Protocol for open-source automated universal high-content multiplex fluorescence for RNA in situ analysis
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.
Authors
- Jean Descarpentrie (ORCID: https://orcid.org/0000-0003-3434-359X)
- Ioannis S. Pateras (ORCID: https://orcid.org/0000-0002-6406-9811)
- Océane Martin (ORCID: https://orcid.org/0000-0003-0783-9119)
- Lucie Brisson
- Thomas Mathivet
- Wilfried Souleyreau (ORCID: https://orcid.org/0000-0002-4831-5891)
- María López Chiloeches (ORCID: https://orcid.org/0000-0003-4255-1532)
- Teresa Frisan (ORCID: https://orcid.org/0000-0002-1209-0942)
- Florian Bernard (ORCID: https://orcid.org/0000-0002-6110-2567)
Institutions
- Centre National de la Recherche Scientifique (FR)
- Université de Bordeaux (FR)
- Inserm (FR)
- National and Kapodistrian University of Athens (GR)
- Bordeaux Population Health (FR)
- Régulations Naturelle et Artificielle (FR)
- Institut de Biochimie et Génétique Cellulaires (FR)
- Umeå University (SE)
Publication Details
- Journal
- STAR Protocols
- Published
- 2024-12-01
- DOI
- https://doi.org/10.1016/j.xpro.2024.103508
- Citations
- 1
- Primary Topic
- Molecular Biology Techniques and Applications
- Type
- article
- Field-Weighted Citation Impact
- 0.23
Funders
- Lung Cancer Research Foundation
- Cancerfonden
- Vetenskapsrådet
- Umeå Universitet
- Cancer Research Foundation in Northern Sweden
- Kempestiftelserna