Pub Date : 2026-03-01Epub Date: 2025-12-23DOI: 10.1016/j.ymeth.2025.12.012
Zakia Zinat Choudhury , Samiran Dey , Haoming Wang , Sean Coffey , Tapabrata Chakraborti , Brendan McCane
Medical imaging is fundamental to cardiovascular diagnostics, with modalities such as Transthoracic Echocardiography (TTE) and Cardiac Magnetic Resonance (CMR) offering complementary strengths. TTE provides real-time, non-invasive visualization of cardiac function but is often limited by operator dependency and incomplete views. In contrast, CMR delivers comprehensive, high-resolution structural assessments, although it comes with greater time and cost burdens. To address these limitations, this study explores cross-modal generative modeling techniques for synthesizing CMR-like images directly from TTE. We propose a novel architecture that combines a UNet backbone with a vision transformer, utilizing the UNet for feature extraction and the transformer for global attention to improve image synthesis quality. Quantitative and qualitative evaluations demonstrate the model’s ability to produce realistic and anatomically consistent CMR images, with strong potential to improve diagnostic accuracy and clinical decision-making across multiple image modalities.
{"title":"Cross-modal image generation with uncertainty quantification from echocardiogram to MRI","authors":"Zakia Zinat Choudhury , Samiran Dey , Haoming Wang , Sean Coffey , Tapabrata Chakraborti , Brendan McCane","doi":"10.1016/j.ymeth.2025.12.012","DOIUrl":"10.1016/j.ymeth.2025.12.012","url":null,"abstract":"<div><div>Medical imaging is fundamental to cardiovascular diagnostics, with modalities such as Transthoracic Echocardiography (TTE) and Cardiac Magnetic Resonance (CMR) offering complementary strengths. TTE provides real-time, non-invasive visualization of cardiac function but is often limited by operator dependency and incomplete views. In contrast, CMR delivers comprehensive, high-resolution structural assessments, although it comes with greater time and cost burdens. To address these limitations, this study explores cross-modal generative modeling techniques for synthesizing CMR-like images directly from TTE. We propose a novel architecture that combines a UNet backbone with a vision transformer, utilizing the UNet for feature extraction and the transformer for global attention to improve image synthesis quality. Quantitative and qualitative evaluations demonstrate the model’s ability to produce realistic and anatomically consistent CMR images, with strong potential to improve diagnostic accuracy and clinical decision-making across multiple image modalities.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"247 ","pages":"Pages 1-11"},"PeriodicalIF":4.3,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145831859","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
DNA Fluorescence In Situ Hybridization (DNA FISH) is an essential technique to study chromosome biology and genetics, enabling precise visualization of specific genomic loci to study structural abnormalities, gene mapping, and chromosomal rearrangements. High-Throughput Imaging (HTI) can automate the analysis of DNA FISH chromosome images, but the accurate and automated segmentation of mitotic chromosomes and simultaneous colocalization of DNA FISH signals remains a challenge. While several commercial automated karyotyping tools partially solve these issues, open-source software that effectively combines robust chromosome segmentation with comprehensive colocalization analysis capabilities remains necessary. To address this unmet need, we developed MetaChrome, an open-source software platform built around a graphical user interface and explicitly designed for automated metaphase chromosome analysis. MetaChrome leverages fine-tuned deep learning models to automate metaphase chromosome segmentation, together with colocalization analysis of chromosome-specific FISH probes and immunofluorescent-labeled proteins. Importantly, MetaChrome achieves enhanced segmentation accuracy compared to traditional image processing methods by adopting a Cellpose segmentation model fine-tuned with manually annotated metaphase chromosome datasets. The fine-tuned model ensures the precise assignment of DNA FISH spots to individual chromosomes in an automated manner. This facilitates rapid identification of chromosomal abnormalities, reduces human error, and advances high-throughput chromosome analysis workflows, addressing a key bottleneck in chromosome biology research.
