Nature Protocols最新文献

{"title":"Synthesis of radioafterglow nanoprobes for deep-tissue imaging of hydrogen peroxide.","authors":"Shuaige Bai, Youshi Lin, Cheng Xu, Kanyi Pu, Yan Zhang","doi":"10.1038/s41596-026-01421-2","DOIUrl":"https://doi.org/10.1038/s41596-026-01421-2","url":null,"abstract":"<p><p>Afterglow imaging eliminates the need for real-time excitation light, thereby greatly reducing tissue autofluorescence to achieve highly sensitive bioimaging. However, the clinical translation of existing photoafterglow and sonoafterglow probes developed so far is hampered by their limited tissue penetration. To address this, we recently developed radioafterglow nanoprobes (RANPs) on the basis of a cascade X-ray energy-converting mechanism for cancer-specific imaging and image-guided surgery. RANPs are constructed by co-assembling radioabsorbers, radiosensitizers and radioafterglow substrates into the amphiphilic polymers using the film rehydration method. This approach is rapid (<10 min), offers precise control over the concentration of encapsulated compounds and is highly scalable. Upon X-ray irradiation, RANPs exhibit near-infrared radioafterglow emission (peaked at 788 nm), with long half-life (~4.8 min) and high brightness. These properties enable an excitation depth of the RANPs up to 15 cm, three times deeper than that of photoafterglow. Here we further develop a tumor-specific RANP for precision molecular imaging that activates its radioafterglow signal specifically in the presence of hydrogen peroxide, a key biomarker of the tumor microenvironment. This design enables the ultrasensitive detection (signal-to-background ratio of 169) and surgical resection of small tumors (1 mm³) under an X-ray dosage 20 times lower than that required for inorganic materials. This protocol is suitable for users with expertise in chemistry, biology and materials science. The typical timeline is 1-2 weeks for nanoprobe construction and characterization, 1-2 weeks for cell assays and 3-4 weeks for animal experiments.</p>","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":""},"PeriodicalIF":18.4,"publicationDate":"2026-08-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148813517","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Selective heavy-metal sequestration and rare-earth element recovery using metal-organic frameworks.","authors":"Dhruv Menon, Prathmesh Bhadane, Priya Mahato, Prateek Goyal, Iseult Lynch, Swaroop Chakraborty, Superb K Misra","doi":"10.1038/s41596-026-01425-y","DOIUrl":"https://doi.org/10.1038/s41596-026-01425-y","url":null,"abstract":"<p><p>Modern water treatment and resource recovery demand materials that combine high performance with real-world durability. Traditional remediation approaches (e.g., precipitation and coagulation) have drawbacks (e.g., poor selectivity) that can be overcome using adsorption-based strategies. Metal-organic frameworks (MOFs) offer high tunability, high uptake capacities and the potential for regeneration. Translating MOF adsorbents into scalable, reliable technologies requires consistent method reporting and improved mechanistic insight, toward improving long-term stability in realistic water matrices. In this protocol, we describe how to deploy and assess the performance of MOF-based adsorbents for simultaneous heavy-metal sequestration (e.g., Pb(II), Cd(II), Ni(II) and Mn(II)) and rare-earth element recovery (e.g., Nd(III), Y(III) and Dy(III)) from complex water matrices. The workflow is broadly applicable across MOF chemistries and is illustrated using Cu(II)-based frameworks as representative model systems, synthesized at gram scale using commercially available precursors. We stabilize these frameworks through controlled defect engineering (e.g., partial metal substitution) to mitigate hydrolytic degradation and prolong operation time. We further tune morphology (e.g., nanosheets) to enhance surface accessibility and enable recyclability. For industrial applicability, we shape the MOFs into macrobeads via a green process. The procedure comprises: (i) MOF synthesis; (ii) comprehensive pre-adsorption characterization to assess crystallinity, porosity, morphology and composition using powder X-ray diffraction, nitrogen adsorption-desorption, scanning electron microscopy and inductively coupled plasma optical emission spectrometry; (iii) mechanistic adsorption assessment with kinetic, isotherm, thermodynamic, pH and selectivity analyses; (iv) regeneration and recovery workflows; and (v) deployment considerations in complex aqueous matrices, including industrial effluents, saline waters and e-waste leachates. The protocol provides a reproducible framework for implementing MOF-based adsorption technologies in water remediation and circular resource applications.