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Protocol for labeling and manipulating "home"-activated neurons in mice.
IF 1.3
STAR Protocols Pub Date : 2025-03-22 DOI: 10.1016/j.xpro.2025.103704
Xiaoya Su, Junyue He, Yunlong Liu, Keyu Liu, Bo Lei, Yi Zhong
{"title":"Protocol for labeling and manipulating \"home\"-activated neurons in mice.","authors":"Xiaoya Su, Junyue He, Yunlong Liu, Keyu Liu, Bo Lei, Yi Zhong","doi":"10.1016/j.xpro.2025.103704","DOIUrl":"https://doi.org/10.1016/j.xpro.2025.103704","url":null,"abstract":"<p><p>Home-related cues, such as home bedding, influence affective behaviors in mice, including fear relief, anxiety reduction, and defensive aggression. Here, we present a protocol for labeling and manipulating \"home\"-activated neurons in the lateral hypothalamus (LH) using a Tet-Off inducible system. We describe steps for virus injection, postoperative recovery, and combining c-Fos-driven activity labeling with chemogenetics. This protocol integrates neuronal targeting with behavioral assays to provide insights into the functional role of \"home\"-activated neurons in home-driven behavioral adaptations. For complete details on the use and execution of this protocol, please refer to Su et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 2","pages":"103704"},"PeriodicalIF":1.3,"publicationDate":"2025-03-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143693608","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 fabrication and application of a low-water-content polyelectrolyte hydrogel.
IF 1.3
STAR Protocols Pub Date : 2025-03-22 DOI: 10.1016/j.xpro.2025.103713
Zihang Shen, Zheyu Dong, Jiabao Bai, Daochen Yin, Zhi Sheng, Yijie Cai, Zheng Jia
{"title":"Protocol for the fabrication and application of a low-water-content polyelectrolyte hydrogel.","authors":"Zihang Shen, Zheyu Dong, Jiabao Bai, Daochen Yin, Zhi Sheng, Yijie Cai, Zheng Jia","doi":"10.1016/j.xpro.2025.103713","DOIUrl":"https://doi.org/10.1016/j.xpro.2025.103713","url":null,"abstract":"<p><p>Hydrogels are prone to dehydration over prolonged use, leading to changes in their properties. Inspired by the human skin's stratum corneum, we present a protocol for the fabrication and application of a low-water-content polyelectrolyte hydrogel (L-hydrogel) that maintains mechanical and electrical properties over time. We describe steps for preparing a hygroscopic monomer through an ionic exchange reaction and a low-water-content hydrogel by UV crosslinking reaction and testing the material behaviors of L-hydrogel. We then detail procedures for preparing and testing L-hydrogel-based devices. For complete details on the use and execution of this protocol, please refer to Shen et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 2","pages":"103713"},"PeriodicalIF":1.3,"publicationDate":"2025-03-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143701700","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 transgene-free naive human induced pluripotent stem cells from somatic cells using modified Sendai viral system.
IF 1.3
STAR Protocols Pub Date : 2025-03-21 DOI: 10.1016/j.xpro.2025.103700
Kaho Washizu, Shinya Yamanaka, Akira Kunitomi
{"title":"Protocol for generating transgene-free naive human induced pluripotent stem cells from somatic cells using modified Sendai viral system.","authors":"Kaho Washizu, Shinya Yamanaka, Akira Kunitomi","doi":"10.1016/j.xpro.2025.103700","DOIUrl":"https://doi.org/10.1016/j.xpro.2025.103700","url":null,"abstract":"<p><p>Sendai virus (SeV) vector represents a powerful tool for generating naive and primed human induced pluripotent stem cells (iPSCs) from somatic cells. Here, we present a protocol for the generation of transgene-free naive human iPSCs from human dermal fibroblasts (HDFs) and human peripheral mononuclear cells (PBMCs) using a modified SeV vector system. We describe steps for thawing the HDFs or PBMCs, reseeding HDFs, SeV vector infection, reseeding the SeV-infected HDFs or PBMCs on an irradiated mouse embryonic fibroblast (iMEF) plate, switching to t2iLGö+Y medium, passaging the generated naive iPSCs, removing the SeV vectors, and cryopreserving the naive iPSCs. For complete details on the use and execution of this protocol, please refer to Kunitomi et al.<sup>1</sup><sup>,</sup><sup>2</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 2","pages":"103700"},"PeriodicalIF":1.3,"publicationDate":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143693631","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
Examining cellular heterogeneity in human DNA methylation studies: Overview and recommendations.
