Optimization and Parallelization of Sorting by Interfacial Tension (SIFT) for High-Throughput Metabolic Cell Sorting

A systematic optimization of throughput and operational stability in Sorting by Interfacial Tension (SIFT) is presented. Reducing droplet size and enabling a broader distribution of droplet trajectories increased the number of droplets processed per sorting element, resulting in about a four fold improvement in throughput from 30 to 125 droplets per second. Throughput was further enhanced through device parallelization, with devices incorporating two and four independent sorting regions demonstrated. These configurations distributed droplets evenly across sorting elements that exhibited comparable pH sorting thresholds, indicating similar flow conditions and drag forces within each region. Among the designs evaluated, the two-element configuration provided the optimal balance of throughput, robustness, and simplicity, achieving maximum throughputs of about 250 droplets per second. Throughput and pH sorting thresholds were preserved throughout two hours of continuous sorting. The improved platform was applied to examine the relationship between cellular glycolysis and iron homeostasis at the single-cell level for Jurkat cells, revealing a subpopulation of highly glycolytic cells with significantly elevated iron uptake, consistent with prior reports linking iron regulation and T cell metabolism. Collectively, these advances expand the scale, stability, and biological applicability of SIFT, enabling large-scale functional studies while facilitating the capture of rare and metabolically distinct cell populations.