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Protocol for generating human assembloids to investigate thalamocortical and corticothalamic synaptic transmission and plasticity.
IF 1.3
STAR Protocols Pub Date : 2025-02-07 DOI: 10.1016/j.xpro.2025.103630
Anjana Nityanandam, Mary H Patton, Ildar T Bayazitov, Kyle D Newman, Kristen T Thomas, Stanislav S Zakharenko
{"title":"Protocol for generating human assembloids to investigate thalamocortical and corticothalamic synaptic transmission and plasticity.","authors":"Anjana Nityanandam, Mary H Patton, Ildar T Bayazitov, Kyle D Newman, Kristen T Thomas, Stanislav S Zakharenko","doi":"10.1016/j.xpro.2025.103630","DOIUrl":"https://doi.org/10.1016/j.xpro.2025.103630","url":null,"abstract":"<p><p>Human induced pluripotent stem cells (hiPSCs) can be used to generate assembloids that recreate thalamocortical circuitry displaying short-term and long-term synaptic plasticity. Here, we describe a protocol for differentiating hiPSCs into thalamic and cortical organoids and then fusing them to generate thalamocortical assembloids. We detail the steps for using whole-cell patch-clamp electrophysiology to investigate the properties of synaptic transmission and synaptic plasticity in this model system. For complete details on the use and execution of this protocol, please refer to Patton et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103630"},"PeriodicalIF":1.3,"publicationDate":"2025-02-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143374795","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Protocol for predicting the single-cell network-based gene activity landscape during human B cell development.
IF 1.3
STAR Protocols Pub Date : 2025-02-07 DOI: 10.1016/j.xpro.2025.103614
Xin Huang, Xuetong Hou, Yizhen Li, Jun J Yang, Jiyang Yu
{"title":"Protocol for predicting the single-cell network-based gene activity landscape during human B cell development.","authors":"Xin Huang, Xuetong Hou, Yizhen Li, Jun J Yang, Jiyang Yu","doi":"10.1016/j.xpro.2025.103614","DOIUrl":"https://doi.org/10.1016/j.xpro.2025.103614","url":null,"abstract":"<p><p>Owing to inconsistencies in human B cell classification and the difficulty in distinguishing heterogeneous subpopulations, we present a protocol to construct gene regulatory networks and gene activity landscapes for human B cell developmental stages. We describe steps for acquiring bone marrow data; conducting single-cell downstream analysis; and leveraging the St. Jude Algorithm for the Reconstruction of Accurate Cellular Networks (SJARACNe), Network-based Bayesian Inference of Drivers (NetBID2), and single-cell Mutual Information-based Network Engineering Ranger (scMINER) algorithms for network-based analysis. Our protocol elucidates the biological characteristics of developmental stages in human B cells. For complete details on the use and execution of this protocol, please refer to Huang et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103614"},"PeriodicalIF":1.3,"publicationDate":"2025-02-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143374797","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Protocol for direct cDNA cap analysis of gene expression for paired-end patterned flow cell sequencing.
IF 1.3
STAR Protocols Pub Date : 2025-02-06 DOI: 10.1016/j.xpro.2024.103594
Diane Delobel, Hiromi Nishiyori-Sueki, Ilaria Nisoli, Hideya Kawaji, Pauline Robbe, Piero Carninci, Hazuki Takahashi
{"title":"Protocol for direct cDNA cap analysis of gene expression for paired-end patterned flow cell sequencing.","authors":"Diane Delobel, Hiromi Nishiyori-Sueki, Ilaria Nisoli, Hideya Kawaji, Pauline Robbe, Piero Carninci, Hazuki Takahashi","doi":"10.1016/j.xpro.2024.103594","DOIUrl":"https://doi.org/10.1016/j.xpro.2024.103594","url":null,"abstract":"<p><p>Cap analysis of gene expression (CAGE) is a technique that facilitates the assessment of the 5'-end of RNA transcript starting site (TSS) of both coding and non-coding genes. Here, we present a protocol for using CAGE on Illumina patterned flow cell technology with dual indexes on mouse and human samples. We describe steps for sequencing, automated data processing, and complete analytical framework ensuring CAGE operability for the determination of TSS and enhancers on the ever-evolving Illumina sequencing platforms.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103594"},"PeriodicalIF":1.3,"publicationDate":"2025-02-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143374761","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Protocol to assess changes in brain network resistance to perturbation during offline processing using TMS-EEG.
