Nature Protocols最新文献_第2页

Sialic acid aptamer and RNA in situ hybridization-mediated proximity ligation assay for spatial imaging of glycoRNAs in single cells. 唾液酸适体和RNA原位杂交介导的糖核糖核酸空间成像的近距离连接实验。

IF 13.1 1区生物学

Nature Protocols Pub Date : 2025-07-01 Epub Date: 2025-01-08 DOI: 10.1038/s41596-024-01103-x

Weijie Guo, Yuan Ma, Quanbing Mou, Xiangli Shao, Mingkuan Lyu, Valeria Garcia, Linggen Kong, Whitney Lewis, Zhenglin Yang, Shuya Lu, Yi Lu

{"title":"Sialic acid aptamer and RNA in situ hybridization-mediated proximity ligation assay for spatial imaging of glycoRNAs in single cells.","authors":"Weijie Guo, Yuan Ma, Quanbing Mou, Xiangli Shao, Mingkuan Lyu, Valeria Garcia, Linggen Kong, Whitney Lewis, Zhenglin Yang, Shuya Lu, Yi Lu","doi":"10.1038/s41596-024-01103-x","DOIUrl":"10.1038/s41596-024-01103-x","url":null,"abstract":"Glycosylated RNAs (glycoRNAs) have recently emerged as a new class of molecules of substantial interest owing to their potential roles in cellular processes and diseases. However, studying glycoRNAs is challenging owing to the lack of effective research tools including, but not limited to, imaging techniques to study the spatial distribution of glycoRNAs. Recently, we reported the development of a glycoRNA imaging technique, called sialic acid aptamer and RNA in situ hybridization-mediated proximity ligation assay (ARPLA), to visualize sialic acid-containing glycoRNAs with high sensitivity and specificity. Here we describe the experimental design principles and detailed step-by-step procedures for ARPLA-assisted glycoRNA imaging across multiple cell types. The procedure includes details for target selection, oligo design and preparation, optimized steps for RNA in situ hybridization, glycan recognition, proximity ligation, rolling circle amplification and a guideline for image acquisition and analysis. With properly designed probe sets and cells prepared, ARPLA-based glycoRNA imaging can typically be completed within 1 d by users with expertise in biochemistry and fluorescence microscopy. The ARPLA approach enables researchers to explore the spatial distribution, trafficking and functional contributions of glycoRNAs in various cellular processes.","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":"1930-1950"},"PeriodicalIF":13.1,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12234808/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142951995","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A multifunctional sensor for cell traction force, matrix remodeling and biomechanical assays in self-assembled 3D tissues in vitro. 一种多功能传感器，用于体外自组装3D组织的细胞牵引力、基质重塑和生物力学分析。

IF 13.1 1区生物学

Nature Protocols Pub Date : 2025-07-01 Epub Date: 2025-01-24 DOI: 10.1038/s41596-024-01106-8

Bashar Emon, Md Saddam Hossain Joy, William C Drennan, M Taher A Saif

{"title":"A multifunctional sensor for cell traction force, matrix remodeling and biomechanical assays in self-assembled 3D tissues in vitro.","authors":"Bashar Emon, Md Saddam Hossain Joy, William C Drennan, M Taher A Saif","doi":"10.1038/s41596-024-01106-8","DOIUrl":"10.1038/s41596-024-01106-8","url":null,"abstract":"Cell-matrix interactions, mediated by cellular force and matrix remodeling, result in dynamic reciprocity that drives numerous biological processes and disease progression. Currently, there is no available method for directly quantifying cell traction force and matrix remodeling in three-dimensional matrices as a function of time. To address this long-standing need, we developed a high-resolution microfabricated device that enables longitudinal measurement of cell force, matrix stiffness and the application of mechanical stimulation (tension or compression) to cells. Here a specimen comprising of cells and matrix self-assembles and self-integrates with the sensor. With primary fibroblasts, cancer cells and neurons we have demonstrated the feasibility of the sensor by measuring single or multiple cell force with a resolution of 1 nN and changes in tissue stiffness due to matrix remodeling by the cells. The sensor can also potentially be translated into a high-throughput system for clinical assays such as patient-specific drug and phenotypic screening. We present the detailed protocol for manufacturing the sensors, preparing experimental setup, developing assays with different tissues and for imaging and analyzing the data. Apart from microfabrication of the molds in a cleanroom (one time operation), this protocol does not require any specialized skillset and can be completed within 4-5 h.","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":"2005-2033"},"PeriodicalIF":13.1,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143039181","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}

