Deterministic scRNA-seq captures variation in intestinal crypt and organoid composition
Open accessAbstract
Single-cell RNA sequencing (scRNA-seq) approaches have transformed our ability to resolve cellular properties across systems, but are currently tailored toward large cell inputs (>1,000 cells). This renders them inefficient and costly when processing small, individual tissue samples, a problem that tends to be resolved by loading bulk samples, yielding confounded mosaic cell population read-outs. Here, we developed a deterministic, mRNA-capture bead and cell co-encapsulation dropleting system, DisCo, aimed at processing low-input samples (<500 cells). We demonstrate that DisCo enables precise particle and cell positioning and droplet sorting control through combined machine-vision and multilayer microfluidics, enabling continuous processing of low-input single-cell suspensions at high capture efficiency (>70%) and at speeds up to 350 cells per hour. To underscore DisCo's unique capabilities, we analyzed 31 individual intestinal organoids at varying developmental stages. This revealed extensive organoid heterogeneity, identifying distinct subtypes including a regenerative fetal-like Ly6a+ stem cell population that persists as symmetrical cysts, or spheroids, even under differentiation conditions, and an uncharacterized 'gobloid' subtype consisting predominantly of precursor and mature (Muc2+) goblet cells. To complement this dataset and to demonstrate DisCo's capacity to process low-input, in vivo-derived tissues, we also analyzed individual mouse intestinal crypts. This revealed the existence of crypts with a compositional similarity to spheroids, which consisted predominantly of regenerative stem cells, suggesting the existence of regenerating crypts in the homeostatic intestine. These findings demonstrate the unique power of DisCo in providing high-resolution snapshots of cellular heterogeneity in small, individual tissues.
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Droplet microfluidics can print one organoid per well and enable rapid generation of intra-organoid heterogeneity.
“Droplet microfluidics can print one organoid per well and enable the rapid generation of intra-organoid heterogeneity75.”
From Organoids (2022) · cites this paper as [75]
The Results describe a droplet-microfluidic system that encapsulates cells and beads for single-cell RNA sequencing, not one that prints organoids into wells. The Introduction and Results describe measuring heterogeneity in existing intestinal organoids; they do not address rapid generation of intra-organoid heterogeneity. Searches for one organoid per well and rapid generation found no supporting passage (page numbers unavailable). (LLM, conf=0.76)
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- Authors
- Bues, J., Biočanin, M., Pezoldt, J., Dainese, R., Chrisnandy, A., Rezakhani, S., Saelens, W., Gardeux, V., Gupta, R., Sarkis, R., Russeil, J., Saeys, Y., Amstad, E., Claassen, M., Lutolf, M. P., & Deplancke, B.
- Affiliations
- SIB Swiss Institute of Bioinformatics (CH)
- École Polytechnique Fédérale de Lausanne (CH)
- Universitätsklinikum Tübingen (DE)
- Ghent University (BE)
- Corresponding author
- Deplancke, Bart
- ORCID
- Venue
- Nature Methods
- Published
- 2022-02-14
- Volume
- 19
- Issue
- 3
- Pages
- 323-330
- ISSN
- 1548-7091 (ISSN-L)
- 1548-7105
- OpenAlex
- W4212799764
- Cited by
- 71
- References
- 63
- Citations by year
- 2026: 8 · 2025: 18 · 2024: 13 · 2023: 16 · 2022: 16
- FWCI
- 5.15
- Type
- journal article
- Language
- en
- Open access
- green
- hdl.handle.net
- Licence
- Locations
- Nature Methods (publishedVersion)
- PubMed (publishedVersion)
- Ghent University Academic Bibliography (Ghent University) (acceptedVersion, other-oa)
- Infoscience (Ecole Polytechnique Fédérale de Lausanne) (submittedVersion, cc-by-nc-nd)
- Funders
- Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (IZLIZ3_156815, CRSK-3_190627)
- Precision Health & related Technologies (PHRT-502) EPFL SV Interdisciplinary PhD Funding Program
- EC | Horizon 2020 Framework Programme (754462)
- ISAC Marylou Ingram scholar
- EPFL SV Interdisciplinary PhD Funding Program
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Single-cell and spatial transcriptomics
- Physical Sciences › Engineering › Biomedical Engineering › Innovative Microfluidic and Catalytic Techniques Innovation
- Physical Sciences › Engineering › Biomedical Engineering › Microfluidic and Bio-sensing Technologies
- Keywords
- single-cell RNA sequencing, intestinal organoids, intestinal crypts, organoid heterogeneity, goblet cells, intestinal stem cells, cysts
- MeSH terms
- Animals, Cell Differentiation, Intestinal Mucosa, Organoids, Stem Cells, Mice, Single-Cell Analysis
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