Work

Protocol for generating human cerebral organoids from two-dimensional cultures of pluripotent stem cells bypassing embryoid body aggregation

Year: 2025

Type: article

Abstract: Human cerebral organoids (hCOs) provide an excellent model for the study of human brain development and disease. Here, we present a protocol to obtain hCOs directly from two-dimensional (2D) pluripotent stem cell (PSC) cultures, avoiding cell dissociation and posterior embryoid body (EB) aggregation. We describe steps for subjecting 2D cultures to a neural fate and subsequently developing hCOs. We then detail the evaluation of different cellular types. For complete details on the use and execution of this protocol, please refer to González-Sastre et al. 1 • Steps for generating hCOs from 2D colonies of PSCs bypassing EB aggregation • Instructions for highly efficient neuroepithelial induction • Guidance on the characterization of hCOs with cellular diversity Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Human cerebral organoids (hCOs) provide an excellent model for the study of human brain development and disease. Here, we present a protocol to obtain hCOs directly from two-dimensional (2D) pluripotent stem cell (PSC) cultures, avoiding cell dissociation and posterior embryoid body (EB) aggregation. We describe steps for subjecting 2D cultures to a neural fate and subsequently developing hCOs. We then detail the evaluation of different cellular types.

Source: STAR Protocols

DOI: 10.1016/j.xpro.2025.103678