IF 1.3
STAR Protocols Pub Date : 2025-02-06 DOI: 10.1016/j.xpro.2025.103622
Martina Bracco, Tuomas P Mutanen, Domenica Veniero, Gregor Thut, Edwin M Robertson
{"title":"Protocol to assess changes in brain network resistance to perturbation during offline processing using TMS-EEG.","authors":"Martina Bracco, Tuomas P Mutanen, Domenica Veniero, Gregor Thut, Edwin M Robertson","doi":"10.1016/j.xpro.2025.103622","DOIUrl":"https://doi.org/10.1016/j.xpro.2025.103622","url":null,"abstract":"<p><p>Transcranial magnetic stimulation (TMS) perturbs specific brain regions and, combined with electroencephalography (EEG), enables the assessment of activity within their connected networks. We present a resting-state TMS-EEG protocol, combined with a controlled experimental design, to assess changes in brain network activity during offline processing, following a behavioral task. We describe steps for experimental design planning, setup preparation, data collection, and analysis. This approach minimizes biases inherent to TMS-EEG, ensuring an accurate assessment of changes within the network. For complete details of the use and execution of this protocol, please refer to Bracco et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103622"},"PeriodicalIF":1.3,"publicationDate":"2025-02-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143371236","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Protocol for the site-specific isolation of mouse endothelial cells using the modified Häutchen technique.
IF 1.3
STAR Protocols Pub Date : 2025-02-06 DOI: 10.1016/j.xpro.2025.103620
Alexander Brückner, Adrian Brandtner, Sarah Rieck, Bernd K Fleischmann, Daniela Wenzel
{"title":"Protocol for the site-specific isolation of mouse endothelial cells using the modified Häutchen technique.","authors":"Alexander Brückner, Adrian Brandtner, Sarah Rieck, Bernd K Fleischmann, Daniela Wenzel","doi":"10.1016/j.xpro.2025.103620","DOIUrl":"https://doi.org/10.1016/j.xpro.2025.103620","url":null,"abstract":"<p><p>Endothelial cells (ECs) play a major role in vascular homeostasis and pathology. They show a pronounced heterogeneity between different organs as well as within the same organ. Here, we present a protocol for the rapid site-specific isolation of murine aortic ECs. We describe steps for vessel preparation and the isolation of EC layers by freezing on glass coverslips using a modified Häutchen technique. This protocol can be adapted to investigate surface cells from other tissues as well. For complete details on the use and execution of this protocol, please refer to Brückner et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103620"},"PeriodicalIF":1.3,"publicationDate":"2025-02-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143374801","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Protocol for a patient-derived preclinical platform to model tumor-immune interactions and evaluate therapeutic efficacy.
IF 1.3
STAR Protocols Pub Date : 2025-02-06 DOI: 10.1016/j.xpro.2025.103623
Wen-Hsin Chang, Andrew I Chin, Ching-Hsien Chen
{"title":"Protocol for a patient-derived preclinical platform to model tumor-immune interactions and evaluate therapeutic efficacy.","authors":"Wen-Hsin Chang, Andrew I Chin, Ching-Hsien Chen","doi":"10.1016/j.xpro.2025.103623","DOIUrl":"https://doi.org/10.1016/j.xpro.2025.103623","url":null,"abstract":"<p><p>Here, we present a protocol for a preclinical ex vivo platform combining experimental flexibility with preservation of the tumor microenvironment. We outline steps for isolating human peripheral blood mononuclear cells (PBMCs), preparing patient-derived precision-cut tumor slices (PCTSs), cryopreserving the samples, and setting up the co-culture system. We provide instructions for treatment applications, interactions, and analyzing therapy responses. By preserving tumor architecture and heterogeneity, this model is applicable for evaluating tumor characteristics, immune interactions, and treatment efficacy in translational cancer research.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103623"},"PeriodicalIF":1.3,"publicationDate":"2025-02-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143371212","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Protocol to study the role of medial entorhinal cortex-basolateral amygdala circuit in context-induced retrieval of morphine withdrawal memory in mice.
IF 1.3
STAR Protocols Pub Date : 2025-02-06 DOI: 10.1016/j.xpro.2024.103542
Yali Fu, Ting Ye, Ming Chen, Bin Lai, Ping Zheng
{"title":"Protocol to study the role of medial entorhinal cortex-basolateral amygdala circuit in context-induced retrieval of morphine withdrawal memory in mice.","authors":"Yali Fu, Ting Ye, Ming Chen, Bin Lai, Ping Zheng","doi":"10.1016/j.xpro.2024.103542","DOIUrl":"https://doi.org/10.1016/j.xpro.2024.103542","url":null,"abstract":"<p><p>The medial entorhinal cortex (MEC) is crucial for context memory, yet its role in context-induced retrieval of morphine withdrawal memory remains to be investigated. Here, we present a protocol to evaluate the importance of projection neurons from the MEC to the basolateral amygdala (BLA) (MEC<sup>-BLA</sup> neurons) in mice during context-induced retrieval of morphine withdrawal memory using a conditioned place aversion (CPA) model. We describe steps for surgical procedure and behavioral experiments. Then, we detail procedures of immunofluorescence staining and image analysis. For complete details on the use and execution of this protocol, please refer to Fu et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103542"},"PeriodicalIF":1.3,"publicationDate":"2025-02-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143374820","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Protocol for the intracerebral injection of AAV-α-synuclein plus poly(ADP-ribose) to generate a Parkinson's disease monkey model.
