Optimizing microfluidic chip for continuous perfusion of retinal organoids

Abstract

Purpose : Stem-cell derived retinal organoids are a potential transplantable therapeutic for patients with advanced retinal degeneration. However, retinal organoid manufacturing methods are highly heterogeneous. Autonomous perfusion-based microfluidic bioreactors can facilitate the manufacturing, quality control, and selection of organoids for downstream applications. We evaluated whether parallel-based microfluidic would more homogenously perfuse organoids than serial-based microfluidics.

Methods : A microfluidic chip mold was designed in SolidWorks and fabricated using Formlabs Form 3B 3D-Printer. Polydimethylsiloxane (PDMS) was cast from the mold. Plasma cleaning of the PDMS device promoted a hydrophilic surface and enhanced adhesion with an adhesive film. We performed a dye perfusion test in which the microfluidic devices were prefilled with yellow dye and green dye was subsequently perfused through the device. Perfusion was performed for 45 minutes at a flow rate of 150 μL/hr followed by a degassing step to remove any residual bubbles. Photographs of dye-perfused chips were evaluated for grayscale values using ImageJ. Box-and-whisker plots were generated to evaluate perfusion uniformity in parallel and serial perfused organoid culture wells

Results : Parallel-based perfusion consistently showed significantly less data variability throughout the 45 minute perfusion test when compared to serial-based perfusion. Thin layers of air bubble were seen in some wells of both parallel and serial design chips.

Conclusions : Based on the perfusion dye test, parallel-based microfluidic design allows more uniform perfusion conditions than serial perfusion. Further exploration is needed to identify effective methods to permanently eliminate air bubbles in organoid culture chambers.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

 

Figure 1: Perfusion data. A) Diagram of parallel and serial microfluidic chips with flow arrows. B) Post-experiment degassing images of parallel and serial chips. C) Box-and-whisker plots assessing the perfusion uniformity for both chips.