发布求助
文献互助 智能选刊 最新文献

STAR Protocols最新文献

筛选
英文 中文
Protocol for assessing T cell receptor-mediated human T cell cytotoxicity.
IF 1.3
STAR Protocols Pub Date : 2025-01-04 DOI: 10.1016/j.xpro.2024.103541
Kathryne E Marks, Ifeoluwakiisi Adejoorin, John Sowerby, Deepak A Rao
{"title":"Protocol for assessing T cell receptor-mediated human T cell cytotoxicity.","authors":"Kathryne E Marks, Ifeoluwakiisi Adejoorin, John Sowerby, Deepak A Rao","doi":"10.1016/j.xpro.2024.103541","DOIUrl":"https://doi.org/10.1016/j.xpro.2024.103541","url":null,"abstract":"<p><p>Measuring the cytotoxicity of human T cells is a valuable but challenging task. We present a protocol for assessing the ability of polyclonal human T cells to kill target cells upon T cell receptor (TCR) triggering. We describe steps for preparing L and T cells for the assay and incubation of target cells with effector cells. We detail procedures for assessing cell death by flow cytometry. This protocol can be applied to small numbers of effector cells isolated from patient samples. For complete details on the use and execution of this protocol, please refer to Wang et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103541"},"PeriodicalIF":1.3,"publicationDate":"2025-01-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142932886","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 generating splice isoform-specific mouse mutants using CRISPR-Cas9 and a minigene splicing reporter.
IF 1.3
STAR Protocols Pub Date : 2025-01-04 DOI: 10.1016/j.xpro.2024.103543
Yudong Teng, Kelsey Arbogast, Harald Junge, Zhe Chen
{"title":"Protocol for generating splice isoform-specific mouse mutants using CRISPR-Cas9 and a minigene splicing reporter.","authors":"Yudong Teng, Kelsey Arbogast, Harald Junge, Zhe Chen","doi":"10.1016/j.xpro.2024.103543","DOIUrl":"https://doi.org/10.1016/j.xpro.2024.103543","url":null,"abstract":"<p><p>Here, we present a protocol to alter the production of alternatively spliced mRNA variants, without affecting the overall gene expression, through CRISPR-Cas9-engineered genomic mutations in mice. We describe steps for designing guide RNA to direct Cas9 endonuclease to consensus splice sites, producing transgenic mice through pronuclear injection, and screening for desired mutations in cultured mammalian cells using a minigene splicing reporter. Splice isoform-specific mouse mutants provide valuable tools for genetic analyses beyond loss-of-function and transgenic alleles. For complete details on the use and execution of this protocol, please refer to Dailey-Krempel et al.<sup>1</sup> and Johnson et al.<sup>2</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103543"},"PeriodicalIF":1.3,"publicationDate":"2025-01-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142932890","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 capturing a full transcriptome from single preimplantation embryos using So-Smart-seq.
IF 1.3
STAR Protocols Pub Date : 2025-01-04 DOI: 10.1016/j.xpro.2024.103540
Chunyao Wei, Jeannie T Lee
{"title":"Protocol for capturing a full transcriptome from single preimplantation embryos using So-Smart-seq.","authors":"Chunyao Wei, Jeannie T Lee","doi":"10.1016/j.xpro.2024.103540","DOIUrl":"https://doi.org/10.1016/j.xpro.2024.103540","url":null,"abstract":"<p><p>Strand-optimized Smart-seq (So-Smart-seq) can capture a comprehensive transcriptome from low-input samples. This technique detects both polyadenylated and non-polyadenylated RNAs, inclusive of repetitive RNAs, while excluding highly abundant ribosomal RNAs. So-Smart-seq preserves strand information and minimizes 5' to 3' coverage bias. We describe steps for the analysis of single mouse preimplantation embryos, including embryo isolation, library preparation, ribosomal cDNA depletion, and initial data processing. The protocol may be adapted for other low-input samples and the detection of small RNAs of <200 nt. For complete details on the use and execution of this protocol, please refer to Wei et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103540"},"PeriodicalIF":1.3,"publicationDate":"2025-01-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142932888","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 detect neutral lipids with BODIPY staining in myeloid-derived suppressor cells in mouse mammary tumors.
