Abstract
Living cells reside within anisotropic microenvironments that orchestrate a broad range of polarized responses through physical and chemical cues. To unravel how localized chemical signals influence complex behaviors, tools must be developed for establishing patterns of chemical gradients that vary over subcellular dimensions. Here, we present a strategy for addressing this critical need in which an arbitrary number of chemically distinct, subcellular dosing streams are created in real time within a microfluidic environment. In this approach, cells are cultured on a thin polymer membrane that serves as a barrier between the cell-culture environment and a reagent chamber containing multiple reagent species flowing in parallel under low Reynolds number conditions. Focal ablation of the membrane creates pores that allow solution to flow from desired regions within this reagent pattern into the cell-culture chamber, resulting in narrow, chemically distinct dosing streams. Unlike previous dosing strategies, this system provides the capacity to tailor arbitrary patterns of reagents on the fly to suit the geometry and orientation of specific cells.