</p>","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":""},"PeriodicalIF":18.4,"publicationDate":"2026-08-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148813528","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Author Correction: Considerations for the design of impactful citizen-science projects in microbiome research.","authors":"Sarah Ahannach, Sandra Condori-Catachura, Jelle Dillen, Caroline Dricot, Thies Gehrmann, Stijn Wittouck, Josiane Marie Kenfack, Wannes Van Beeck, Ilke De Boeck, Tom Eilers, Monica Ticlla, Agustina Santullo Latorre, Wenke Smets, Jari Temmermans, Enya Arconada Nuin, Sandra Van Puyvelde, Irina Spacova, Veronique Verhoeven, Sarah Lebeer","doi":"10.1038/s41596-026-01448-5","DOIUrl":"10.1038/s41596-026-01448-5","url":null,"abstract":"","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":""},"PeriodicalIF":18.4,"publicationDate":"2026-08-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148795827","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Using MetaboAnalyst 6.0 for exposomics data analysis-from LC-MS2 spectra processing to dose-response modeling and causal inference.","authors":"Zhiqiang Pang, Yao Lu, Guangyan Zhou, Huiting Ou, Charles Viau, Fumihiko Matsuda, Niladri Basu, Jianguo Xia","doi":"10.1038/s41596-026-01415-0","DOIUrl":"https://doi.org/10.1038/s41596-026-01415-0","url":null,"abstract":"<p><p>Exposomics is an emerging field of research that aims to comprehensively investigate individuals' environmental exposures and how these exposures relate to health outcomes. Liquid chromatography-tandem mass spectrometry is widely used in exposomics studies. MetaboAnalyst ( https://www.metaboanalyst.ca/ ) is a widely used platform for statistical and functional analysis of metabolomics data. The current MetaboAnalyst 6.0 release incorporates updates to meet the needs of exposomics studies, including improved support for tandem mass spectrometry compound identification, exposome annotation, dose-response analysis and linking to genetics and functions. Here we extend our 2022 Nature Protocol by providing step-by-step instructions on how to use MetaboAnalyst 6.0 for exposomics data analysis, including: liquid chromatography-tandem mass spectrometry spectra processing and compound identification (Stage 1), exposomics data processing and exploratory analysis (Stage 2), dose-response modeling to study metabolic responses to exposure levels (Stage 3) and leveraging known genetic associations for causal inference (Stage 4). We demonstrate Stages 1-3 using data from a recent blood exposomics study concerning electronic waste exposure. Stage 4 is illustrated through an investigation of the potential causal link between ʟ-isoleucine and type 2 diabetes. Stage 1 may take ~2 h to complete depending on server load, and the remaining stages may be executed in a total of ~90 min.</p>","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":""},"PeriodicalIF":18.4,"publicationDate":"2026-08-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148795787","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Centralized processing of frozen tumor tissues for global cancer genomics.","authors":"Behnoush Abedi-Ardekani, Arash Nikmanesh, Masoud Sotoudeh, Valerie Gaborieau, Farid Azmoudeh-Ardalan, Hiva Saffar, Stefano Serra, Christine Carreira, Priscilia Chopard, Sophie Ferlicot, Jean-Yves Scoazec, Christophe Lallemand, Elodie Colney, Sophie Guillot, Fatemeh Ghasemi-Kebria, Ana Carolina de Carvalho, Aida Ferreiro-Iglesias, Sandra Perdomo, Paul Brennan, Manuel Salto-Tellez, Gholamreaz Roshandel","doi":"10.1038/s41596-026-01426-x","DOIUrl":"https://doi.org/10.1038/s41596-026-01426-x","url":null,"abstract":"<p><p>Here we present the pathology workflow of the Mutographs project, a 7-year, multicancer genomics initiative led by the Sanger Institute and funded by Cancer Research UK. Mutographs aims to elucidate global differences in cancer incidence through whole-genome sequencing and mutational signature analysis of selected cancers with unexplained variation of incidence rate worldwide. The Protocol was developed by the International Agency for Research on Cancer, where pathology assessment and paired tumor-blood DNA extraction were performed. We provide details for handling frozen cancer tissues for multistep pathology evaluation and quality control, tumor-enrichment methods (such as macrodissecion and laser-capture microdissection) for certain tumor types, and manual and automated DNA extraction. This Protocol has achieved an overall success rate of 88% in providing high-quality DNA for whole-genome sequencing, aiding in the decision to exclude low-quality or noneligible tissues. This Protocol provides a reproducible, fully integrated pathology-molecular pipeline in challenging conditions to ensure high-quality DNA and reliable data interpretation at an unprecedented multinational scale and could be considered a model for high-quality tissue processing in international genomics initiatives. It is based on routine practice of pathology and requires practical experience of processing frozen tissues and morphological assessment of frozen tissues by digital pathology.