IF 1.3
STAR Protocols Pub Date : 2025-01-03 DOI: 10.1016/j.xpro.2024.103485
Ukjin Kim, Rumela Chakrabarti
{"title":"Protocol to detect neutral lipids with BODIPY staining in myeloid-derived suppressor cells in mouse mammary tumors.","authors":"Ukjin Kim, Rumela Chakrabarti","doi":"10.1016/j.xpro.2024.103485","DOIUrl":"https://doi.org/10.1016/j.xpro.2024.103485","url":null,"abstract":"<p><p>Neutral lipids affect the immunosuppressive function of myeloid-derived suppressor cells (MDSCs). Here, we present a protocol for measuring neutral lipids in MDSCs using BODIPY from mouse mammary tumor derived from triple-negative breast cancer cells, 4T1, which is applicable to other mammary tumors of interest. We describe steps for 4T1 cell culture, single-cell isolation from tumors, staining of cells with antibodies and BODIPY, and flow cytometry. Furthermore, we introduce alternative protocols with MDSC sorting to overcome risk of cell death by BODIPY. For complete details on the use and execution of this protocol, please refer to Kim et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103485"},"PeriodicalIF":1.3,"publicationDate":"2025-01-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142928288","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 semisynthesis of histone H4 with site-specific modifications using irreversible sortase-mediated ligation.
IF 1.3
STAR Protocols Pub Date : 2025-01-03 DOI: 10.1016/j.xpro.2024.103527
Yihang Xiao, Kun Zou, Mingxuan Wu
{"title":"Protocol for semisynthesis of histone H4 with site-specific modifications using irreversible sortase-mediated ligation.","authors":"Yihang Xiao, Kun Zou, Mingxuan Wu","doi":"10.1016/j.xpro.2024.103527","DOIUrl":"https://doi.org/10.1016/j.xpro.2024.103527","url":null,"abstract":"<p><p>Post-translational modifications (PTMs) of histone H4 play significant roles in the regulation of chromatin status. Here, we present a protocol for semisynthesis of histone H4 by sortase-mediated ligation (SML). We describe steps for solid-phase peptide synthesis of H4R40C(1-42), recombinant expression and purification of H4(41-102), expression and purification of eSrt(2A-9), and preparation of acrylamidine. We then detail procedures for SML of histone H4. This protocol can also be applied to the preparation of homogenous proteins with PTMs. For complete details on the use and execution of this protocol, please refer to Xiao et al.<sup>1</sup><sup>,</sup><sup>2</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103527"},"PeriodicalIF":1.3,"publicationDate":"2025-01-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142928235","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
4D light sheet imaging, computational reconstruction, and cell tracking in mouse embryos.
IF 1.3
STAR Protocols Pub Date : 2025-01-02 DOI: 10.1016/j.xpro.2024.103515
Martin H Dominguez, Jonathon M Muncie-Vasic, Benoit G Bruneau
{"title":"4D light sheet imaging, computational reconstruction, and cell tracking in mouse embryos.","authors":"Martin H Dominguez, Jonathon M Muncie-Vasic, Benoit G Bruneau","doi":"10.1016/j.xpro.2024.103515","DOIUrl":"https://doi.org/10.1016/j.xpro.2024.103515","url":null,"abstract":"<p><p>As light sheet fluorescence microscopy (LSFM) becomes widely available, reconstruction of time-lapse imaging will further our understanding of complex biological processes at cellular resolution. Here, we present a comprehensive workflow for in toto capture, processing, and analysis of multi-view LSFM experiments using the ex vivo mouse embryo as a model system of development. Our protocol describes imaging on a commercial LSFM instrument followed by computational analysis in discrete segments, using open-source software. Quantification of migration and morphodynamics is included. For complete details on the use and execution of this protocol, please refer to Dominguez et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103515"},"PeriodicalIF":1.3,"publicationDate":"2025-01-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142928229","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 production of tonic CAR T cells with dasatinib.
IF 1.3
STAR Protocols Pub Date : 2024-12-30 DOI: 10.1016/j.xpro.2024.103529
Léa Rosselle, Thibault Leray, Sandy Joaquina, Benjamin Caulier, Emmet McCormack, Pascal Gelebart, Sébastien Wälchli, Else Marit Inderberg
{"title":"Protocol for production of tonic CAR T cells with dasatinib.","authors":"Léa Rosselle, Thibault Leray, Sandy Joaquina, Benjamin Caulier, Emmet McCormack, Pascal Gelebart, Sébastien Wälchli, Else Marit Inderberg","doi":"10.1016/j.xpro.2024.103529","DOIUrl":"https://doi.org/10.1016/j.xpro.2024.103529","url":null,"abstract":"<p><p>Chimeric antigen receptors (CARs) are synthetic molecules composed of an extracellular antigen-binding domain and an intracellular signaling domain, leading to tonic signaling and manufacturing challenges. We present a protocol for the expansion of tonic CARs by using a Food and Drug Administration (FDA)-approved kinase inhibitor, dasatinib. We report steps for T cell transduction with retrovirus, expansion and verification of CAR quality using flow cytometry, and killing assay. At only 30 nM, dasatinib improves tonic CAR T cell proliferation and quality after expansion. For complete details on the use and execution of this protocol, please refer to Caulier et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103529"},"PeriodicalIF":1.3,"publicationDate":"2024-12-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142910945","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 fluorescently stain vacuoles in Arabidopsis root cells.