IF 1.3
STAR Protocols Pub Date : 2025-02-06 DOI: 10.1016/j.xpro.2025.103621
Shuyi Liu, Wei Si, Zhengbo Wang
{"title":"Protocol for the intracerebral injection of AAV-α-synuclein plus poly(ADP-ribose) to generate a Parkinson's disease monkey model.","authors":"Shuyi Liu, Wei Si, Zhengbo Wang","doi":"10.1016/j.xpro.2025.103621","DOIUrl":"https://doi.org/10.1016/j.xpro.2025.103621","url":null,"abstract":"<p><p>The lack of an etiologic Parkinson's disease (PD) model in non-human primates has inhibited our understanding of the disease and the development of therapies. Here, we present a protocol for generating a PD model in Macaca fascicularis. We describe steps for localizing the substantia nigra and striatum using magnetic resonance imaging (MRI). We then detail a technique for the intracerebral administration of adeno-associated virus (AAV)-mediated overexpression of α-synuclein into the substantia nigra plus the injection of poly(ADP-ribose) (PAR) into the striatum. For complete details on the use and execution of this protocol, please refer to Liu et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103621"},"PeriodicalIF":1.3,"publicationDate":"2025-02-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143371234","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Protocol to study the impact of breast cancer on colonization resistance of mouse microbiota using network node manipulation.
IF 1.3
STAR Protocols Pub Date : 2025-02-06 DOI: 10.1016/j.xpro.2025.103618
Ana Laura Cano-Argüelles, Alejandra Wu-Chuang, Lourdes Mateos-Hernandez, Lianet Abuin-Denis, Apolline Maitre, Janet Avellanet, Arlem García, Dasha Fuentes, Alejandro Cabezas-Cruz
{"title":"Protocol to study the impact of breast cancer on colonization resistance of mouse microbiota using network node manipulation.","authors":"Ana Laura Cano-Argüelles, Alejandra Wu-Chuang, Lourdes Mateos-Hernandez, Lianet Abuin-Denis, Apolline Maitre, Janet Avellanet, Arlem García, Dasha Fuentes, Alejandro Cabezas-Cruz","doi":"10.1016/j.xpro.2025.103618","DOIUrl":"https://doi.org/10.1016/j.xpro.2025.103618","url":null,"abstract":"<p><p>Network analysis is a powerful tool for investigating complex interactions between different microbial taxa within a community. We present a protocol to study the gut microbial community in a mouse model of breast cancer using a network-based approach. Here, we describe the procedures for tumor cell production and inoculation and 16S rRNA data processing. We then detail steps for constructing co-occurrence networks based on correlations between microbial abundances. For complete details on the use and execution of this protocol, please refer to Wu-Chuang et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103618"},"PeriodicalIF":1.3,"publicationDate":"2025-02-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143374816","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Protocol for perfusing human axillary lymph nodes ex vivo to study structure and function in real time.
IF 1.3
STAR Protocols Pub Date : 2025-02-05 DOI: 10.1016/j.xpro.2025.103624
Amy Llewellyn, Rachel Barrow-McGee, Julia Stevenson, Jasmine Gore, Kalnisha Naidoo
{"title":"Protocol for perfusing human axillary lymph nodes ex vivo to study structure and function in real time.","authors":"Amy Llewellyn, Rachel Barrow-McGee, Julia Stevenson, Jasmine Gore, Kalnisha Naidoo","doi":"10.1016/j.xpro.2025.103624","DOIUrl":"https://doi.org/10.1016/j.xpro.2025.103624","url":null,"abstract":"<p><p>Lymph nodes regulate immunity and maintain fluid balance in health and disease. Here, we present a protocol that uses normothermic perfusion to sustain patient-derived lymph nodes ex vivo for up to 24 h to study their structure and function. We describe steps for setting up both thermoregulatory and perfusion circuits, cannulating human lymph nodes, and perfusion. This protocol can be used to study how human lymph nodes change in cancer and other diseases, and/or in response to perturbations, including drugs. For complete details on the use and execution of this protocol, please refer to Barrow-McGee et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103624"},"PeriodicalIF":1.3,"publicationDate":"2025-02-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143366015","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","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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