The differentiation of pluripotent stem cells in three-dimensional cultures can recapitulate key aspects of brain development, but protocols are prone to variable results. Here we differentiated multiple human pluripotent stem cell lines for over 100 d using our previously developed approach to generate brain-region-specific organoids called cortical spheroids and, using several assays, found that spheroid generation was highly reliable and consistent. We anticipate the use of this approach for large-scale differentiation experiments and disease modeling. A protocol adapted to xeno- and feeder-free conditions is shown to generate reliable and consistent cortical brain organoids across differentiations and source stem cell lines, making it suitable for disease modeling and other applications.

Reliability of human cortical organoid generation