引用次数: 0

Synthesis of bicyclo[3.1.1]heptanes, meta-substituted arene isosteres, from [3.1.1]propellane. 由[3.1.1]丙烷合成间取代芳烃异构双环[3.1.1]庚烷。

IF 13.1 1区生物学

Nature Protocols Pub Date : 2025-07-01 Epub Date: 2025-02-17 DOI: 10.1038/s41596-024-01109-5

Bhaskar Paul, Ayan Dasgupta, Nils Frank, Jeremy Nugent, Edward A Anderson

{"title":"Synthesis of bicyclo[3.1.1]heptanes, meta-substituted arene isosteres, from [3.1.1]propellane.","authors":"Bhaskar Paul, Ayan Dasgupta, Nils Frank, Jeremy Nugent, Edward A Anderson","doi":"10.1038/s41596-024-01109-5","DOIUrl":"10.1038/s41596-024-01109-5","url":null,"abstract":"The use of saturated small-ring bridged hydrocarbons as bioisosteres for aromatic rings has become a popular tactic in drug discovery. Perhaps the best known of such hydrocarbons is bicyclo[1.1.1]pentane, for which the angle between the exit vectors of the bridgehead substituents is identical to that of a para-substituted arene (180°). The development of meta-arene (bio)isosteres is much less explored due to the challenge of identifying an accurate geometric mimic (substituent exit vector angle ~120°, dihedral angle ~0°). To address this, we recently reported straightforward access to bicyclo[3.1.1]heptanes (BCHeps), which exactly meet these geometric properties, via radical ring-opening reactions of [3.1.1]propellane. This required the development of a scalable synthesis of [3.1.1]propellane, as well as the implementation of various ring-opening reactions and derivatizations. Here we describe methodology for a multigram scale synthesis of [3.1.1]propellane in five steps from commercially available ethyl 4-chlorobutanoate, which proceeds in an overall yield of 26-37%. We also describe the functionalization of [3.1.1]propellane to three key classes of BCHep iodides by photocatalyzed-atom transfer radical addition reactions using 456 nm blue light. We further report protocols for the elaboration of these products to other useful derivatives, via iron-catalyzed Kumada coupling with aryl Grignard reagents and conversion of a pivalate ester to a carboxylic acid through hydrolysis/oxidation. The total times required to synthesize [3.1.1]propellane, the BCHep iodides and the BCHep carboxylic acid are ~53, 6-8 and 40 h, respectively, requiring an average level of synthetic chemistry expertise (for example, masters and/or graduate students).","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":"2056-2082"},"PeriodicalIF":13.1,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143441539","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}

引用次数: 0

Single-cell CRISPR screening in mouse brain. 小鼠脑内单细胞CRISPR筛选。

IF 13.1 1区生物学

Nature Protocols Pub Date : 2025-07-01 DOI: 10.1038/s41596-024-01128-2

Eugenia V Pankevich, Christoph Bock

引用次数: 0

Massively parallel in vivo Perturb-seq screening. 大规模并行体内Perturb-seq筛选。

IF 13.1 1区生物学

Nature Protocols Pub Date : 2025-07-01 Epub Date: 2025-02-12 DOI: 10.1038/s41596-024-01119-3