{"title":"MetaChrome: an open-source, user-friendly tool for automated metaphase chromosome analysis","authors":"Md Abdul Kader Sagar , Yamini Dalal , Gianluca Pegoraro , Ganesan Arunkumar","doi":"10.1016/j.ymeth.2025.12.013","DOIUrl":"10.1016/j.ymeth.2025.12.013","url":null,"abstract":"<div><div>DNA Fluorescence In Situ Hybridization (DNA FISH) is an essential technique to study chromosome biology and genetics, enabling precise visualization of specific genomic loci to study structural abnormalities, gene mapping, and chromosomal rearrangements. High-Throughput Imaging (HTI) can automate the analysis of DNA FISH chromosome images, but the accurate and automated segmentation of mitotic chromosomes and simultaneous colocalization of DNA FISH signals remains a challenge. While several commercial automated karyotyping tools partially solve these issues, open-source software that effectively combines robust chromosome segmentation with comprehensive colocalization analysis capabilities remains necessary. To address this unmet need, we developed MetaChrome, an open-source software platform built around a graphical user interface and explicitly designed for automated metaphase chromosome analysis. MetaChrome leverages fine-tuned deep learning models to automate metaphase chromosome segmentation, together with colocalization analysis of chromosome-specific FISH probes and immunofluorescent-labeled proteins. Importantly, MetaChrome achieves enhanced segmentation accuracy compared to traditional image processing methods by adopting a Cellpose segmentation model fine-tuned with manually annotated metaphase chromosome datasets. The fine-tuned model ensures the precise assignment of DNA FISH spots to individual chromosomes in an automated manner. This facilitates rapid identification of chromosomal abnormalities, reduces human error, and advances high-throughput chromosome analysis workflows, addressing a key bottleneck in chromosome biology research.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"247 ","pages":"Pages 12-24"},"PeriodicalIF":4.3,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145877422","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-03-01Epub Date: 2026-01-27DOI: 10.1016/j.ymeth.2026.01.009
Yevheniia Kharkivska , Olha Shkel , Yun Kyung Kim
Syntenin is a multifunctional PDZ-domain adaptor protein that orchestrates membrane trafficking, cytoskeletal remodeling, and exosome biogenesis. Initially identified as a syndecan-binding molecule, syntenin has since emerged as a central hub connecting membrane receptors to intracellular signaling pathways that regulate adhesion, motility, immune signaling, and cellular plasticity. While extensively studied in cancer and neural development, recent discoveries reveal that a wide range of viruses exploit syntenin to facilitate their replication, assembly, or dissemination. This review consolidates current evidence across diverse viral infections to elucidate the molecular mechanisms underlying the interaction between syntenin and viruses. Coronaviruses utilize syntenin to link PDZ-binding motifs to p38 MAPK-driven inflammation and endosomal entry. Papillomaviruses and Epstein–Barr virus hijack the CD63-syntenin-ALIX complex to control vesicle-mediated trafficking. Hepatitis C virus employs it to secrete E2-coated, antibody-resistant exosomes. Dengue virus harnesses its mosquito homolog AeSyntenin to package sfRNA for transmission. Human T-cell leukemia virus type 1 employs its Tax-1 oncoprotein to bind the PDZ domains of syntenin, remodel extracellular vesicle cargo, and promote viral spread. In contrast, during human immunodeficiency virus infection, syntenin restricts viral fusion at the plasma membrane, though the nucleocapsid mimics its PDZ tandem to promote virion release. Collectively, these findings establish syntenin as a dynamic regulator at the host–virus interface, capable of exerting both proviral and antiviral effects. Emerging pharmacological strategies targeting syntenin PDZ domains further underscore its potential as a broad-spectrum, host-directed antiviral target.
{"title":"Syntenins at the crossroads of host–virus interactions","authors":"Yevheniia Kharkivska , Olha Shkel , Yun Kyung Kim","doi":"10.1016/j.ymeth.2026.01.009","DOIUrl":"10.1016/j.ymeth.2026.01.009","url":null,"abstract":"<div><div>Syntenin is a multifunctional PDZ-domain adaptor protein that orchestrates membrane trafficking, cytoskeletal remodeling, and exosome biogenesis. Initially identified as a syndecan-binding molecule, syntenin has since emerged as a central hub connecting membrane receptors to intracellular signaling pathways that regulate adhesion, motility, immune signaling, and cellular plasticity. While extensively studied in cancer and neural development, recent discoveries reveal that a wide range of viruses exploit syntenin to facilitate their replication, assembly, or dissemination. This review consolidates current evidence across diverse viral infections to elucidate the molecular mechanisms underlying the interaction between syntenin and viruses. Coronaviruses utilize syntenin to link PDZ-binding motifs to p38 MAPK-driven inflammation and endosomal entry. Papillomaviruses and Epstein–Barr virus hijack the CD63-syntenin-ALIX complex to control vesicle-mediated trafficking. Hepatitis C virus employs it to secrete E2-coated, antibody-resistant exosomes. Dengue virus harnesses its mosquito homolog AeSyntenin to package sfRNA for transmission. Human T-cell leukemia virus type 1 employs its Tax-1 oncoprotein to bind the PDZ domains of syntenin, remodel extracellular vesicle cargo, and promote viral spread. In contrast, during human immunodeficiency virus infection, syntenin restricts viral fusion at the plasma membrane, though the nucleocapsid mimics its PDZ tandem to promote virion release. Collectively, these findings establish syntenin as a dynamic regulator at the host–virus interface, capable of exerting both proviral and antiviral effects. Emerging pharmacological strategies targeting syntenin PDZ domains further underscore its potential as a broad-spectrum, host-directed antiviral target.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"247 ","pages":"Pages 175-183"},"PeriodicalIF":4.3,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"146074260","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-03-01Epub Date: 2026-01-08DOI: 10.1016/j.ymeth.2026.01.004
