IF 4.2 3区生物学

Methods Pub Date : 2025-04-07 DOI: 10.1016/j.ymeth.2025.04.003

Xingjian Zhou, Karoline Faust

{"title":"A high-throughput and time-efficient Nanopore full-length 16S rRNA gene sequencing protocol for synthetic microbial communities","authors":"Xingjian Zhou, Karoline Faust","doi":"10.1016/j.ymeth.2025.04.003","DOIUrl":"10.1016/j.ymeth.2025.04.003","url":null,"abstract":"<div><div>Next-generation sequencing (NGS) has transitioned from primarily research-focused applications to a mature technology. However, resolving microbial community composition on the species level based on the 16S rRNA gene is impeded by several critical bottlenecks that limit the efficiency and scalability of analyses. Specifically, standard MiSeq sequencing suffers from read-length limitation; library preparation requires multiple labour-intensive steps from DNA isolation to amplification and barcoding; and prolonged turnaround times delay results. These challenges underscore the need for improved methods, which our study aims to address. Recent advances in Oxford Nanopore long-read sequencing technology (ONT), including a smaller and cheaper benchtop instrument and support for diverse sample types, have enabled faster sequencing in-house with reduced costs. To address the need for standardized, reproducible workflows, we present an optimized and state-of-the-art protocol for full-length 16S rRNA gene sequencing using the ONT MinION sequencing device. Furthermore, we quantified the reproducibility and accuracy of our protocol and compared it with previous MiSeq results. The results showed that the accuracy of our sequencing pipeline for synthetic communities is significantly higher than for MiSeq pipeline. In summary, our protocol elucidates the composition of synthetic microbial communities in an easy, fast and accurate manner while ensuring reproducible results.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"240 ","pages":"Pages 14-20"},"PeriodicalIF":4.2,"publicationDate":"2025-04-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143828114","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}

引用次数: 0

Transforming breast cancer diagnosis and treatment with large language Models: A comprehensive survey 用大语言模型转化乳腺癌的诊断和治疗：一项综合调查

IF 4.2 3区生物学

Methods Pub Date : 2025-04-06 DOI: 10.1016/j.ymeth.2025.04.001

Mohsen Ghorbian , Mostafa Ghobaei-Arani , Saied Ghorbian

{"title":"Transforming breast cancer diagnosis and treatment with large language Models: A comprehensive survey","authors":"Mohsen Ghorbian , Mostafa Ghobaei-Arani , Saied Ghorbian","doi":"10.1016/j.ymeth.2025.04.001","DOIUrl":"10.1016/j.ymeth.2025.04.001","url":null,"abstract":"<div><div>Breast cancer (BrCa), being one of the most prevalent forms of cancer in women, poses many challenges in the field of treatment and diagnosis due to its complex biological mechanisms. Early and accurate diagnosis plays a fundamental role in improving survival rates, but the limitations of existing imaging methods and clinical data interpretation often prevent optimal results. Large Language Models (LLMs), which are developed based on advanced architectures such as transformers, have brought about a significant revolution in data processing and medical decision-making. By analyzing a large volume of medical and clinical data, these models enable early diagnosis by identifying patterns in images and medical records and provide personalized treatment strategies by integrating genetic markers and clinical guidelines. Despite the transformative potential of these models, their use in BrCa management faces challenges such as data sensitivity, algorithm transparency, ethical considerations, and model compatibility with the details of medical applications that need to be addressed to achieve reliable results. This review systematically reviews the impact of LLMs on BrCa treatment and diagnosis. This study’s objectives include analyzing the role of LLM technology in diagnosing and treating this disease. The findings indicate that the application of LLMs has resulted in significant improvements in various aspects of BrCa management, such as a 35% increase in the Efficiency of Diagnosis and BrCa Treatment (EDBC), a 30% enhancement in the System’s Clinical Trust and Reliability (SCTR), and a 20% improvement in the quality of patient education and information (IPEI). Ultimately, this study demonstrates the importance of LLMs in advancing precision medicine for BrCa and paves the way for effective patient-centered care solutions.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"239 ","pages":"Pages 85-110"},"PeriodicalIF":4.2,"publicationDate":"2025-04-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143791482","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}

引用次数: 0

RNA aptamer-induced fluorescence enhancement for NADH monitoring in cellular environment RNA适体诱导的荧光增强用于细胞环境中NADH的监测。