Xinhe Zheng, Patrick C Thompson, Cassandra M White, Xin Jin

{"title":"Massively parallel in vivo Perturb-seq screening.","authors":"Xinhe Zheng, Patrick C Thompson, Cassandra M White, Xin Jin","doi":"10.1038/s41596-024-01119-3","DOIUrl":"10.1038/s41596-024-01119-3","url":null,"abstract":"Advances in genomics have identified thousands of risk genes impacting human health and diseases, but the functions of these genes and their mechanistic contribution to disease are often unclear. Moving beyond identification to actionable biological pathways requires dissecting risk gene function and cell type-specific action in intact tissues. This gap can in part be addressed by in vivo Perturb-seq, a method that combines state-of-the-art gene editing tools for programmable perturbation of genes with high-content, high-resolution single-cell genomic assays as phenotypic readouts. Here we describe a detailed protocol to perform massively parallel in vivo Perturb-seq using several versatile adeno-associated virus (AAV) vectors and provide guidance for conducting successful downstream analyses. Expertise in mouse work, AAV production and single-cell genomics is required. We discuss key parameters for designing in vivo Perturb-seq experiments across diverse biological questions and contexts. We further detail the step-by-step procedure, from designing a perturbation library to producing and administering AAV, highlighting where quality control checks can offer critical go-no-go points for this time- and cost-expensive method. Finally, we discuss data analysis options and available software. In vivo Perturb-seq has the potential to greatly accelerate functional genomics studies in mammalian systems, and this protocol will help others adopt it to answer a broad array of biological questions. From guide RNA design to tissue collection and data collection, this protocol is expected to take 9-15 weeks to complete, followed by data analysis.","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":"1733-1767"},"PeriodicalIF":13.1,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12237601/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143409116","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A guide to building a low-cost centrifuge force microscope module for single-molecule force experiments. 用于单分子力实验的低成本离心力显微镜模块的构建指南。

IF 13.1 1区生物学

Nature Protocols Pub Date : 2025-07-01 Epub Date: 2024-12-30 DOI: 10.1038/s41596-024-01102-y

Jibin Abraham Punnoose, Andrew Hayden, Chai S Kam, Ken Halvorsen

{"title":"A guide to building a low-cost centrifuge force microscope module for single-molecule force experiments.","authors":"Jibin Abraham Punnoose, Andrew Hayden, Chai S Kam, Ken Halvorsen","doi":"10.1038/s41596-024-01102-y","DOIUrl":"10.1038/s41596-024-01102-y","url":null,"abstract":"The ability to apply controlled forces to individual molecules or molecular complexes and observe their behaviors has led to many important discoveries in biology. Instruments capable of probing single-molecule forces typically cost >US$100,000, limiting the use of these techniques. The centrifuge force microscope (CFM) is a low-cost and easy-to-use instrument that enables high-throughput single-molecule studies. By combining the imaging capabilities of a microscope with the force application of a centrifuge, the CFM enables the simultaneous probing of hundreds to thousands of single-molecule interactions using tethered particles. Here we present a comprehensive set of instructions for building a CFM module that fits within a commercial benchtop centrifuge. The CFM module uses a 3D-printed housing, relies on off-the-shelf optical and electrical components, and can be built for less than US$1,000 in about 1 day. We also provide detailed instructions for setting up and running an experiment to measure force-dependent shearing of a short DNA duplex, as well as the software for CFM control and data analysis. The protocol is suitable for users with basic experience in analytical biochemistry and biophysics. The protocol enables the use of CFM-based experiments and may facilitate access to the single-molecule research field.","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":"1951-1975"},"PeriodicalIF":13.1,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12206937/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142910030","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Photothermal nanofiber-mediated photoporation for gentle and efficient intracellular delivery of macromolecules. 光热纳米纤维介导的温和高效的大分子细胞内递送的光穿孔。

IF 13.1 1区生物学

Nature Protocols Pub Date : 2025-07-01 Epub Date: 2025-01-22 DOI: 10.1038/s41596-024-01115-7

Dongyang Miao, Yuanyuan Song, Stijn De Munter, Huining Xiao, Bart Vandekerckhove, Stefaan C De Smedt, Chaobo Huang, Kevin Braeckmans, Ranhua Xiong

