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

Publikation: Beitrag in Buch/Konferenzbericht/Sammelband/GutachtenBeitrag in Buch/Sammelband/GutachtenBeigetragenBegutachtung

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.

Details

OriginalspracheEnglisch
TitelRetinal Degeneration
Seiten119-141
Seitenumfang23
Band1834
ISBN (elektronisch)978-1-4939-8669-9
PublikationsstatusVeröffentlicht - 2019
Peer-Review-StatusJa

Publikationsreihe

ReiheMethods in molecular biology (Clifton, N.J.)
ISSN1064-3745

Externe IDs

ORCID /0000-0001-5624-1717/work/142239055
ORCID /0000-0002-0926-6556/work/142250486
Scopus 85054892358

Schlagworte

Schlagwörter

  • Animals, Cell Culture Techniques, Cell Differentiation, Cryopreservation, Fluorescent Antibody Technique, Genetic Vectors/genetics, Immunohistochemistry, Mice, Microscopy, Fluorescence, Mouse Embryonic Stem Cells/cytology, Organ Culture Techniques, Organoids/cytology, Retina/cytology, Transduction, Genetic