Non‐Invasive Quality Control of Organoid Cultures Using Mesofluidic CSTR Bioreactors and High‐Content Imaging

Abstract Human brain organoids produce anatomically relevant cellular structures and recapitulate key aspects of in vivo brain function, which holds great potential to model neurological diseases and screen therapeutics. However, the long growth time of 3D systems complicates the culturing of brain organoids and results in heterogeneity across samples hampering their applications. An integrated platform is developed to enable robust and long‐term culturing of 3D brain organoids. A mesofluidic bioreactor device is designed based on a reaction‐diffusion scaling theory, which achieves robust media exchange for sufficient nutrient delivery in long‐term culture. This device is integrated with longitudinal tracking and machine learning‐based classification tools to enable non‐invasive quality control of live organoids. This integrated platform allows for sample pre‐selection for downstream molecular analysis. Transcriptome analyses of organoids revealed that the mesofluidic bioreactor promoted organoid development while reducing cell death. This platform thus offers a generalizable tool to establish reproducible culture standards for 3D cellular systems for a variety of applications beyond brain organoids.