the application of STICS for the quantification of multiple protein flow dynamics in complex biological systems including the actomyosin cytoskeleton.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":" ","pages":""},"PeriodicalIF":1.5,"publicationDate":"2024-07-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141498280","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":"Study on line-scan profile for a trapezoid line under varying sample temperatures through Monte–Carlo simulation","authors":"Peng Zhang","doi":"10.1111/jmi.13343","DOIUrl":"10.1111/jmi.13343","url":null,"abstract":"<p>This study investigates the influence of the sample inherent temperature on the line-scan profile for a silicon trapezoid line with different sidewall angles by Monte–Carlo simulation. This study demonstrates that the profile varies with temperature, particularly focusing on the ‘shoulder’, which becomes more pronounced with larger sidewall angles. The contrast of the secondary electron profile increases at low primary electron energy but decreases at relatively high PE energy as the temperature rises. The trend of the backscattering electron profile is similar but less noticeable. The underlying mechanism is discussed in detail. This study has potential to provide valuable insights into thermometry in nanostructures using SEMs.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"296 1","pages":"63-69"},"PeriodicalIF":1.5,"publicationDate":"2024-06-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141457475","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":"Epifluorescence microscopy study of a quadruple node of triple junctions of grain boundaries in a Eu2+-decorated highly textured composite of (Cl, Br)(K, Rb) and I(K, Rb) solid solutions","authors":"A. E. Cordero-Borboa,&nbsp;R. Unda-Angeles","doi":"10.1111/jmi.13341","DOIUrl":"10.1111/jmi.13341","url":null,"abstract":"<p>The structural nature and geometry, as well as the lattice-relative orientation, of an arrangement of crystal defects in a highly textured Eu²⁺-doped composite of two alkali-halide solid solutions was studied by epifluorescence microscopy (EFM) using the doping ion as a fluorochrome. A three-dimensional reconstruction and a skeleton type model, as built from a sequence of EFM images of different optical cross-sections of this arrangement, are presented. Structurally, this arrangement is a quadruple node (QN) of triple junctions of grain boundaries. The QN core geometry is that of a tetragonal tristetrahedron (TTTH), centred at the QN site, whose tetrahedron vertices and edges are on the QN triple junctions and grain boundaries, respectively, whereas the tristetrahedron tetragonal axis is nearly parallel to the lattice [001]-axis. The measured values of the angles between triple junctions and between the grain boundaries forming them are reported. The distinct chemical compositions of the composite solid solutions are discussed to be responsible, in last instance, for the tristetrahedron departure from a cubic configuration. Collaterally, certain families of translationally periodic almost-parallel (TPAP)-wall-like regions which consist of TPAP-columns of TPAP-spindle-like singularities, as well as certain zigzag arrays of columns of this like, existing into the QN grains, are reported to be observed. Three-dimensional reconstructions of typical individuals of these families and arrays as well as of their constituent parts are presented and geometrically analysed. These families and arrays are discussed to be families of tilt subboundaries, whose constituent dislocations are decorated by cylindrical second-phase europium di-halide precipitates, and regularly faceted tilt subboundaries, respectively. Crystal growing and sample preparation, composite structural characterisation by powder and single-slab X-ray diffraction (PXRD and SSXRD, respectively), microscopy and fluorescence-cube unit optics, image processing, electronic three-dimensional reconstruction and measuring methodologies, are all described in detail.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"296 1","pages":"48-62"},"PeriodicalIF":1.5,"publicationDate":"2024-06-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141427080","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":"TOC - Issue Information","authors":"","doi":"10.1111/jmi.13200","DOIUrl":"https://doi.org/10.1111/jmi.13200","url":null,"abstract":"","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"295 1","pages":"1-2"},"PeriodicalIF":2.0,"publicationDate":"2024-06-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.13200","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141424797","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}