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Traject3d for studying 3D cellular heterogeneity

Nature Reviews Cancer volume 23, page 577 (2023)Cite this article

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3D in vitro culture can be used to study the genetic mechanisms that give rise to multicellular structures from the interactions of individual cells. However, the heterogeneity of the structures as a function of time presents an inherent challenge to this approach. Although single-cell profiling can provide insights into function, this is a snapshot of cell populations at a given time rather than capturing dynamic heterogeneity during state transitions. To address this, we developed Traject3d, a workflow to identify co-occurring morphological phenotypes using label-free, time-lapse imaging.

In brief, a single-cell suspension is cultured within extracellular matrix and live-imaged for 3–7 days as cells divide and form 3D spheroids. The CellProfiler open-source software is used to track structures within this time-lapse, measuring size, shape and movement. Using these metrics, spheroids are first classified based on phenotype, as determined by user-defined criteria or a data-driven approach, and then ordered using real-time tracking data to identify behavioural patterns over time.

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References

Original article

  • Freckmann, E. C. et al. Traject3d allows label-free identification of distinct co-occurring phenotypes within 3D culture by live imaging. Nat. Commun. 13, 5317 (2022)

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  1. Eva C. Freckmann

    Present address: CRUK Beatson Institute, Glasgow, UK

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  1. University of Glasgow, Glasgow, UK

    Eva C. Freckmann

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  1. Eva C. Freckmann
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Correspondence to Eva C. Freckmann.

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Competing interests

The author was supported by a University of Glasgow Industrial Partnership PhD scheme co-funded by Essen Bioscience, Sartorius Group.

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Freckmann, E.C. Traject3d for studying 3D cellular heterogeneity. Nat Rev Cancer 23, 577 (2023). https://doi.org/10.1038/s41568-023-00592-4

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  • DOI: https://doi.org/10.1038/s41568-023-00592-4

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