</p>","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":""},"PeriodicalIF":18.4,"publicationDate":"2026-08-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148795766","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Synthesis of diiron subcluster [Fe₂[(μ-SCH₂)₂NH](CN)₂(CO)₄]^2- for artificial maturation of [FeFe]-hydrogenases.","authors":"Xin Yu, Fang Wei, Thomas B Rauchfuss","doi":"10.1038/s41596-026-01418-x","DOIUrl":"https://doi.org/10.1038/s41596-026-01418-x","url":null,"abstract":"<p><p>The [FeFe]-hydrogenases are widely distributed enzymes that efficiently make and use hydrogen (H₂). They have attracted intense interest as potential biocatalysts as well as models for biomimetic catalysis. Substrate turnover occurs at an Fe-based active site featuring an azadithiolate cofactor. The biosynthesis of the active site is remarkably elaborate. The production of these enzymes is greatly simplified by replacing the biosynthesis of this active site with reconstitution of the enzyme with chemically synthesized [Fe₂[(μ-SCH₂)₂NH](CN)₂(CO)₄]^2- ([1]^2-), the subject of this protocol. Two approaches to [1]^2- are described: a 'condensation route' and an 'Fmoc route'. Each of the two routes requires ~20 h of hands-on time. The condensation route entails the reaction of Fe₂(μ-SH)₂(CO)₆ and N₄(CH₂)₆ followed by cyanation to give [1]^2-. The most demanding step in this route is the preparation of Fe₂(μ-S₂)(CO)₆. The second route focuses on fluorenylmethoxycarbonyl (Fmoc)-protected derivatives of azadithiolate cofactor. This route proceeds via Fe₂[(μ-SCH₂)₂NFmoc](CO)₆. Both the condensation and Fmoc routes allow isotopic labeling of the active site, but ⁵⁷Fe labeling is especially simple using the Fmoc route.</p>","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":""},"PeriodicalIF":18.4,"publicationDate":"2026-08-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148760819","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"ONE microscopy.","authors":"Ali H Shaib, Mohamad Mahdi Alawieh, Silvio O Rizzoli","doi":"10.1038/s41596-026-01399-x","DOIUrl":"https://doi.org/10.1038/s41596-026-01399-x","url":null,"abstract":"<p><p>The introduction of expansion microscopy (ExM), a decade ago, marked a shift in super-resolution imaging, by physically separating fluorophores to bypass the diffraction limit. Numerous ExM developments have extended the method's reach since, yet molecular-scale resolution remained inaccessible. We recently developed one-step nanoscale ExM, which combines ExM with fluctuation-based super-resolution analysis to enable the direct visualization of individual protein shapes, using conventional fluorescence microscopes, a capability that was previously limited to cryo-electron microscopy and averaging-based techniques. Here we provide detailed procedures for gel embedding, labeling, expansion, image acquisition and data analysis. We also introduce a stable, user-friendly software package for efficient fluctuation analysis. Although one-step nanoscale ExM is broadly applicable to a range of samples, including purified proteins, cells and tissues, its most distinctive contribution lies in making single-protein shape analysis accessible and reproducible. Overall, we provide a practical framework for protein imaging on conventional equipment.</p>","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":""},"PeriodicalIF":18.4,"publicationDate":"2026-08-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148713199","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Direct printing of metasurfaces using formulated optical materials.","authors":"Hyunjung Kang, Eunji Lee, Yujin Park, Hongyoon Kim, Dong Kyo Oh, Jaekyung Kim, Junsuk Rho","doi":"10.1038/s41596-026-01407-0","DOIUrl":"https://doi.org/10.1038/s41596-026-01407-0","url":null,"abstract":"<p><p>Optical metasurfaces represent a promising technology for compact, lightweight and multifunctional optical devices; yet, their practical implementation remains challenging because of the need for high-refractive-index (high-index) materials, complex fabrication processes and limited substrate compatibility. Conventional top-down approaches relying on deposition, electron-beam lithography and etching are expensive and typically restricted to rigid substrates, hindering scalability and integration with flexible platforms. Nanoimprint lithography is a cost-effective, high-throughput alternative for metasurface fabrication; nevertheless, the low refractive index of conventional imprint resins fundamentally limits device performance. Here, we describe the fabrication of optical metasurfaces using a printable high-index composite material known as nanoparticle-embedded resin (nanoPER). By embedding high-index nanoparticles into a curable resin matrix, nanoPER achieves an effective refractive index above 1.8 at the target wavelength and enables single-step replication of functional nanostructures on a wide range of substrates, including flexible and curved surfaces. The procedure focuses on TiO₂ nanoPER and provides a reproducible and comprehensive guide covering resin formulation, nanoimprint lithography process parameters and optical characterization. By emphasizing scalability and versatility, this protocol supports the translation of metasurface research into real-world applications such as light detection and ranging, compact imaging and integrated photonics. The entire process can be completed within 1-2 days and can be performed by researchers with experience in nanofabrication and optical measurements.