Human brain organoids provide opportunities to produce three-dimensional (3D) brain-like tissues for
biomedical research and translational drug discovery, toxicology, and tissue replacement. Here we describe
a protocol for rapid and defined induction of brain organoids from human induced pluripotent stem cells
(iPSCs), using commercially available culture and differentiation media and a cheap, easy to handle and
clinically approved semisynthetic hydrogel. Importantly, the methodology is uncomplicated, well-defined,
and reliable for reproducible and scalable organoid generation, and amendable to principles of current good
laboratory practice (cGLP), with the potential for prospective adaptation to current good manufacturing
practice (cGMP) toward clinical compliance.