Matrigel Solution Whole Brain Cerebral Organoids for Increased Throughput | AIChE

Matrigel Solution Whole Brain Cerebral Organoids for Increased Throughput


Estridge, R. C. - Presenter, North Carolina State University
Keung, A. J., North Carolina State University
Rao, B., North Carolina State University
Human cerebral organoids have become a popular model for studying neurodevelopment and diseases as they can be readily generated from embryonic and induced pluripotent stem cells. Established whole brain cerebral organoids require a process of Matrigel, a gelatinous mixture of extracellular matrix proteins and growth factors secreted by Engelbreth-Holm-Swarm mouse sarcoma cells that promotes polarization and supports organoid buds, embedding. However, this tedious, most time-intensive part of whole brain cerebral organoid generation process restricts the throughput, limiting the number of samples and/or conditions that can be tested. To address this, we instead cultured whole brain cerebral organoids in various concentrations of Matrigel solution and lengths of time. The Lancaster protocol was followed until the Matrigel embedding step. A small amount of Matrigel was then mixed with cerebral organoid differentiation media, added to the organoids, and the organoids were placed on an orbital shaker. The organoids were then imaged over time using a brightfield microscope and their size calculated using ImageJ. The organoids cultured in the Matrigel solution exhibited faster growth and larger size than embedded organoids. Moreover, some organoid conditions exhibited large, fluid-filled cavities. Additionally, organoids at days 25 and 50 were immunostained for maturation (SOX2 and TUJ1), cell type (MAP2, TBR1, FOXA2, FOXG1), and cell death (Caspase-3) markers. The Matrigel solution organoids displayed organized structures and similar cellular diversity to the embedded control. The larger, fluid filled organoids exhibited increased cell death. Using Matrigel solution to culture human whole brain cerebral organoids will allow for expanded research avenues of human neurodevelopment.