Víctor Pola-Véliz , Sebastián B. Arredondo , Yennyfer Arancibia , Juan Ahumada , Sebastián Estay , Nicole Vidal , Paola A. Haeger , Marco Fuenzalida , Lorena Varela-Nallar , Fernando J. Bustos , Martin Montecino , Brigitte van Zundert
Memory formation activates a relatively sparse population of engram cells that store long-term memories. Changes in the epigenetic landscape and 3D chromatin architecture have been proposed as key candidate regulators of transcriptional waves that control gene expression in engram cells; however, isolating chromatin efficiently from engram cells has remained challenging. Double-transgenic Targeted Recombination in Active Populations (dTRAP) mice have enabled indelible EYFP labeling of hippocampal engram cells expressing the immediate-early gene (IEG) Arc when ArcCreERT2 mice are crossed with R26R-STOP-floxed-EYFP mice and exposed to learning paradigms. A major limitation of dTRAP mice is that labeling of activated hippocampal Arc+ neurons with soluble EYFP compromises the efficiency of fluorescence-activated nuclear sorting (FANS) of engram nuclei, and hence isolation of chromatin. Here, we used viral-mediated delivery of GFP-KASH (AAV-PHP.eB-FLEX-EGFP-KASH) to ArcCreERT2 mice -generating vkTRAP mice- to enable precise and robust endogenous perinuclear fluorescent tagging of activated hippocampal neurons following contextual fear conditioning (CFC). At 24 h post-CFC (24 h-CFC), vkTRAP mice exhibited a robust freezing behavior. Electrophysiological recordings in CA1 hippocampal slices showed occluded long-term potentiation (LTP). Efficient FANS-based isolation of hippocampal engram nuclei enabled chromatin immunoprecipitation (ChIP) assays (detecting H3K4me3, H3K9ac and H3K27ac) at promoters of immediate-early (Egr1) and plasticity-related (Dlg4/PSD95) genes. Expression peaks of both Egr1 and Dlg4/PSD95 transcripts during memory acquisition (1 h-CFC) and consolidation (24 h-CFC) were accompanied by active epigenetic histone mark profiles. We conclude that vkTRAP provides a robust model to study epigenomic regulation in engram cells.
{"title":"Viral-mediated fluorescent labelling of activated hippocampal memory engrams to study epigenetic dynamics associated with gene expression","authors":"Víctor Pola-Véliz , Sebastián B. Arredondo , Yennyfer Arancibia , Juan Ahumada , Sebastián Estay , Nicole Vidal , Paola A. Haeger , Marco Fuenzalida , Lorena Varela-Nallar , Fernando J. Bustos , Martin Montecino , Brigitte van Zundert","doi":"10.1016/j.ymeth.2026.01.004","DOIUrl":"10.1016/j.ymeth.2026.01.004","url":null,"abstract":"<div><div>Memory formation activates a relatively sparse population of engram cells that store long-term memories. Changes in the epigenetic landscape and 3D chromatin architecture have been proposed as key candidate regulators of transcriptional waves that control gene expression in engram cells; however, isolating chromatin efficiently from engram cells has remained challenging. Double-transgenic <em>T</em>argeted <em>R</em>ecombination in <em>A</em>ctive <em>P</em>opulations (dTRAP) mice have enabled indelible EYFP labeling of hippocampal engram cells expressing the immediate-early gene (IEG) <em>Arc</em> when ArcCreER<sup>T2</sup> mice are crossed with R26R-STOP-floxed-EYFP mice and exposed to learning paradigms. A major limitation of dTRAP mice is that labeling of activated hippocampal Arc<sup>+</sup> neurons with soluble EYFP compromises the efficiency of fluorescence-activated nuclear sorting (FANS) of engram nuclei, and hence isolation of chromatin. Here, we used viral-mediated delivery of GFP-KASH (AAV-PHP.eB-FLEX-EGFP-KASH) to ArcCreER<sup>T2</sup> mice -generating vkTRAP mice- to enable precise and robust endogenous perinuclear fluorescent tagging of activated hippocampal neurons following contextual fear conditioning (CFC). At 24 h post-CFC (24 h-CFC), vkTRAP mice exhibited a robust freezing behavior. Electrophysiological recordings in CA1 hippocampal slices showed occluded long-term potentiation (LTP). Efficient FANS-based isolation of hippocampal engram nuclei enabled chromatin immunoprecipitation (ChIP) assays (detecting H3K4me3, H3K9ac and H3K27ac) at promoters of immediate-early (<em>Egr1</em>) and plasticity-related (<em>Dlg4</em>/PSD95) genes. Expression peaks of both <em>Egr1</em> and <em>Dlg4/</em>PSD95 transcripts during memory acquisition (1 h-CFC) and consolidation (24 h-CFC) were accompanied by active epigenetic histone mark profiles. We conclude that vkTRAP provides a robust model to study epigenomic regulation in engram cells.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"247 ","pages":"Pages 85-94"},"PeriodicalIF":4.3,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145942002","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-03-01Epub Date: 2026-01-24DOI: 10.1016/j.ymeth.2026.01.008
