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Multiplexed single-cell analysis of organoid signaling networks

Sufi, J; Qin, X; Rodriguez, FC; Bu, YJ; Vlckova, P; Zapatero, MR; Nitz, M; (2021) Multiplexed single-cell analysis of organoid signaling networks. Nature Protocols , 16 pp. 4897-4918. 10.1038/s41596-021-00603-4. Green open access

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Abstract

Organoids are biomimetic tissue models comprising multiple cell types and cell states. Post-translational modification (PTM) signaling networks control cellular phenotypes and are frequently dysregulated in diseases such as cancer. Although signaling networks vary across cell types, there are limited techniques to study cell type–specific PTMs in heterocellular organoids. Here, we present a multiplexed mass cytometry (MC) protocol for single-cell analysis of PTM signaling and cell states in organoids and organoids co-cultured with fibroblasts and leukocytes. We describe how thiol-reactive organoid barcoding in situ (TOBis) enables 35-plex and 126-plex single-cell comparison of organoid cultures and provide a cytometry by time of flight (CyTOF) signaling analysis pipeline (CyGNAL) for computing cell type–specific PTM signaling networks. The TOBis MC protocol takes ~3 d from organoid fixation to data acquisition and can generate single-cell data for >40 antibodies from millions of cells across 126 organoid cultures in a single MC run.

Type: Article
Title: Multiplexed single-cell analysis of organoid signaling networks
Open access status: An open access version is available from UCL Discovery
DOI: 10.1038/s41596-021-00603-4
Publisher version: https://doi.org/10.1038/s41596-021-00603-4
Language: English
Additional information: This version is the author accepted manuscript. For information on re-use, please refer to the publisher's terms and conditions.
Keywords: Cancer models, Cell signalling, Gastrointestinal models, Proteomic analysis
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Cancer Institute
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Cancer Institute > Research Department of Oncology
URI: https://discovery.ucl.ac.uk/id/eprint/10135626
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