Enhanced contactless dielectrophoresis enrichment and isolation platform via cell-scale microstructures

Abstract

We designed a new microfluidic device that uses pillars on the same order as the diameter of a cell (20 μm) to isolate and enrich rare cell samples from background. These cell-scale microstructures improve viability, trapping efficiency, and throughput while reducing pearl chaining. The area where cells trap on each pillar is small, such that only one or two cells trap while fluid flow carries away excess cells. We employed contactless dielectrophoresis in which a thin PDMS membrane separates the cell suspension from the electrodes, improving cell viability for off-chip collection and analysis. We compared viability and trapping efficiency of a highly aggressive Mouse Ovarian Surface Epithelial (MOSE) cell line in this 20 μm pillar device to measurements in an earlier device with the same layout but pillars of 100 μm diameter. We found that MOSE cells in the new device with 20 μm pillars had higher viability at 350 VRMS, 30 kHz, and 1.2 ml/h (control 77%, untrapped 71%, trapped 81%) than in the previous generation device (untrapped 47%, trapped 42%). The new device can trap up to 6 times more cells under the same conditions. Our new device can sort cells with a high flow rate of 2.2 ml/h and throughput of a few million cells per hour while maintaining a viable population of cells for off-chip analysis. By using the device to separate subpopulations of tumor cells while maintaining their viability at large sample sizes, this technology can be used in developing personalized treatments that target the most aggressive cancerous cells.

FIG. 1.

20 μm pillar contactless dielectrophoresis device. (a) Top view photo and (b) exploded schematic of a cross section across a row of pillars with white space added to visually separate layers. The main channel is colored green and contains inlets for both cell suspension and DEP buffer. Electrode channels are colored purple, they are filled with 10 × PBS and connected to the high voltage generator. An electric field is applied across each chamber, perpendicular to the direction of fluid flow.

FIG. 2.

Cross-section of the ratio of the gradient electric field squared to electric field $(\nabla {\left|E\right|}^2)/E$ in front of a pillar. (a) Diagram of the pillar layout in the device. (b) The ratio $(\nabla {\left|E\right|}^2)/E$ in front of 20 μm and 100 μm pillars. At the same voltage to distance ratio, the gradient is much steeper in the 20 μm pillar device than in 100 μm pillar device, reducing accumulation of excessive cells at each pillar. Note that both designs have a 60 μm gap between pillars.

FIG. 3.

Gradient of electric field squared $\nabla {\left|E\right|}^2$ (dark blue arrows) and velocity of fluid flow u_f (light red arrows) in a device with ((a) and (c)) 100 μm pillars and ((b) and (d)) 20 μm pillars, when no cells are present ((a) and (b)) and when one cell is already trapped ((c) and (d)). Insulating pillars are shaded with yellow and cells with light blue. Log values of both vectors are plotted however they are scaled arbitrarily (not to compare size of $\nabla {\left|E\right|}^2$ and u_f) while the same scale is used in all panels. If a cell is already trapped in front of a pillar the next cells will be exposed to different forces. In front of a large 100 μm pillar there is enough space that cells can form a chain, while in front of a small 20 μm pillar the balance of DEP and drag force is different and cells are unlikely to form a chain.

FIG. 4.

Photo of trapped cells in cDEP devices. Direction of fluid flow is from top to bottom, while electric field is applied horizontally. (a) High density of cell suspension in a device with large (100 μm) pillars results in pearl-chaining and unspecific trapping. (b) In a device with small 20 μm pillars only one or two cells can trap on each pillar. When saturated, excessive cells will not accumulate, but continue to next available location.

FIG. 5.

Viability of MOSE-L-_FFLv cells in devices with 100 and 20 μm pillars at flow rate of 20, 28, and 36 μl/min, voltage of 0, 250, 300, and 350 V_RMS and frequency of 30 kHz. The experiment was repeated 3 times for each set of parameters. (a) 100 μm pillar device, (b) 20 μm pillar device, (c) Viability of trapped cells, grouped by flow rate. 20 μm pillar device has little effect on viability at all voltages and flow rates, while the 100 μm pillar device significantly reduces cell viability. In both devices cell viability decreases with increasing voltage and flow rate.

FIG. 6.

Percentage of trapped MOSE-L-_FFLv cells in devices with 100 and 20 μm pillars. At the same voltage, flow rate and density of cell suspension the device with small pillars can trap up to 6 times more cells than the device with large pillars.