Angga Hermawan , Asmi Aris , Munawar Khalil , Ni Luh Wulan Septiani , Teti Estiasih , Hamidie Ronald Daniel Ray , Miguel Palma , Widiastuti Setyaningsih
Nanomaterial-enabled electrochemical sensors are nearing the performance and practicality needed for routine, on-site monitoring of phenolic compounds in foods and beverages. Advances in nanomaterial dimensionality and hybrid architectures, from atomically doped nanoparticles and zero-dimensional clusters to two- and three-dimensional porous frameworks, have enhanced electron-transfer kinetics, expanded electroactive surface area, and enabled more selective surface chemistries. These gains align with progress in molecular recognition using enzymes, aptamers, molecularly imprinted polymers, and permselective antifouling coatings, as well as electrode-engineering strategies that translate nanoscale activity into reliable printed-electrodes. Although laboratory detection limits are often impressive (micromolar to low-nanomolar in controlled media), challenges remain in reproducibility, shelf life, and performance in complex matrices such as wine, olive oil, and fermented foods. Closing these gaps requires integrated solutions that unite printable, stable nanomaterial inks with simple on-cartridge sample conditioning, modular recognition layers, and robust on-board calibration and data-handling routines. To enable practical deployment, we propose a development pathway focused on scalable manufacturing and quality control of nanomaterial inks and electrodes, harmonized validation against chromatographic reference methods, durable antifouling and self-cleaning strategies, and an ecosystem approach that uses smartphone connectivity and cloud analytics to convert electrochemical signals into traceable, defensible decisions for industry, regulators, and consumers.
{"title":"Nanomaterial-based electrochemical sensors for phenolic antioxidants in foods and beverages: From design to device translation","authors":"Angga Hermawan , Asmi Aris , Munawar Khalil , Ni Luh Wulan Septiani , Teti Estiasih , Hamidie Ronald Daniel Ray , Miguel Palma , Widiastuti Setyaningsih","doi":"10.1016/j.ymeth.2026.01.008","DOIUrl":"10.1016/j.ymeth.2026.01.008","url":null,"abstract":"<div><div>Nanomaterial-enabled electrochemical sensors are nearing the performance and practicality needed for routine, on-site monitoring of phenolic compounds in foods and beverages. Advances in nanomaterial dimensionality and hybrid architectures, from atomically doped nanoparticles and zero-dimensional clusters to two- and three-dimensional porous frameworks, have enhanced electron-transfer kinetics, expanded electroactive surface area, and enabled more selective surface chemistries. These gains align with progress in molecular recognition using enzymes, aptamers, molecularly imprinted polymers, and permselective antifouling coatings, as well as electrode-engineering strategies that translate nanoscale activity into reliable printed-electrodes. Although laboratory detection limits are often impressive (micromolar to low-nanomolar in controlled media), challenges remain in reproducibility, shelf life, and performance in complex matrices such as wine, olive oil, and fermented foods. Closing these gaps requires integrated solutions that unite printable, stable nanomaterial inks with simple on-cartridge sample conditioning, modular recognition layers, and robust on-board calibration and data-handling routines. To enable practical deployment, we propose a development pathway focused on scalable manufacturing and quality control of nanomaterial inks and electrodes, harmonized validation against chromatographic reference methods, durable antifouling and self-cleaning strategies, and an ecosystem approach that uses smartphone connectivity and cloud analytics to convert electrochemical signals into traceable, defensible decisions for industry, regulators, and consumers.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"247 ","pages":"Pages 132-160"},"PeriodicalIF":4.3,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"146049740","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-22DOI: 10.1016/j.ymeth.2026.02.012
Chau Ha Pham, Birgitte Kasin Hønsvall, Nivedhitha Jothinarayanan, Erik Andrew Johannessen, Bao Quoc Ta
Environmental DNA (eDNA) analysis is a non-invasive method for discovering and identifying rare and endangered species in a variety of ecosystems including aquatic environments. However, conventional eDNA analysis tends to be time consuming and labor-intensive, which limits their efficiency. In this study, a novel eDNA capture system is made from stacked layers of membranes with a binding pad of borosilicate glass paper embedded with a triple layer of high molecular weight chitosan and silica nanoparticles. The system is designed to enhance DNA capture efficiency and is part of a portable system for on-site eDNA extraction that measures approx. 22 x 70 mm using sample volumes up to 3 mL and enables DNA extraction under 10 min. The DNA recovery rate of 86% is comparable to silica bead-based laboratory methods at a significantly lower cost, and the Ct values from the nucleic acid amplification step are reached within 18 to 36 min. Real water samples were tested, yielding results comparable to those obtained with the silica bead-based laboratory methods. Thus, enables downstream applications, such as species detection through a portable system that has the potential to improve the accessibility of eDNA analysis for effective environmental monitoring and conservation practices.