IF 4.2 3区生物学

Methods Pub Date : 2025-04-05 DOI: 10.1016/j.ymeth.2025.04.002

Mohammad Faysal Al Mazid , Syafira Rizqi Eskasalam , Jun-Seok Lee

{"title":"RNA aptamer-induced fluorescence enhancement for NADH monitoring in cellular environment","authors":"Mohammad Faysal Al Mazid , Syafira Rizqi Eskasalam , Jun-Seok Lee","doi":"10.1016/j.ymeth.2025.04.002","DOIUrl":"10.1016/j.ymeth.2025.04.002","url":null,"abstract":"<div><div>Cellular redox homeostasis is tightly regulated by the oxidation–reduction reactions of nicotinamide metabolites, including NAD(H) and NADP(H), which serve as essential cofactors in enzymatic processes related to energy metabolism. Monitoring intracellular NADH levels is therefore of significant interest. Most chemosensor designs to date rely on fluorescence turn-on mechanisms triggered by NADH oxidation, but these reaction-based sensors are inherently limited by NADH concentration and reaction kinetics. While NADH exhibits intrinsic fluorescence, its low quantum yield has led to the development of redox-sensitive substrates that emit fluorescence upon NADH oxidation. Here, we report an alternative fluorescence enhancement strategy based on an NADH-binding RNA aptamer. The interaction between NADH and a 49-base-pair RNA aptamer induces a 1.4-fold increase in fluorescence emission <em>in vitro</em> and an 1.8-fold increase in live-cell imaging. This fluorescence enhancement arises from aptamer-induced structural rigidity, analogous to the mechanism by which 4-(p-hydroxybenzylidene)-5-imidazolidinone (HBI) enhances fluorescence in green fluorescent protein. Using our aptamer-based assay, we established a live-cell fluorescence emission assay for real-time monitoring of cellular NADH dynamics.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"240 ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-04-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143802131","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}

引用次数: 0

Predicting genes associated with ossification of the posterior longitudinal ligament using graph attention network 利用图注意网络预测后纵韧带骨化相关基因。

IF 4.2 3区生物学

Methods Pub Date : 2025-04-04 DOI: 10.1016/j.ymeth.2025.03.014

Fanyu Kong , Han Liu , Xiaoqi Liu , Lei Shi

{"title":"Predicting genes associated with ossification of the posterior longitudinal ligament using graph attention network","authors":"Fanyu Kong , Han Liu , Xiaoqi Liu , Lei Shi","doi":"10.1016/j.ymeth.2025.03.014","DOIUrl":"10.1016/j.ymeth.2025.03.014","url":null,"abstract":"<div><div>Ossification of the posterior longitudinal ligament is a degenerative disease that severely impacts the spine, with a complex pathogenesis involving the interplay of multiple genes. This study utilizes a combination of graph neural networks and deep neural networks to systematically analyze genes associated with OPLL, leveraging genomics and bioinformatics techniques. By integrating gene data from the DisGeNET and HumanNetV2 databases, we constructed a GNN model to identify potential pathogenic genes for OPLL and validated the expression characteristics and mechanisms of these genes in different cell types. The findings indicate that the GNN model achieves remarkable accuracy and reliability in predicting genes associated with OPLL. Additionally, cellular trajectory analysis and immune cell infiltration studies uncovered distinct cellular environments and immune features in OPLL patients, emphasizing the significant roles of fibroblasts and mesenchymal stem cells in the disease's progression. Drug sensitivity analysis also sheds light on future personalized treatment options. This study not only enhances the understanding of OPLL's molecular mechanisms but also suggests new avenues for diagnostic and targeted therapy development.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"240 ","pages":"Pages 47-53"},"PeriodicalIF":4.2,"publicationDate":"2025-04-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143794190","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}

引用次数: 0

Optimized toolkit for the manipulation of immortalized axolotl fibroblasts 永生化蝾螈成纤维细胞的优化工具箱。

IF 4.2 3区生物学

Methods Pub Date : 2025-04-03 DOI: 10.1016/j.ymeth.2025.03.019

Benjamin J. Tajer , Glory Kalu , Sarah Jay , Eric Wynn , Antoine Decaux , Paul Gilbert , Hani D. Singer , Maddeline D. Kidd , Jeffery A. Nelson , Noora Harake , Noah J. Lopez , Nathan R. Souchet , Anna G. Luong , Aaron M. Savage , Sangwon Min , Alparslan Karabacak , Sebastian Böhm , Ryan T. Kim , Tim Froitzheim , Konstantinos Sousounis , Jessica L. Whited

