(175at) A Triple-Input Microfluidic Droplet Trapping Array for Multiplexed Single Cell Analysis

An evolving technology to control the cellular environment utilizes the principles of droplet-based microfluidics, where an aqueous flow is segregated into individual droplets within an immiscible carrier fluid to encapsulate cells, organic molecules and reagents. Droplet microfluidic devices offer a wide-range of applications such as high-throughput drug screening, rare cell detection, and single cell DNA amplification. While droplet microfluidic devices have numerous applications, they are oftentimes limited to a single input condition that prevents them from being able to analyze multiple input parameters (e.g., combinations of cellular treatments) in a single experiment. Previous work from our group demonstrated the feasibility of utilizing photoluminescent rare earth (RE)-doped nanoparticles (NPs) as optical tags for simultaneous droplet tracking as a method to identify droplets generated from different input conditions. Similarly, we demonstrated the utility of a microfluidic droplet trapping array capable of generating 70 µm aqueous droplet in a 775member array with ~40-50% single cell trapping efficiency. The goal of this work is to expand upon the previous studies by fabricating a triple-input microfluidic droplet trapping array capable of high-throughput single cell analysis. The multi-input microfluidic device incorporates four input streams: one continuous oil inlet (containing 2% w/w fluorosurfactant) and three aqueous inlets each of which contains three different samples with three different RE-doped NPs. Each of the three aqueous inlets intersects with the oil channel at three T-junctions positioned in series upstream of a large overhead trapping array to collect single droplet for on-chip interrogation and analysis. This device utilized gravity-induced flow for droplet generation to reduce the number of syringe pumps needed to operate the system. In order to determine optimal operating parameters, a combination of mathematical modeling and experimental studies of the fluid properties were performed to investigate the effect of the capillary number (Ca), the channel geometry (cross section aspect ratio­), and the flow rate (disperse-to-continuous phases) ratio on the dynamics of the droplet breakup. The spacing between the three T-junctions and the dimensions of the device were further optimized using COMSOL Multiphysics to ensure rapid and monodisperse droplet generation. The downstream trapping array has a capacity for isolating ~3500 droplets, which are separated from the bulk solution due to the difference in densities between the aqueous and oil phases. This array allows for time-dependent single cell analysis without the need for expensive high-speed cameras. The work described here is a first step towards a fully characterized, multi-input microfluidic droplet trapping device capable of performing high-throughput screening of individual cellular behavior under a variety of cancer therapeutics simultaneously in a time and cost efficient manner.