{"title":"An Affordable paper based platform for eDNA Filtration, Concentration, and nucleic acid extraction.","authors":"Chau Ha Pham, Birgitte Kasin Hønsvall, Nivedhitha Jothinarayanan, Erik Andrew Johannessen, Bao Quoc Ta","doi":"10.1016/j.ymeth.2026.02.012","DOIUrl":"10.1016/j.ymeth.2026.02.012","url":null,"abstract":"<p><p>Environmental DNA (eDNA) analysis is a non-invasive method for discovering and identifying rare and endangered species in a variety of ecosystems including aquatic environments. However, conventional eDNA analysis tends to be time consuming and labor-intensive, which limits their efficiency. In this study, a novel eDNA capture system is made from stacked layers of membranes with a binding pad of borosilicate glass paper embedded with a triple layer of high molecular weight chitosan and silica nanoparticles. The system is designed to enhance DNA capture efficiency and is part of a portable system for on-site eDNA extraction that measures approx. 22 x 70 mm using sample volumes up to 3 mL and enables DNA extraction under 10 min. The DNA recovery rate of 86% is comparable to silica bead-based laboratory methods at a significantly lower cost, and the Ct values from the nucleic acid amplification step are reached within 18 to 36 min. Real water samples were tested, yielding results comparable to those obtained with the silica bead-based laboratory methods. Thus, enables downstream applications, such as species detection through a portable system that has the potential to improve the accessibility of eDNA analysis for effective environmental monitoring and conservation practices.</p>","PeriodicalId":390,"journal":{"name":"Methods","volume":" ","pages":"77-86"},"PeriodicalIF":4.3,"publicationDate":"2026-02-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147281528","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-01Epub Date: 2025-12-18DOI: 10.1016/j.ymeth.2025.12.007
Jacob E. Labovitz , Patrick Kulaga , Eric M. DuBois, Kunyu Li, Timothy M. O’Shea
Traumatic injury to the healthy central nervous system (CNS) causes mechanical tissue damage that results in localized cell death and blood–brain-barrier (BBB) disruption. CNS tissue damage stimulates a multicellular wound response to limit the extent of damage but fails to reestablish the normal function of injured tissue. There is strong interest in developing new strategies to augment regeneration after CNS injury. To enable therapy development, reliable assays to screen and identify molecular approaches to augment glial-based wound responses over fibrotic scarring are needed. Scratch assays, which involve mechanically removing cells from an in vitro culture, allow for the simulation of wounds with high throughput and tight control over applied treatments to mechanistically study cell migration and proliferation functions that are critical to effective repair. Current methods require researchers to individually scratch each well with a pipette tip, resulting in low throughput as well as inconsistent scratch widths, straightness, and efficacy within and between wells. Here, we describe the design of a quickly assembled (<30 min), inexpensive (<$110) scratch assay rig that readily creates uniform scratches that are straight (average tortuosity < 1.1), have tunable widths (730–1100 µm), and fully remove damaged cells from the simulated wound region. Designed for a 24-well plate, the rig allows for high-throughput screening of varied experimental conditions or for testing many replicates. Application of the scratch assay device on an in vitro culture of neural progenitor cells (NPC) demonstrates the ability to detect differences in wound closure rate for three unique media conditions. These results support the implementation of this high-throughput scratch assay rig as a method to standardize and improve the efficiency of in vitro wound healing studies.
