(K-412) Development and Fabrication of a Microfluidic Device for Trapping and Analysis of Hydrogel Encapsulated Cells

Microfluidics presents promising potential for replicating biological functions and conducting experiments in a controlled environment. An advantage of using microfluidics is the versatility of the designs and fabrication process that optimizes the localization and real-time analysis of an object of interest. In this study, a cytocompatible microfluidic device is developed to trap hydrogel-encapsulated cells under flow conditions, enabling real-time analysis of the cells and the collection of biologics for multiple days (Figure 1A). The primary objective of the study is to overcome challenges associated with conventional microfluidic devices by improving the transparency of 3D-printed fluidic chips, minimizing leakage during long-term perfusion, and optimizing capsule trapping efficiency. The final prototype of the device uses a gravity-driven mechanism to enhance the loading and imaging of encapsulated cells, as well as enable the collection of trapped capsules after study completion for further analysis. The study will outline the design and fabrication process, as well as data demonstrating the cytocompatibility of trapped encapsulated cells and functional uses such as in-situ imaging and measurements of biologics collected under flow conditions.

We designed and fabricated a transparent microfluidic device for gravity-driven trapping of spherical alginate hydrogel capsules and culture under flow conditions. The device is designed with Fusion360 and 3D printed from a biocompatible resin using a Formlabs Form 3B stereolithography printer. After printing, the fluidic chips were post-processed with isopropyl alcohol and UV cured to ensure complete curing of the resin and cytocompatibility. To test the device’s performance, we validated robust leak-free microchannel perfusion under various media flow rates. We then measured the efficiency of trapping hydrogel capsules and the maximum flow rate at which capsules would remain in the microchambers. To ensure cytocompatibility of the device design and material, we trapped and cultured encapsulated human epithelial cells (ARPE-19) for 24 hours under constant flow and compared the cell viability of microfluidic cultured capsules with capsules cultured under normal static conditions. Cell viability was analyzed using live-dead staining and imaging, as well as counting live and dead cells from lysed capsules with an automated cell counter. Finally, we demonstrated the capability for analysis of secreted biologics from collected media fractions using ELISA. 

Results & Discussions

The final prototype contains one microchannel and 11 microchambers that vertically branch off from the channel (Figure 1A). This vertical orientation improved the trap efficiency and stability of 1.5mm hydrogel capsules under a higher media flow rate (Figure 1A). A long-term perfusion study demonstrated it is possible to run an overnight study without leakage and capsules escaping the microchambers. Furthermore, there was no statistical difference in cell viability from capsules cultured under normal static conditions compared to capsules cultured within the microfluidic device under flow conditions (Figure 1B, 1C). We measured the concentration of IgG antibodies secreted from encapsulated therapeutic cells in flow fraction samples to compare differences in productivity and release rate between two different alginates. We discovered that both alginates had similar release kinetics but the modified alginate demonstrated enhanced per-cell productivity.

The microfluidic device and fabrication methodology employed herein can provide a multitude of advantages for experiments requiring analysis of encapsulated cell biologic release kinetics and potency. Furthermore, a glass slide that was attached at the bottom of the device allowed real-time imaging of the capsules without retrieving them from the device. Specifically tailored to optimize capsule localization, prevent leakage, and facilitate imaging, this device provides a platform for real-time analysis of the rate of therapeutic productions under flow conditions for multiple days. 

Conclusion

The study aimed to develop a microfluidic device for the characterization of the release kinetic of biologics from cells by addressing existing challenges in microfluidic devices designed to measure cell potency. The localization of the capsules was facilitated by gravity, which was achieved through the vertical geometry of the chambers. Additionally, the microfluidic device was designed as an integrated device, reducing leakage during experiments. Cytocompatibility tests successfully demonstrated high cell viability. Building upon previous microfluidic devices for cell potency measurement, the presented approach offers a potential platform for the measurement of therapeutic production by encapsulated cells that pushes beyond established methods and facilitates kinetic analysis of media flow fraction.