High Density 3D Printed Microfluidics for Cell-Based Biomedical Applications

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Project Summary Concentration gradients of growth factors, other cell-signaling molecules, and nutrients drive a wide range of critical biological processes, including immune cell migration, angiogenesis, wound healing, cancer metastasis, and organism development. Microfluidic devices are extensively used to both create the relevant concentration gradient, and to monitor cellular behavior in response to that gradient. Unfortunately, most concentration gradient microfluidic devices are designed assuming exclusively diffusional transport processes, while incorporating advective transport decoupling strategies that are largely ineffective and/or slow. This approach results in specific and important drawbacks, including low dynamic range in the gradient, unstable gradients over the lifetime of the measurement, and inconsistency in the gradient between different spatial regions of the device. As a result, existing microfluidic approaches to concentration gradient construction do not adequately mimic the diffusion- generated concentration gradients found in-vivo. Hence, there is a large unmet need for microfluidic devices that can rapidly create stable and flexible concentration gradients that allow sensitive monitoring of critical cellular processes. This renewal proposal focuses on leveraging and extending sophisticated high resolution 3D printing technology for microfluidics to create integrated devices that generate concentration gradients with large dynamic range that are switchable between multiple source and sink solutions with selectable concentration and rapid set up time (few minutes) to enable temporal multiplexing of sequences of stable concentration gradients. Research efforts will consist of three specific aims. First, 3D simulation will be employed to evaluate a wide range of concentration gradient formation geometries based on a new opposing-flow concept for source and sink fluids with the objective of decoupling advective from diffusive mass transport, where the former is needed to replenish source and sink fluids while the latter is required to generate the concentration gradient. Next, a variety of candidate geometries will be 3D printed and tested to iteratively optimize concentration gradient dynamic range, set up time, stability, and uniformity. The best candidates will be integrated with on-chip pumps, valves, serial diluters, and reservoirs to create integrated systems. Finally, such devices will be used to analyze chemotaxis of metabolically engineered bacteria. The new capabilities developed over the grant period are designed to allow important questions to be answered with respect to developmental biology, cellular response to nutritive cues, as well as angiogenesis, and cancer growth and invasiveness.