{"title":"In vitro wound simulation: A high-throughput device for scratch assays","authors":"Jacob E. Labovitz , Patrick Kulaga , Eric M. DuBois, Kunyu Li, Timothy M. O’Shea","doi":"10.1016/j.ymeth.2025.12.007","DOIUrl":"10.1016/j.ymeth.2025.12.007","url":null,"abstract":"<div><div>Traumatic injury to the healthy central nervous system (CNS) causes mechanical tissue damage that results in localized cell death and blood–brain-barrier (BBB) disruption. CNS tissue damage stimulates a multicellular wound response to limit the extent of damage but fails to reestablish the normal function of injured tissue. There is strong interest in developing new strategies to augment regeneration after CNS injury. To enable therapy development, reliable assays to screen and identify molecular approaches to augment glial-based wound responses over fibrotic scarring are needed. Scratch assays, which involve mechanically removing cells from an <em>in vitro</em> culture, allow for the simulation of wounds with high throughput and tight control over applied treatments to mechanistically study cell migration and proliferation functions that are critical to effective repair. Current methods require researchers to individually scratch each well with a pipette tip, resulting in low throughput as well as inconsistent scratch widths, straightness, and efficacy within and between wells. Here, we describe the design of a quickly assembled (<30 min), inexpensive (<$110) scratch assay rig that readily creates uniform scratches that are straight (average tortuosity < 1.1), have tunable widths (730–1100 µm), and fully remove damaged cells from the simulated wound region. Designed for a 24-well plate, the rig allows for high-throughput screening of varied experimental conditions or for testing many replicates. Application of the scratch assay device on an <em>in vitro</em> culture of neural progenitor cells (NPC) demonstrates the ability to detect differences in wound closure rate for three unique media conditions. These results support the implementation of this high-throughput scratch assay rig as a method to standardize and improve the efficiency of <em>in vitro</em> wound healing studies.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"246 ","pages":"Pages 223-235"},"PeriodicalIF":4.3,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145800327","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-02-01Epub Date: 2025-11-21DOI: 10.1016/j.ymeth.2025.10.010
Pradip Moon , Weizi Li , Antoni Chan , Bing Wang , Eghosa Bazuaye
Psoriatic arthritis (PsA) is a chronic inflammatory disease characterised by unpredictable flare-ups that are difficult to forecast, particularly in patients without an acute phase response. In this paper, we propose and apply an explainable, multimodal machine learning framework that jointly leverages structured temporal electronic patient records (EPRs) – sequential blood tests, disease activity scores, comorbidity burden, medications, and demographics – and unstructured clinical referral letters pre-processed with large language models ((LLMs, (Qwen-2.5 family)) to predict PsA flares. Gradient boosting models, Light Gradient Boosting Machine (LGBM) and eXtreme Gradient Boosting (XGBoost) were used to predict PsA flares, achieving the highest predictive performance 3 months before a clinic visit (accuracy = 92.8 %, AUROC = 0.94). Model performance gradually declined for longer timeframes (6 months: 78.2 %, AUROC = 0.80; 9 months: 76.6 %, AUROC = 0.78; 12 months: 72.2 %, AUROC = 0.75). LLMs applied to unstructured GP referral letters had limited standalone predictive value, but enhanced sensitivity and specificity when combined with the structured models in an ensemble approach. SHapley Additive exPlanations (SHAP) helped explain the prediction and demonstrated comorbidity count, disease scores, and immunosuppressive medications as the top predictors. Our results show that integrating both structured longitudinal data with unstructured clinical narratives using interpretable multimodal artificial intelligence can enable time-sensitive, personalised management of PsA flares and early clinical intervention.
