THE CELLXPRESS.AI AUTOMATED CELL CULTURE SYSTEM FOR AUTOMATED, ROBUST FOREBRAIN AND MIDBRAIN ORGANOID GENERATION

Human brain organoids are powerful in vitro models for studying neurodegenerative diseases, but their adoption is limited by labor‑intensive workflows, and poor scalability. Precise control of seeding, differentiation timing, long‑term culture conditions, and media handling over extended periods remains a challenge.
We present fully automated workflows for robust forebrain and midbrain organoid generation using the CellXpress.ai™ Automated Cell Culture System. Forebrain organoids were generated from pluripotent stem cells seeded into 96‑well U‑bottom plates for embryoid body formation and neural induction, followed by automated transfer to 6‑well plates for long‑term maturation on in‑incubator rocking racks. Continuous media agitation supported uniform growth and neuronal activity comparable to standard agitation methods.
Midbrain organoids were generated from small‑molecule neural precursor cells (smNPCs) entirely in a 96‑well format. Complex differentiation protocols, including accurate seeding, timed media exchanges, daily maintenance, and imaging were automated, minimizing operator‑dependent variability.
The CellXpress.ai system’s cell journey feature enabled continuous tracking of individual organoids from seeding through maturation, providing integrated quality control and improved reproducibility. Organoids showed homogeneous morphology and reduced inter‑and intra‑batch variability. Differentiation was confirmed by whole‑mount immunostaining for key neural markers, and neuronal activity was assessed using calcium imaging on the ImageXpress® Confocal HT.ai High‑Content Imaging System.
Finally, we demonstrate an automated drug‑screening workflow integrating robotic plate handling and high‑content imaging, highlighting the feasibility of high‑throughput organoid‑based assays for neurodegenerative disease modeling and drug discovery.