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Brain organoids derived from human induced pluripotent stem cells can recapitulate the early stages of brain development, representing a powerful in vitro system for modeling brain development and diseases. However, the existing methods for brain organoid formation often require time-consuming procedures, including the initial formation of embryoid bodies (EBs) from hiPSCs, and subsequent neural induction and differentiation companied by multi-steps of cell transfer and encapsulation in a 3D matrix. Herein, we propose a simple strategy to enable in situ formation of massive brain organoids from hiPSCs on a micropillar array without tedious manual procedures. The optimized micropillar configurations allow for controlled EB formation, neural induction and differentiation, and generation of functional human brain organoids in 3D culture on a single device. The generated brain organoids were examined to …