{"title":"Explainable machine learning-based prediction of psoriatic arthritis flares using heterogenous real-world data for personalised patient care","authors":"Pradip Moon , Weizi Li , Antoni Chan , Bing Wang , Eghosa Bazuaye","doi":"10.1016/j.ymeth.2025.10.010","DOIUrl":"10.1016/j.ymeth.2025.10.010","url":null,"abstract":"<div><div>Psoriatic arthritis (PsA) is a chronic inflammatory disease characterised by unpredictable flare-ups that are difficult to forecast, particularly in patients without an acute phase response. In this paper, we propose and apply an explainable, multimodal machine learning framework that jointly leverages structured temporal electronic patient records (EPRs) – sequential blood tests, disease activity scores, comorbidity burden, medications, and demographics – and unstructured clinical referral letters pre-processed with large language models ((LLMs, (Qwen-2.5 family)) to predict PsA flares. Gradient boosting models, Light Gradient Boosting Machine (LGBM) and eXtreme Gradient Boosting (XGBoost) were used to predict PsA flares, achieving the highest predictive performance 3 months before a clinic visit (accuracy = 92.8 %, AUROC = 0.94). Model performance gradually declined for longer timeframes (6 months: 78.2 %, AUROC = 0.80; 9 months: 76.6 %, AUROC = 0.78; 12 months: 72.2 %, AUROC = 0.75). LLMs applied to unstructured GP referral letters had limited standalone predictive value, but enhanced sensitivity and specificity when combined with the structured models in an ensemble approach. SHapley Additive exPlanations (SHAP) helped explain the prediction and demonstrated comorbidity count, disease scores, and immunosuppressive medications as the top predictors. Our results show that integrating both structured longitudinal data with unstructured clinical narratives using interpretable multimodal artificial intelligence can enable time-sensitive, personalised management of PsA flares and early clinical intervention.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"246 ","pages":"Pages 95-106"},"PeriodicalIF":4.3,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145585760","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Molecular dynamics (MD) simulations and experimental analysis were performed on MgO–SrO containing bioactive glasses (MSBGs) with the composition of 60SiO2–(31–x)CaO–4P2O5–5MgO–xSrO (mol%) (x = 0, 1, 3, 5, 8, 10, 15, 20; M5S0–M5S20) to evaluate the structural properties, ion clustering, and dissolution behavior as a function of SrO content. Simulation employed the Buckingham potential for short-range interactions and Coulombic potentials for long-range forces. MSBGs had Si–O and P–O bond lengths of 1.609 Å and 1.491 Å, with O–Si–O and O–P–O bond angles centered at ∼109.3° and 109.4°, respectively, confirming tetrahedral SiO4/PO4 coordination. Across all compositions, Si–O–Si bonds dominated the majority of the distribution (88–89 %), with Si–O–P at 11–12 % and P–O–P negligible (∼0.3 %). Densities decreased from 2.913 g·cm−3 (M5S20) to 2.631 g·cm−3 (M5S0), reflecting network loosening with SrO substitution. Qn distribution remained stable, with Q3/Q4 fractions of 38–43 % and 26–30 % for Si-based tetrahedra. R-factor analysis revealed optimal homogeneity for M5S5 ( = 0.838252), balancing reduced cation clustering and moderate network stability. ICP-AES showed M5S5 with a sustained release of Si4+, Mg2+, and Sr2+ over 24 h. Meanwhile, antibacterial study resulted in statistically significant increase in efficiency for M5S5 compared to M5S0 (***p < 0.001). The combined computational and experimental findings identify M5S5 as the most promising candidate for biomedical applications requiring structural benefits, controlled ion release, and antibacterial efficiency.
{"title":"Investigation on structure–property relationships of MgO-SrO containing silicate-based bioactive glasses: An experimental and molecular dynamics simulation study","authors":"Amirhossein Moghanian , Ramin Farmani , Niloufar Kolivand , Arman Tayebi , Sirus Safaee","doi":"10.1016/j.ymeth.2025.12.001","DOIUrl":"10.1016/j.ymeth.2025.12.001","url":null,"abstract":"<div><div>Molecular dynamics (MD) simulations and experimental analysis were performed on MgO–SrO containing bioactive glasses (MSBGs) with the composition of 60SiO<sub>2</sub>–(31–x)CaO–4P<sub>2</sub>O<sub>5</sub>–5MgO–xSrO (mol%) (x = 0, 1, 3, 5, 8, 10, 15, 20; M5S0–M5S20) to evaluate the structural properties, ion clustering, and dissolution behavior as a function of SrO content. Simulation employed the Buckingham potential for short-range interactions and Coulombic potentials for long-range forces. MSBGs had Si–O and P–O bond lengths of 1.609 Å and 1.491 Å, with O–Si–O and O–P–O bond angles centered at ∼109.3° and 109.4°, respectively, confirming tetrahedral SiO<sub>4</sub>/PO<sub>4</sub> coordination. Across all compositions, Si–O–Si bonds dominated the majority of the distribution (88–89 %), with Si–O–P at 11–12 % and P–O–P negligible (∼0.3 %). Densities decreased from 2.913 g·cm<sup>−3</sup> (M5S20) to 2.631 g·cm<sup>−3</sup> (M5S0), reflecting network loosening with SrO substitution. Q<sup>n</sup> distribution remained stable, with Q<sup>3</sup>/Q<sup>4</sup> fractions of 38–43 % and 26–30 % for Si-based tetrahedra. R-factor analysis revealed optimal homogeneity for M5S5 (<span><math><mrow><msubsup><mi>R</mi><mrow><mi>S</mi><mi>i</mi><mo>/</mo><mi>P</mi></mrow><mrow><mi>S</mi><mi>r</mi></mrow></msubsup></mrow></math></span> = 0.838252), balancing reduced cation clustering and moderate network stability. ICP-AES showed M5S5 with a sustained release of Si<sup>4+</sup>, Mg<sup>2+</sup>, and Sr<sup>2+</sup> over 24 h. Meanwhile, antibacterial study resulted in statistically significant increase in efficiency for M5S5 compared to M5S0 (***p < 0.001). The combined computational and experimental findings identify M5S5 as the most promising candidate for biomedical applications requiring structural benefits, controlled ion release, and antibacterial efficiency.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"246 ","pages":"Pages 116-129"},"PeriodicalIF":4.3,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145712861","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Liquid biopsy, particularly the analysis of circulating tumor DNA (ctDNA), offers immense potential for non-invasive cancer diagnosis and monitoring. It provides a less invasive alternative to traditional tissue biopsies, enabling earlier detection and real-time assessment of disease progression. However, a significant hurdle in its widespread adoption is the extremely low concentration of ctDNA in biological samples, especially during the early stages of cancer, making sensitive and specific detection challenging. This work addresses the critical problem of developing a highly sensitive and specific method for low abundance ctDNA detection.