</p>","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":""},"PeriodicalIF":18.4,"publicationDate":"2026-08-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148713267","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Standardized assays for evaluating superoxide dismutase-like and catalase-like activities of nanozymes.","authors":"Ruofei Zhang, Hanqing Zhao, Sijie Zhang, Xiyun Yan, Kelong Fan","doi":"10.1038/s41596-026-01417-y","DOIUrl":"https://doi.org/10.1038/s41596-026-01417-y","url":null,"abstract":"<p><p>Antioxidant nanozymes are nanomaterials with superoxide dismutase-like or catalase-like activities. They have emerged as promising therapeutics for oxidative stress-related disorders. However, the lack of standardized quantitative assays for activity characterization has limited cross-study reproducibility and direct comparison of reported performance. Here this protocol provides step-by-step procedures for the quantitative characterization of the superoxide dismutase-like and catalase-like activities of nanozymes. Superoxide dismutase-like activity is quantified via a self-prepared xanthine/xanthine oxidase/water-soluble tetrazolium salt-1 system, in which nanozymes catalyze the dismutation of superoxide anions (O₂^•-), thereby suppressing formazan formation. By tuning O₂^•- generation through controlled xanthine oxidase concentrations, the protocol enables the determination of Michaelis-Menten kinetics for the short-lived O₂^•-. Catalase-like activity is quantified by monitoring hydrogen peroxide (H₂O₂) decomposition at 240 nm via ultraviolet-visible spectrophotometry, which yields specific activity and kinetic constants, whereas a complementary 3,3',5,5'-tetramethylbenzidine oxidation assay excludes interference from peroxidase-like reactions. Representative nanozymes spanning carbon-based, metal oxide, noble metal and single-atom systems are used as case studies to demonstrate the robustness and versatility of this protocol. Functional validation in cellular models using flow cytometry further demonstrates the practical applicability of reactive oxygen species scavenging. This protocol enables reliable and reproducible evaluation of antioxidant nanozymes, facilitating direct comparison across materials, elucidation of structure-activity relationships and rational optimization for biomedical applications. The complete procedure can be performed by researchers with standard training in biochemistry and flow cytometric analysis within 10-12 h.</p>","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":""},"PeriodicalIF":18.4,"publicationDate":"2026-08-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148707122","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Drug self-administration in head-fixed mice.","authors":"Elizabeth M Doncheck, Rachel E Clarke, Adam G Gordon, Josh Boquiren, Logan M Manusky, Jade Baek, Jacqueline E Paniccia, Isabella E Dinu, Eric B Dereschewitz, Roger I Grant, Garret D Stuber, James M Otis","doi":"10.1038/s41596-026-01406-1","DOIUrl":"10.1038/s41596-026-01406-1","url":null,"abstract":"<p><p>Drug self-administration has the greatest construct and predictive validity of the preclinical models for substance use disorder, providing landmark insights into the neurobiology of addiction. However, these experiments have traditionally been performed in freely moving animals, which can prohibit the incorporation of emerging neurotechnologies that require or are greatly facilitated by head restraint. Recently, we developed and validated a head-restrained approach in mice for intravenous and oral self-administration of drug and nondrug rewards. Here we present a step-by-step protocol for these experiments, including custom equipment construction, open-source software implementation and adaptation, catheter implantation, and unique considerations for conducting head-fixed self-administration experiments. To ensure that each component can be implemented by a wide range of audiences, detailed descriptions are provided so that this Protocol may serve as a standalone guide for researchers with varying levels of experience.</p>","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":""},"PeriodicalIF":18.4,"publicationDate":"2026-08-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148707087","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}