An Air Bubble-Isolating Rotating Wall Vessel Bioreactor for Improved Spheroid/Organoid Formation

Rotating wall vessel (RWV) bioreactors have been used to produce cell spheroids and organoids at a faster rate than in other bioreactor devices and with higher structural and functional fidelity. One of the limitations of traditional RWV systems is the well-documented tendency for air bubble formation during operation. The presence of these bubbles negates key features of the RWV environment, such as zero headspace, low-shear, and simulated microgravity. In this article, we describe the design, construction, and testing of a novel RWV capable of constantly removing air bubbles from the system without interfering with the fluid dynamics that produce optimized cell culture conditions. We modeled this capacity using computational fluid dynamics and then validated the model with alginate beads and spheroid cultures of A549 human lung adenocarcinoma cells. The areas of spheroids assembled from A549 cells in the novel bioreactor in the presence of air bubbles were an order of magnitude larger than in conventional bioreactors when bubbles were present. Our results demonstrate the ability of the novel design to remove and isolate bubbles while avoiding damage to spheroid assembly, as observed in conventional RWV bioreactors in the presence of bubbles. We anticipate that the novel design will increase experimental reproducibility and consistency when using RWV bioreactors.