{"title":"Optimized toolkit for the manipulation of immortalized axolotl fibroblasts","authors":"Benjamin J. Tajer , Glory Kalu , Sarah Jay , Eric Wynn , Antoine Decaux , Paul Gilbert , Hani D. Singer , Maddeline D. Kidd , Jeffery A. Nelson , Noora Harake , Noah J. Lopez , Nathan R. Souchet , Anna G. Luong , Aaron M. Savage , Sangwon Min , Alparslan Karabacak , Sebastian Böhm , Ryan T. Kim , Tim Froitzheim , Konstantinos Sousounis , Jessica L. Whited","doi":"10.1016/j.ymeth.2025.03.019","DOIUrl":"10.1016/j.ymeth.2025.03.019","url":null,"abstract":"<div><div>The axolotl salamander model has broad utility for regeneration studies, but this model is limited by a lack of efficient cell-culture-based tools. The Axolotl Limb-1 (AL-1) fibroblast line, the only available immortalized axolotl cell line, was first published over 20 years ago, but many established molecular biology techniques, such as lipofectamine transfection, CRISPR-Cas9 mutagenesis, and antibiotic selection, work poorly or remain untested in AL-1 cells. Innovating technologies to manipulate AL-1 cells in culture and study their behavior following transplantation into the axolotl will complement <em>in-vivo</em> studies, decrease the number of animals used, and enable the faster, more streamlined investigation of regenerative biology questions. Here, we establish transfection, mutagenesis, antibiotic selection, and <em>in-vivo</em> transplantation techniques in axolotl AL-1 cells. These techniques will enable efficient culture with AL-1 cells and guide future tool development for the culture and manipulation of other salamander cell lines.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"240 ","pages":"Pages 21-34"},"PeriodicalIF":4.2,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143787640","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}

引用次数: 0

3D printing of calcium doped Isomalt via custom-made Extruder: Facile approach for creating blood vascular like networks within tissue mimicking hydrogel matrix 通过定制挤出机三维打印掺钙异麦芽酮：在组织模拟水凝胶基质中创建类似血管网络的简便方法。

IF 4.2 3区生物学

Methods Pub Date : 2025-04-02 DOI: 10.1016/j.ymeth.2025.03.018

Amar Dhwaj , Nimisha Roy , Amit Prabhakar , Deepti Verma

{"title":"3D printing of calcium doped Isomalt via custom-made Extruder: Facile approach for creating blood vascular like networks within tissue mimicking hydrogel matrix","authors":"Amar Dhwaj , Nimisha Roy , Amit Prabhakar , Deepti Verma","doi":"10.1016/j.ymeth.2025.03.018","DOIUrl":"10.1016/j.ymeth.2025.03.018","url":null,"abstract":"<div><div>3D printing domain has witnessed rapid advancements with immense applications in various fields ranging from aerospace to 3D printed organs. This study describes a facile biofabrication approach for creating an Artificial blood vascular network inside the Hydrogel matrix by using Isomalt sugar (Sugar Alcohol) as a sacrificial component inside a composite-Hydrogel matrix. Conventional 3D-printers have extruder and hot-end assembly, whereas Bioprinters use pneumatic-piston, and piezoelectric-driven extrusion mechanisms. In this study, we describe the design and operation of a custom-made miniature precision lead screw-based syringe-pump extruder mechanism with integrated temperature-controlled heat-block. We are currently using this integrated setup for melt Isomalt-based 3D printing, which can be easily mounted over the Z-axis and is driven using a geared stepper motor with high torque, providing controlled extrusion of highly viscous polymers where sugar structures are used as sacrificial materials for making Artificial blood vascular like networks in the microfluidics domain within the composite Hydrogel matrix. Computational studies using COMSOL Multiphysics were performed to predict the diffusion pattern of the DMEM culture medium to estimate the rate of mass flow through a porous media. Furthermore, Cell based testing is performed using Human Adipose Derived Mesenchymal Stem Cells (HAD-MSC’s) which were cultured over the vascular Hydrogel matrix perfused with culture media with defined flowrates to mimic the natural function of the Nutrient and gaseous exchange inside human tissues. The proposed can be used to produce equivalent Tissue models which could be potentially used in On-chip drug testing platforms, drug discovery and regenerative medicine domains.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"239 ","pages":"Pages 72-84"},"PeriodicalIF":4.2,"publicationDate":"2025-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143787637","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}

引用次数: 0

Multi-Omics clustering by integrating clinical features from large language model 整合大型语言模型临床特征的多组学聚类