IF 1.3
STAR Protocols Pub Date : 2024-12-21 DOI: 10.1016/j.xpro.2024.103537
Wylie Kristopher Tiernan, Graciela Veronica Castro, Kurtis Shipman, Cecilia Rodriguez-Furlan
{"title":"Protocol to fluorescently stain vacuoles in Arabidopsis root cells.","authors":"Wylie Kristopher Tiernan, Graciela Veronica Castro, Kurtis Shipman, Cecilia Rodriguez-Furlan","doi":"10.1016/j.xpro.2024.103537","DOIUrl":"https://doi.org/10.1016/j.xpro.2024.103537","url":null,"abstract":"<p><p>Plant vacuoles are essential organelles that respond to developmental and environmental signals. Here, we present a protocol for staining the vacuole lumen in Arabidopsis root cells, enabling precise visualization of vacuolar dynamics. We describe steps for preparing plant material and staining with commonly used fluorescent dyes. We then detail procedures for visualizing vacuoles in the blue, green, and red emission spectra, allowing for their combined use with a variety of compatible fluorescent-tagged protein markers. For complete details on the use and execution of this protocol, please refer to Dubrovsky et al.,<sup>1</sup> Fricker,<sup>2</sup> Bassil et al.,<sup>3</sup> Grzam et al.,<sup>4</sup> and Stefano et al.<sup>5</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103537"},"PeriodicalIF":1.3,"publicationDate":"2024-12-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142878122","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 monitor live-cell, real-time, mitochondrial respiration in mouse muscle cells using the Resipher platform. 使用 Resipher 平台实时监测小鼠肌肉细胞线粒体呼吸的活细胞方案。
IF 1.3
STAR Protocols Pub Date : 2024-12-20 Epub Date: 2024-09-20 DOI: 10.1016/j.xpro.2024.103330
Matthew Triolo, Mireille Khacho
{"title":"Protocol to monitor live-cell, real-time, mitochondrial respiration in mouse muscle cells using the Resipher platform.","authors":"Matthew Triolo, Mireille Khacho","doi":"10.1016/j.xpro.2024.103330","DOIUrl":"10.1016/j.xpro.2024.103330","url":null,"abstract":"<p><p>Mitochondrial function is typically assessed by measuring oxygen consumption at a given time point. However, this approach cannot monitor respiratory changes that occur over time. Here, we present a protocol to measure mitochondrial respiration in freshly isolated muscle stem cells, primary skeletal muscle, and immortalized C2C12 myoblasts in real time using the Resipher platform. We describe steps for preparing and plating cells, performing media changes, setting up the software and device, and analyzing data. This method can be adapted to other cell types. For complete details on the use and execution of this protocol, please refer to Triolo et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"5 4","pages":"103330"},"PeriodicalIF":1.3,"publicationDate":"2024-12-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11459069/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142297147","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Protocol for generating a 3D culture of epiblast stem cells. 生成上胚层干细胞三维培养液的方案。
IF 1.3
STAR Protocols Pub Date : 2024-12-20 Epub Date: 2024-09-27 DOI: 10.1016/j.xpro.2024.103347
Viviane S Rosa, Nanami Sato, Marta N Shahbazi
{"title":"Protocol for generating a 3D culture of epiblast stem cells.","authors":"Viviane S Rosa, Nanami Sato, Marta N Shahbazi","doi":"10.1016/j.xpro.2024.103347","DOIUrl":"10.1016/j.xpro.2024.103347","url":null,"abstract":"<p><p>Mouse gastrulation entails concomitant changes in cell fate, tissue shape, and embryo size. The use of a reproducible in vitro system is crucial for dissecting the mechanisms that coordinate these events. Here, we present a protocol for generating a 3D culture of epiblast stem cells (3D EpiSCs), which grow as epithelial spheroids mimicking key features of the gastrulating mouse embryonic epiblast. We describe steps for spheroid formation, growth, and passaging, followed by imaging or further downstream analyses. For complete details on the use and execution of this protocol, please refer to Sato et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"5 4","pages":"103347"},"PeriodicalIF":1.3,"publicationDate":"2024-12-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11465143/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142355529","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
下一页 尾页
0
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整，为了账户安全，请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
现在去查看 取消
×
提示
确定
请完成安全验证×
相关产品
×
本文献相关产品
联系我们：info@booksci.cn Book学术提供免费学术资源搜索服务，方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号：481959085
Book学术官方微信