There are a number of techniques for the formation of human organoids and neurospheres in culture, and they typically fall within the patterned or unpatterned categories. Patterned methods implement factors to regulate Wnt, BMP, TGFβ, and other signaling pathways to push differentiation toward specific lineages23. Unpatterned methods, such as the one described here, take advantage of the propensity for induced pluripotent stem cells (iPSCs) and human embryonic stem cells (hESCs) to differentiate toward a neuroectodermal lineage by default24. After approximately three weeks of differentiation, the resulting organoids consist of large, biomimetic neuroepithelial structures that contain several cell types that are observed in the early developing brain.
