Reproducible Staged Differentiation of Retinal Organoids
Source Capowski et al. · University of Wisconsin-Madison, Madison, WI, USA · 10.1242/dev.171686
Abstract
Capowski et al. established a reproducible, standardized protocol for generating retinal organoids from human pluripotent stem cells with defined morphological staging criteria enabling rigorous quality control. The Gamm lab Wisconsin protocol validated differentiation across multiple cell lines and provides the reference framework for retinal organoid research and clinical-grade manufacturing.
Protocol overview
20 steps across 4 phases
- 1 Maintain pluripotent stem cells
- 2 Lift embryoid bodies
- 3 Wean EBs into Neural Induction Medium
- 4 Add BMP4 to culture
- 5 Plate EBs on Matrigel
- 6 Perform sequential half media changes
- 1 Switch to Retinal Differentiation Medium
- 2 Monitor optic vesicle formation
- 3 Dissect optic vesicles
- 1 Transfer organoids to poly-HEMA-coated flasks
- 2 Feed organoids with 3D-RDM twice weekly
- 3 Add all-trans retinoic acid
- 4 Remove retinoic acid
- 5 Monitor organoid morphology by light microscopy
- 6 Culture organoids for extended maturation
- 1 Perform optical coherence tomography (OCT)
- 2 Perform immunocytochemistry
- 3 Perform electron microscopy (TEM)
- 4 Perform multi-photon imaging
- 5 Perform optical metabolic imaging (OMI)
Full SOP
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Attribution
This SOP was authored by Organthis based on the published method in Capowski et al.. The originating laboratory holds no rights in this SOP and has not endorsed it unless marked Verified.
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