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Retinal Degeneration pp 119–141Cite as

The Mouse Retinal Organoid Trisection Recipe: Efficient Generation of 3D Retinal Tissue from Mouse Embryonic Stem Cells

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1834))

Abstract

The introduction of stem cell-based technologies for the derivation of three-dimensional retinal tissues, the so-called retinal organoids, offers many new possibilities for vision research: Organoids facilitate studies on retinal development and in vitro retinal disease modeling, as well as being valuable for drug testing. Further, retinal organoids also provide an unlimited cell source for cell replacement therapies. Here, we describe our protocol for efficiently differentiating large, stratified retinal organoids from mouse embryonic stem cells: unbiased manual dissection of the developing retinal organoid at an early stage into three evenly sized neuroepithelial portions (trisection step) doubles the yield of high-quality organoids. We also describe some useful applications of the protocol, e.g., generation of rod- or cone-enriched retinal organoids, AAV transfection, and cell birth dating. In addition, we provide details of how to process retinal organoids for single organoid gene expression analysis, immunohistochemistry, and electron microscopy.

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Acknowledgments

This work was supported by funding programs for DZNE Helmholtz and TU Dresden CRTD (M.K.), DFG (KA2794/3-1; SPP1738) (M.K.), and MedDrive TU Dresden UKD-Medical Faculty (M.K.). We thank Sara Oakeley (Basel, Switzerland) and Felix Wagner (DZNE, Dresden) for helpful comments on the manuscript.

Author information

Authors and Affiliations

  1. German Center for Neurodegenerative Diseases Dresden (DZNE), Dresden, Germany

    Manuela Völkner & Mike O. Karl

  2. Technische Universität Dresden, Center for Molecular and Cellular Bioengineering (CMCB), Center for Regenerative Therapies Dresden (CRTD), Dresden, Germany

    Thomas Kurth & Mike O. Karl

Authors
  1. Manuela Völkner
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  2. Thomas Kurth
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  3. Mike O. Karl
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Corresponding author

Correspondence to Mike O. Karl .

Editor information

Editors and Affiliations

  1. Institute of Human Genetics, University of Regensburg, Regensburg, Germany

    Bernhard H. F. Weber

  2. Department of Ophthalmology, University of Cologne, Cologne, Germany

    Thomas Langmann

1 Electronic Supplementary Material

Organoid trisection using single-step cutting technique (MP4 58363 kb)

Organoid trisection using two-step cutting technique (MP4 91134 kb)

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Völkner, M., Kurth, T., Karl, M.O. (2019). The Mouse Retinal Organoid Trisection Recipe: Efficient Generation of 3D Retinal Tissue from Mouse Embryonic Stem Cells. In: Weber, B.H.F., Langmann, T. (eds) Retinal Degeneration. Methods in Molecular Biology, vol 1834. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-8669-9_9

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  • DOI: https://doi.org/10.1007/978-1-4939-8669-9_9

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-8668-2

  • Online ISBN: 978-1-4939-8669-9

  • eBook Packages: Springer Protocols

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