{"title":"Photothermal nanofiber-mediated photoporation for gentle and efficient intracellular delivery of macromolecules.","authors":"Dongyang Miao, Yuanyuan Song, Stijn De Munter, Huining Xiao, Bart Vandekerckhove, Stefaan C De Smedt, Chaobo Huang, Kevin Braeckmans, Ranhua Xiong","doi":"10.1038/s41596-024-01115-7","DOIUrl":"10.1038/s41596-024-01115-7","url":null,"abstract":"Photoporation with free photothermal nanoparticles (NPs) is a promising technology for gentle delivery of functional biomacromolecules into living cells, offering great flexibility in terms of cell types and payload molecules. However, the translational use of photoporation, such as for transfecting patient-derived cells for cell therapies, is hampered by safety and regulatory concerns as it relies on direct contact between cells and photothermal NPs. A solution is to embed the photothermal NPs in electrospun nanofibers, which form a substrate for cell culture. Here we present a protocol for photothermal electrospun nanofiber (PEN)-mediated photoporation that induces membrane permeabilization by photothermal effects and enables efficient intracellular delivery of payload molecules into various cell types. By incorporating photothermal NPs within biocompatible electrospun nanofibers, direct cellular contact with NPs is avoided, thus largely mitigating safety or regulatory issues. Importantly, PEN photoporation is gentler to cells compared with electroporation, the most commonly used physical transfection method, resulting in higher-quality genetically engineered cells with better therapeutic potential. According to this protocol, it takes 2-3 d to prepare PEN culture wells with the desired cells, 3-4 d to optimize PEN photoporation parameters for intracellular delivery of payload molecules into different cell types in vitro and 4-5 weeks to evaluate the in vivo therapeutic efficacy of PEN-photoporated T cells. The protocol also provides details on how to construct the laser-based setup for performing photoporation experiments.","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":"1810-1845"},"PeriodicalIF":13.1,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143024023","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}

引用次数: 0

Generation of human fetal brain organoids and their CRISPR engineering for brain tumor modeling. 人类胎儿脑类器官的生成及其用于脑肿瘤建模的CRISPR工程。

IF 13.1 1区生物学

Nature Protocols Pub Date : 2025-07-01 Epub Date: 2025-02-06 DOI: 10.1038/s41596-024-01107-7

Anna Pagliaro, Francesco Andreatta, Roxy Finger, Benedetta Artegiani, Delilah Hendriks

{"title":"Generation of human fetal brain organoids and their CRISPR engineering for brain tumor modeling.","authors":"Anna Pagliaro, Francesco Andreatta, Roxy Finger, Benedetta Artegiani, Delilah Hendriks","doi":"10.1038/s41596-024-01107-7","DOIUrl":"10.1038/s41596-024-01107-7","url":null,"abstract":"The developing human brain displays unique features that are difficult to study in animal models. Current in vitro models based on human brain tissue face several challenges, including the limited cellular heterogeneity in two- or three-dimensional neural stem cell cultures, while tissue slice cultures suffer from short survival. We recently established culture conditions to derive organoid cultures directly from human fetal brain tissue by preserving tissue integrity, which can be long-term expanded and display cellular heterogeneity and complex organization. In this Protocol, we describe detailed procedures to establish human fetal brain organoids (FeBOs) that broadly retain regional characteristics, along with procedures for their passaging and characterization. In addition, we describe genome engineering approaches applied to FeBOs to generate mutant FeBO lines that serve as versatile bottom-up brain cancer models. Lastly, we exemplify various downstream applications applicable to both healthy and mutant FeBOs. Scientists with experience in tissue culture can expect the establishment of human FeBO cultures to take 2-3 weeks, while genome engineering of FeBOs takes 2-4 months.","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":"1846-1883"},"PeriodicalIF":13.1,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143365257","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}

引用次数: 0

Live-cell synthesis of biocompatible quantum dots. 生物相容性量子点的活细胞合成。

IF 13.1 1区生物学

Nature Protocols Pub Date : 2025-07-01 Epub Date: 2025-03-17 DOI: 10.1038/s41596-024-01133-5