IF 1.3
STAR Protocols Pub Date : 2025-03-21 Epub Date: 2025-02-12 DOI: 10.1016/j.xpro.2025.103638
Maggie Po-Yuan Fu, Sarah Martin Merrill, Keegan Korthauer, Michael Steffen Kobor
{"title":"Examining cellular heterogeneity in human DNA methylation studies: Overview and recommendations.","authors":"Maggie Po-Yuan Fu, Sarah Martin Merrill, Keegan Korthauer, Michael Steffen Kobor","doi":"10.1016/j.xpro.2025.103638","DOIUrl":"10.1016/j.xpro.2025.103638","url":null,"abstract":"<p><p>Intersample cellular heterogeneity (ISCH) is one of the largest contributors to DNA methylation (DNAme) variability. It is imperative to account for ISCH to accurately interpret analysis results in epigenome-wide association studies. We compiled this primer based on the current literature to guide researchers through the process of estimating and accounting for ISCH in DNA methylation studies. This primer outlines the procedure of bioinformatic ISCH prediction, including using reference-based and reference-free algorithms. It then follows with descriptions of several methods to account for ISCH in downstream analyses, including robust linear regression and principal-component-analysis-based adjustments. Finally, we outlined three methods for estimating differential DNAme signals in a cell-type-specific manner. Throughout the primer, we provided statistical and biological justification for our recommendations, as well as R code examples for ease of implementation.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103638"},"PeriodicalIF":1.3,"publicationDate":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11969412/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143417015","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 isolating extracellular vesicles from alveolar macrophages phagocytosing MRSA in vitro cell culture models. 在体外细胞培养模型中从吞噬MRSA的肺泡巨噬细胞中分离细胞外囊泡的方法。
IF 1.3
STAR Protocols Pub Date : 2025-03-21 Epub Date: 2024-12-20 DOI: 10.1016/j.xpro.2024.103526
Songjie Bai, Ruomeng Hu, Xuehuan Wen, Wenting Zhang, Bingyu Li, Jie Yang, Qing Yu, Xianchang Zeng, Huiqing Xiu, Zhijian Cai, Gensheng Zhang
{"title":"Protocol for isolating extracellular vesicles from alveolar macrophages phagocytosing MRSA in vitro cell culture models.","authors":"Songjie Bai, Ruomeng Hu, Xuehuan Wen, Wenting Zhang, Bingyu Li, Jie Yang, Qing Yu, Xianchang Zeng, Huiqing Xiu, Zhijian Cai, Gensheng Zhang","doi":"10.1016/j.xpro.2024.103526","DOIUrl":"10.1016/j.xpro.2024.103526","url":null,"abstract":"<p><p>Extracellular vesicles (EVs) play a crucial role in delivering bioactive cargo in infectious diseases. Here, we present a protocol for isolating EVs from alveolar macrophages (AMs) that phagocytose methicillin-resistant Staphylococcus aureus (MRSA) in vitro cell culture models. We describe steps for bacterial preparation; infection of AMs with MRSA; and isolation, purification, and characterization of EVs. This protocol provides a valuable perspective for studying EVs derived from pathogen-infected immune cells. For complete details on the use and execution of this protocol, please refer to Bai et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103526"},"PeriodicalIF":1.3,"publicationDate":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11730243/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142872902","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 to detect neutral lipids with BODIPY staining in myeloid-derived suppressor cells in mouse mammary tumors. 用BODIPY染色检测小鼠乳腺肿瘤骨髓源性抑制细胞中性脂的方案。
IF 1.3
STAR Protocols Pub Date : 2025-03-21 Epub 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":"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":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11754504/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142928288","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 and characterizing a nasal epithelial model using imaging with application for respiratory viruses. 使用呼吸道病毒成像生成和表征鼻上皮模型的方案。
IF 1.3
STAR Protocols Pub Date : 2025-03-21 Epub Date: 2025-01-07 DOI: 10.1016/j.xpro.2024.103520
Victor H K Lam, Aleena Ghafoor, Yazan Khan, Shirley Constable, Lane B Buchanan, David Zuanazzi, Reeya Parmar, Zeynep G Tepe, Leigh J Sowerby, Cindy M Liu, Ryan M Troyer, Jessica L Prodger
