A Membraneless Continuous-Flow Filter for High-Throughput Separation and Enrichment of Particles and Cells

There is a critical need for advanced filtration methods adaptable for separation of particles, cells, and cell-sized components from complex fluid mixtures?specifically those offering the capability to rapidly process large sample volumes (> mL/min flow rates). Microfluidic technologies provide a natural platform to address these challenges, but most of these efforts have yet to advance past the proof of concept stage.

Here we describe a new microfluidic-based filtration method capable of performing simultaneous size-based isolation and enrichment of particles and cells. Instead of forcing a particle- or cell-laden suspension to flow through tiny pores in a membrane filter, we are able to construct a filter oriented along the centerline of the microchannel so that it creates a barrier between the left and right hand sides. When this geometry is incorporated into a curved flow path, the resulting centrifugal forces that arise due to fluid motion act to push the suspended components across the centerline barrier from the inside wall to the outside wall, with only those smaller than the barrier gap able to pass across. Consequently, this filtration method does not impose an excessive pressure drop because the barrier is oriented parallel to the flow direction rather than perpendicular to it. Moreover, this approach is most effective at high flow rates because the magnitude of the curvature-induced transverse flow is maximized under these conditions, making it ideally suited for high-throughput analysis of large sample volumes.

To construct microchannel networks incorporating embedded centerline barriers, we have developed a new microfabrication approach that capitalizes on the properties of biodegradable poly(lactic acid) (PLA). The process works by perfusing the surface of a PLA substrate with an enzymatic agent capable of cleaving lysine-lysine bonds (e.g. proteinase-k). By manipulating parameters associated with the laminar flow enzymatic degradation process (e.g., temperature, degradation time, enzyme concentration, etc.), complex microchannel topologies can be precisely etched in PLA sheets without the multiple lithography steps that would otherwise be needed.

Filtration capability was evaluated by injecting a mixture of fluorescent polystyrene beads of diameter 3 and 10 µm into the inner inlet at a flow rate of 1 ml/min. Samples were then collected at outlets positioned at the inner and outer sides of the centerline barrier and analyzed by flow cytometry. The inner outlet contained the 10 µm beads that were unable to pass across the barrier, while the outer outlet contained only 3 µm beads. In addition to demonstrating extremely high selectivity, no clogging effects were observed because the primary flow acts to sweep aggregates downstream.

The design of our membraneless filter uniquely merges the most favorable aspects of (1) continuous operation at high flow rates (necessary for high-throughput analysis of large-volume samples) with (2) the high selectivity of a physical membrane barrier (i.e., eliminating the need for expensive antibody-based affinity capture). In addition to isolation and enrichment, our design can enable viable cells to be recovered for subsequent analysis (a capability not readily available in many comparable devices).