An-An Liu, Ran Cui, Xia Zong, Jianhong Jia, Yusi Hu, Jing-Ya Zhao, Dai-Wen Pang

{"title":"Live-cell synthesis of biocompatible quantum dots.","authors":"An-An Liu, Ran Cui, Xia Zong, Jianhong Jia, Yusi Hu, Jing-Ya Zhao, Dai-Wen Pang","doi":"10.1038/s41596-024-01133-5","DOIUrl":"10.1038/s41596-024-01133-5","url":null,"abstract":"Quantum dots (QDs) exhibit fluorescence properties with promising prospects for biomedical applications; however, the QDs synthesized in organic solvents shows poor biocompatibility, limiting their use in biological systems. We developed an approach for synthesizing QDs in live cells by coupling a series of intracellular metabolic pathways in a precise spatial and temporal sequence. We have validated this approach in yeast (Saccharomyces cerevisiae), Staphylococcus aureus, Michigan Cancer Foundation-7 (MCF-7) and Madin-Darby canine kidney (MDCK) cells. The intracellularly synthesized QDs are inherently stable and biocompatible, making them suitable for the direct in situ labeling of cells and cell-derived vesicles. Here, we provide an optimized workflow for the live-cell synthesis of QDs by using S. cerevisiae, S. aureus or MCF-7 cells. In addition, we detail a cell-free aqueous synthetic system (quasi-biosynthesis) containing enzymes, electrolytes, peptides and coenzymes, which closely mimics the intracellular synthetic conditions used in our cell culture system. In this solution, we synthesize biocompatible ultrasmall QDs that are easier to purify and characterize than those synthesized in cells. The live-cell-synthesized QDs can be used for bioimaging and microvesicle detection, whereas the quasi-biosynthesized QDs are suitable for applications such as biodetection, biolabeling and real-time imaging. The procedure can be completed in 3-4 d for live-cell QD synthesis and 2 h for the quasi-biosynthesis of QDs. The procedure is suitable for users with expertise in chemistry, biology, materials science and synthetic biology. This approach encourages interested researchers to engage in the field of QDs and develop further biomedical applications.","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":"1884-1914"},"PeriodicalIF":13.1,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143649761","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}

引用次数: 0

Fabrication of nonplanar tapered fibers to integrate optical and electrical signals for neural interfaces in vivo. 非平面锥形光纤在体内神经界面集成光、电信号的制备。

IF 13.1 1区生物学

Nature Protocols Pub Date : 2025-07-01 Epub Date: 2025-01-22 DOI: 10.1038/s41596-024-01105-9

Antonio Balena, Marco Bianco, Maria Samuela Andriani, Cinzia Montinaro, Barbara Spagnolo, Marco Pisanello, Filippo Pisano, Bernardo L Sabatini, Massimo De Vittorio, Ferruccio Pisanello

{"title":"Fabrication of nonplanar tapered fibers to integrate optical and electrical signals for neural interfaces in vivo.","authors":"Antonio Balena, Marco Bianco, Maria Samuela Andriani, Cinzia Montinaro, Barbara Spagnolo, Marco Pisanello, Filippo Pisano, Bernardo L Sabatini, Massimo De Vittorio, Ferruccio Pisanello","doi":"10.1038/s41596-024-01105-9","DOIUrl":"10.1038/s41596-024-01105-9","url":null,"abstract":"Implantable multifunctional probes have transformed neuroscience research, offering access to multifaceted brain activity that was previously unattainable. Typically, simultaneous access to both optical and electrical signals requires separate probes, while their integration into a single device can result in the emergence of photogenerated electrical artifacts, affecting the quality of high-frequency neural recordings. Among the nontrivial strategies aimed at the realization of an implantable multifunctional interface, the integration of optical and electrical capabilities on a single, minimally invasive, tapered optical fiber probe has been recently demonstrated using fibertrodes. Fibertrodes require the application of a set of planar microfabrication techniques to a nonplanar system with low and nonconstant curvature radius. Here we develop a process based on multiple conformal depositions, nonplanar two-photon lithography and chemical wet etching steps to obtain metallic patterns on the highly curved surface of the fiber taper. We detail the manufacturing, encapsulation and back end of the fibertrodes. The design of the probe can be adapted for different experimental requirements. Using the optical setup design, it is possible to perform angle selective light coupling with the fibertrodes and their implantation and use in vivo. The fabrication of fibertrodes is estimated to require 5-9 d. Nonetheless, due to the high scalability of a large part of the protocol, the manufacture of multiple fibertrodes simultaneously substantially reduces the required time for each probe. The procedure is suitable for users with expertise in microfabrication of electronics and neural recordings.","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":" ","pages":"1768-1809"},"PeriodicalIF":13.1,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143024020","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}

引用次数: 0