{"title":"Protocol for generating and characterizing a nasal epithelial model using imaging with application for respiratory viruses.","authors":"Victor H K Lam, Aleena Ghafoor, Yazan Khan, Shirley Constable, Lane B Buchanan, David Zuanazzi, Reeya Parmar, Zeynep G Tepe, Leigh J Sowerby, Cindy M Liu, Ryan M Troyer, Jessica L Prodger","doi":"10.1016/j.xpro.2024.103520","DOIUrl":"10.1016/j.xpro.2024.103520","url":null,"abstract":"<p><p>Air-liquid interface (ALI) culture can differentiate airway epithelial cells to recapitulate the respiratory tract in vitro. Here, we present a protocol for isolating and culturing nasal epithelial cells from turbinate tissues for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. We describe steps to overcome challenges of imaging fragile cultures, detect the production of mucus, and quantify intracellular virus post-SARS-CoV-2 infection. We present data on the optimal duration of ALI maturation prior to experimentation and describe which steps can be altered to optimize testing of specific hypotheses.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103520"},"PeriodicalIF":1.3,"publicationDate":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11760824/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142956074","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 the isolation of brain microvessels and visualization of RNA fluorescence in mice and humans. 小鼠和人脑微血管分离和RNA荧光可视化的方案。
IF 1.3
STAR Protocols Pub Date : 2025-03-21 Epub Date: 2025-01-07 DOI: 10.1016/j.xpro.2024.103530
Olivia M Osborne, Oandy Naranjo, Silvia Torices, Sarah Schmidlin, Destiny Tiburcio, Minseon Park, Michal Toborek
{"title":"Protocol for the isolation of brain microvessels and visualization of RNA fluorescence in mice and humans.","authors":"Olivia M Osborne, Oandy Naranjo, Silvia Torices, Sarah Schmidlin, Destiny Tiburcio, Minseon Park, Michal Toborek","doi":"10.1016/j.xpro.2024.103530","DOIUrl":"10.1016/j.xpro.2024.103530","url":null,"abstract":"<p><p>Here, we present a protocol for isolating microvessels from fresh or snap-frozen brain tissue from mice and humans, followed by visualization of RNA utilizing RNAscope hybridization for quantification of mRNA. We describe the steps for sample preparation and isolation, fixation, and hybridization. This protocol was specifically designed to integrate with RNAscope in situ hybridization. Although the protocol was developed in a mouse model, it can be optimized for use in other organisms, including human brain samples.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103530"},"PeriodicalIF":1.3,"publicationDate":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11760319/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142956076","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 3D photogrammetry and morphological digitization of complex skulls. 复杂颅骨三维摄影测量及形态数字化方案。
IF 1.3
STAR Protocols Pub Date : 2025-03-21 Epub Date: 2025-01-16 DOI: 10.1016/j.xpro.2024.103572
Naomi De Leo, Claudio Chimenti, Luigi Maiorano, Davide Tamagnini
{"title":"Protocol for 3D photogrammetry and morphological digitization of complex skulls.","authors":"Naomi De Leo, Claudio Chimenti, Luigi Maiorano, Davide Tamagnini","doi":"10.1016/j.xpro.2024.103572","DOIUrl":"10.1016/j.xpro.2024.103572","url":null,"abstract":"<p><p>Here, we present a protocol for 3D photogrammetry and morphological digitization of skulls, including complex ones with tusks, antlers, and horns, which are challenging to reconstruct digitally. We describe steps for setting up specimens for image acquisition, including camera and lighting configurations, and the subsequent image processing to generate high-quality 3D models. We also outline the extraction of morphological data for accurate geometric morphometric analyses.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103572"},"PeriodicalIF":1.3,"publicationDate":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11787561/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143013051","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 cell image-based spatiotemporal proteomics in budding yeast. 芽殖酵母基于细胞图像的时空蛋白质组学研究。
IF 1.3
STAR Protocols Pub Date : 2025-03-21 Epub Date: 2025-01-16 DOI: 10.1016/j.xpro.2024.103577
Athanasios Litsios, Myra Paz David Masinas, Helena Friesen, Charles Boone, Brenda J Andrews
{"title":"Protocol for cell image-based spatiotemporal proteomics in budding yeast.","authors":"Athanasios Litsios, Myra Paz David Masinas, Helena Friesen, Charles Boone, Brenda J Andrews","doi":"10.1016/j.xpro.2024.103577","DOIUrl":"10.1016/j.xpro.2024.103577","url":null,"abstract":"<p><p>The eukaryotic cell division cycle is a highly conserved process, featuring fluctuations in protein localization and abundance required for key cell cycle transitions. Here, we present a protocol for the spatiotemporal analysis of the proteome during the budding yeast cell division cycle using live-cell imaging. We describe steps for strain construction, cell cultivation, microscopy, and image analysis. Variations of this protocol can be applied for the spatiotemporal analysis of the proteome in different contexts, such as genetic and environmental perturbations. For complete details on the use and execution of this protocol, please refer to Litsios et al.<sup>1</sup>.</p>","PeriodicalId":34214,"journal":{"name":"STAR Protocols","volume":"6 1","pages":"103577"},"PeriodicalIF":1.3,"publicationDate":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11787502/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143013053","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
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