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Protocol for open-source automated universal high-content multiplex fluorescence for RNA in situ analysis
Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
University of Bordeaux, INSERM, U1212, Nucleic Acids: Natural and Artificial Regulations Laboratory, Bordeaux, France.
University of Bordeaux, INSERM, U1312 BRIC, Tumor and Vascular Biology Laboratory, Bordeaux, France.
University of Bordeaux, INSERM, U1312 BRIC, Tumor and Vascular Biology Laboratory, Bordeaux, France.
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2024 (English)In: STAR Protocols, E-ISSN 2666-1667, Vol. 5, no 4, article id 103508Article in journal (Refereed) Published
Abstract [en]

In situ hybridization visualizes RNA in cells, but image analysis is complex. We present a protocol based on open-source software for automated high-content multiplex fluorescence in situ transcriptomics analysis. Steps include nuclei segmentation with a Fiji macro and quantification of up to 14 mRNA probes per image. We describe procedures for storing raw data, quality control images, and the use of a Python app to summarize all the results in one spreadsheet detailing the number of single or co-positive cells.
Place, publisher, year, edition, pages
Elsevier, 2024. Vol. 5, no 4, article id 103508
Keywords [en]
cell biology, health sciences, in situ hybridization
National Category
Cell and Molecular Biology Medical Imaging
Identifiers
URN: urn:nbn:se:umu:diva-232964DOI: 10.1016/j.xpro.2024.103508ISI: 001375026800001PubMedID: 39644494Scopus ID: 2-s2.0-85211160702OAI: oai:DiVA.org:umu-232964DiVA, id: diva2:1924828
Funder
Swedish Research Council, 2021-00960Swedish Cancer Society, 23 2814 PjThe Kempe Foundations, 2021 JCK-3110Cancerforskningsfonden i Norrland, AMP20-993Lions Cancerforskningsfond i Norr, LP 22-2306Lions Cancerforskningsfond i Norr, LP 24-2357Available from: 2025-01-07 Created: 2025-01-07 Last updated: 2025-08-28Bibliographically approved

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fulltext(8021 kB)172 downloads
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Descarpentrie, JeanLopez Chiloeches, MariaFrisan, Teresa

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Descarpentrie, JeanLopez Chiloeches, MariaFrisan, Teresa
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Umeå Centre for Microbial Research (UCMR)Department of Molecular Biology (Faculty of Medicine)Department of Molecular Biology (Faculty of Science and Technology)
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STAR Protocols
Cell and Molecular BiologyMedical Imaging

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