IF 4.2 3区生物学

Methods Pub Date : 2025-04-01 DOI: 10.1016/j.ymeth.2025.03.017

Xiucai Ye, Tianyi Shi, Dong Huang, Tetsuya Sakurai

{"title":"Multi-Omics clustering by integrating clinical features from large language model","authors":"Xiucai Ye, Tianyi Shi, Dong Huang, Tetsuya Sakurai","doi":"10.1016/j.ymeth.2025.03.017","DOIUrl":"10.1016/j.ymeth.2025.03.017","url":null,"abstract":"<div><div>Multi-omics clustering has emerged as a powerful approach for understanding complex biological systems and enabling cancer subtyping by integrating diverse omics data. Existing methods primarily focus on the integration of different types of omics data, often overlooking the value of clinical context. In this study, we propose a novel framework that incorporates clinical features extracted from large language model (LLM) to enhance multi-omics clustering. Leveraging clinical data extracted from pathology reports using a BERT-based model, our framework converts unstructured medical text into structured clinical features. These features are integrated with omics data through an autoencoder, enriching the information content of each omics layer to improve feature extraction. The extracted features are then projected into a latent subspace using singular value decomposition (SVD), followed by spectral clustering to obtain the final clustering result. We evaluate the proposed framework on six cancer datasets on three omics levels, comparing it with several state-of-the-art methods. The experimental results demonstrate that the proposed framework outperforms existing methods in multi-omics clustering for cancer subtyping. Moreover, the results highlight the efficacy of integrating clinical features derived from LLM, significantly enhancing clustering performance. This work underscores the importance of clinical context in multi-omics analysis and showcases the transformative potential of LLM in advancing precision medicine.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"239 ","pages":"Pages 64-71"},"PeriodicalIF":4.2,"publicationDate":"2025-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143769113","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}

引用次数: 0

Strategies for ocular drug delivery 眼部给药策略。

IF 4.2 3区生物学

Methods Pub Date : 2025-03-28 DOI: 10.1016/j.ymeth.2025.03.020

Silvia L. Fialho (Guest editor)

引用次数: 0

Corrigendum to “Production of AFM wedged cantilevers for stress-relaxation experiments: Uniaxial loading of soft, spherical cells” [Methods 236 (2025) 1–9] “用于应力松弛实验的AFM楔形悬臂梁的生产：软球形单元的单轴加载”的勘误表[方法236 (2025)1-9]

IF 4.2 3区生物学

Methods Pub Date : 2025-03-28 DOI: 10.1016/j.ymeth.2025.03.013

Riccardo Campanile , Jonne Helenius , Cristina Scielzo , Lydia Scarfò , Domenico Salerno , Mario Bossi , Marta Falappi , Alessia Saponara , Daniel J. Müller , Francesco Mantegazza , Valeria Cassina

引用次数: 0

A methodological comparison of synthesizing heavy metal substituted bioapatite 合成重金属取代生物磷灰石的方法比较

IF 4.2 3区生物学

Methods Pub Date : 2025-03-25 DOI: 10.1016/j.ymeth.2025.03.016

Kennedy A. Drake, Tyler A. Grubelich, Stephanie Wong, Alix C. Deymier

{"title":"A methodological comparison of synthesizing heavy metal substituted bioapatite","authors":"Kennedy A. Drake, Tyler A. Grubelich, Stephanie Wong, Alix C. Deymier","doi":"10.1016/j.ymeth.2025.03.016","DOIUrl":"10.1016/j.ymeth.2025.03.016","url":null,"abstract":"<div><div>This study evaluates two methods—maturation and direct precipitation—for synthesizing heavy metal substituted biomimetic hydroxyapatite (HA), focusing on their efficacy in mimicking human bone composition and crystallinity. Cobalt (Co) and chromium (Cr) substitutions were investigated due to their relevance to metal-on-metal implant degradation and the potential integration of these ions into bone mineral. The maturation method involves prolonged incubation, producing amorphous and bioresorbable apatites, while the direct precipitation (DP) method achieves rapid synthesis of highly crystalline apatites through controlled titration. Both approaches were characterized using X-ray diffraction (XRD), Raman spectroscopy, and Fourier Transform Infrared (FTIR) spectroscopy, confirming the apatitic nature of the samples and lattice strain induced by metal ion substitution. This study highlights the maturation method’s adaptability for long-term biological interactions and the DP method’s mechanical stability for load-bearing applications. Comparison of the structural and chemical properties of substituted HA from each method provides insights into optimizing synthesis techniques for diverse biomedical applications, such as bone tissue engineering and mitigating the effects of heavy metal ion release on bone health. These findings contribute to advancing hydroxyapatite-based biomaterials tailored for therapeutic and regenerative medicine needs.</div></div>","PeriodicalId":390,"journal":{"name":"Methods","volume":"239 ","pages":"Pages 42-48"},"PeriodicalIF":4.2,"publicationDate":"2025-03-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143714607","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}

引用次数: 0

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