Fig. 1.

Microwell fabrication and use in human cerebral organoid culture in this study. A) Schematic of microwell fabrication and post-treatment. The reverse mold (micropillars) was 3D-printed. By molding in PDMS through soft lithography, the microwell platform was created, followed by a surface coating of mPEG, Lipidure, or BSA, and then the embryonic stem cell (H9) culturing. B) Timeline of the microwell culturing and human cerebral organoid generation. hESCs (H9) were seeded at 0.2 million cells per microwell device and cultured for three days for EB formation. The formed EBs were then transferred to T25 flasks for prolonged culture and neural induction. Cerebral organoid formation and maturation were then assessed on day 20 and day 45 by whole organoid staining and cyto-sectioning, respectively. NIM medium was used throughout the entire culture process, and B27 was added since day 20.