We developed a novel, highly sensitive, and specific method for ctDNA analysis, employing copper-free click chemistry (strain-promoted azide-alkyne cycloaddition, SPAAC) for enzyme-free amplification, coupled with magnetic bead-assisted fluorometric detection. This enzyme-free approach significantly enhanced specificity and reduced background noise. We meticulously optimized parameters, including primer length and annealing temperature, finding that 30-base primers and a 50 °C annealing temperature yielded optimal amplification efficiency. Our method successfully detected ctDNA at concentrations as low as 10 pM (15 bp primer). Agarose gel electrophoresis confirmed highly specific amplification with minimal non-specific products, and the assay demonstrated excellent allelic discrimination, accurately distinguishing single-nucleotide mutations. Importantly, the method proved robust in complex human serum samples, demonstrating its practical applicability.
This innovative, cost-effective, and enzyme-free platform overcomes many limitations of current ctDNA detection technologies. By enabling highly sensitive and specific detection of low abundance ctDNA, this methodology represents a significant leap forward for non-invasive cancer diagnostics, paving the way for earlier disease detection, improved treatment monitoring, and the broader implementation of personalized medicine.
{"title":"Sensitive and specific detection of ctDNA using Copper-Free click chemistry and magnetic bead Technology","authors":"Reza Didarian , Dilek Kanarya , Sonya Sahin , Canan Özyurt , Serap Evran , Dilek Odaci , Nimet Yildirim-Tirgil","doi":"10.1016/j.ymeth.2025.11.010","DOIUrl":"10.1016/j.ymeth.2025.11.010","url":null,"abstract":"<div><div>Liquid biopsy, particularly the analysis of circulating tumor DNA (ctDNA), offers immense potential for non-invasive cancer diagnosis and monitoring. It provides a less invasive alternative to traditional tissue biopsies, enabling earlier detection and real-time assessment of disease progression. However, a significant hurdle in its widespread adoption is the extremely low concentration of ctDNA in biological samples, especially during the early stages of cancer, making sensitive and specific detection challenging. This work addresses the critical problem of developing a highly sensitive and specific method for low abundance ctDNA detection.</div><div>We developed a novel, highly sensitive, and specific method for ctDNA analysis, employing copper-free click chemistry (strain-promoted azide-alkyne cycloaddition, SPAAC) for enzyme-free amplification, coupled with magnetic bead-assisted fluorometric detection. This enzyme-free approach significantly enhanced specificity and reduced background noise. We meticulously optimized parameters, including primer length and annealing temperature, finding that 30-base primers and a 50 °C annealing temperature yielded optimal amplification efficiency. Our method successfully detected ctDNA at concentrations as low as 10 pM (15 bp primer). Agarose gel electrophoresis confirmed highly specific amplification with minimal non-specific products, and the assay demonstrated excellent allelic discrimination, accurately distinguishing single-nucleotide mutations. Importantly, the method proved robust in complex human serum samples, demonstrating its practical applicability.</div><div>This innovative, cost-effective, and enzyme-free platform overcomes many limitations of current ctDNA detection technologies. By enabling highly sensitive and specific detection of low abundance ctDNA, this methodology represents a significant leap forward for non-invasive cancer diagnostics, paving the way for earlier disease detection, improved treatment monitoring, and the broader implementation of personalized medicine.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"246 ","pages":"Pages 83-94"},"PeriodicalIF":4.3,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145659955","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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