Versatile size-dependent sorting of C-elegans nematodes and embryos using a tunable microfluidic filter structure

The roundworm Caenorhabditis elegans (C. elegans) is a powerful model organism for addressing fundamental biological questions related to human disease and aging. Its life cycle consists of an embryo stage, four larval stages that can be clearly distinguished by size and different morphological features, and adulthood. Many worm-based bio-assays require stage-or age-synchronized worm populations, for example for studying the life cycle and aging of worms under different pharmacological conditions or to avoid misinterpretation of results due to overlap of stage-specific response in general. Here, we present a new microfluidic approach for size-dependent sorting of C. elegans nematodes on-chip. We take advantage of the external pressure-deformable profile of polydimethylsiloxane (PDMS) transfer channels that connect two on-chip worm chambers. The pressure-controlled effective cross-section of these channels creates adjustable filter structures that can be easily tuned for a specific worm sorting experiment, without changing the design parameters of the device itself. By optimizing the control pressure settings, we can extract larvae of a specific development stage from a mixed worm culture with an efficiency close to 100% and with a throughput of up to 3.5 worms per second. Our approach also allows us to generate mixed populations of larvae of adjacent stages or to adjust their ratio directly in the microfluidic chamber. Moreover, using the same device, we demonstrated extraction of embryos from adult worm populations for subsequent culture of accurately age-synchronized nematode populations or embryo-based assays. Considering that our sorting device is merely based on geometrical parameters and operated by simple fluidic and pressure control, we believe that it has strong potential for use in advanced, automated, microfluidic C. elegans-based assay platforms.