Characterization and Separation of Live and Dead Yeast Cells Using CMOS-Based DEP Microfluidics

Abstract

This study aims at developing a miniaturized CMOS integrated silicon-based microfluidic system, compatible with a standard CMOS process, to enable the characterization, and separation of live and dead yeast cells (as model bio-particle organisms) in a cell mixture using the DEP technique. DEP offers excellent benefits in terms of cost, operational power, and especially easy electrode integration with the CMOS architecture, and requiring label-free sample preparation. This can increase the likeliness of using DEP in practical settings. In this work the DEP force was generated using an interdigitated electrode arrays (IDEs) placed on the bottom of a CMOS-based silicon microfluidic channel. This system was primarily used for the immobilization of yeast cells using DEP. This study validated the system for cell separation applications based on the distinct responses of live and dead cells and their surrounding media. The findings confirmed the device's capability for efficient, rapid and selective cell separation. The viability of this CMOS embedded microfluidic for dielectrophoretic cell manipulation applications and compatibility of the dielectrophoretic structure with CMOS production line and electronics, enabling its future commercially mass production.

Keywords: CMOS-based lab-on-a-chip; cell characterization; cell separation; dielectrophoresis (DEP); interdigitated electrodes (IDEs); microfluidics.

Figure 1

(a) Microfluidic device with isolated fluidic and electrical interfaces; (b) Schematic cross-sectional view of the device with one IDE structure.

Figure 2

Microfluidic device in operation: cell entrapment at the IDEs after imposing AC and generating DEP.

Figure 3

The Clausius-Mossotti factor as a function of frequency for live and dead yeast cells suspended in: (a) DIW; (b) Tap-water; (c) KCl; (d) PBS; (e) D-PBS. Electrical and geometrical properties of the live and dead yeasts were taken from [79]. Solid lines and dotted lines represent the real and imaginary parts of the Clausius-Mossotti factor, respectively.

Figure 4

FEM simulation results for cell separation. DEP generated by IDEs, shown in black and white segments (marked by −V and +V, respectively). The line contour illustrates the electric potential applied to the IDEs, and red arrows represent the electric field distribution.

Figure 5

Empirically proven pDEP regime and crossover frequencies (fc) vs. numerical predictions for live yeasts suspended in DIW. Colored frequency bands represent trapping behavior based on microscopical observations.

Figure 6

DEP characterization of live yeasts in DIW at 1 µm s⁻¹ flow rate: (a) Trapping rate (yield) approximation by changing frequency based on covered cell area at the IDEs; (b–g) the images show the trapping at various frequencies after 1 min.

Figure 7

Empirically proven pDEP regime and crossover frequencies (fc) vs. numerical predictions for: (a) dead yeast suspension in DIW and live cell suspensions in; (b) Tap water; (c) KCL; (d) D-PBS. Colored frequency bands represent empirically proven trapping behavior based on optical observations.

Figure 8

DEP behavior of live yeasts in different medium solutions at 20 Vpp and 1 µm s⁻¹ flow rate: (a–c) pDEP, which leads to cell trapping and; (d) nDEP and no cell trapping.

Figure 9

Experimentally measured (pink and gray zones) and numerically predicted (lines) affective pDEP for live and dead cells in 0.0002 S/m DIW at different AC field frequencies at the microfluidic channel using pDEP.

Figure 10

Experimentally recorded results on selectively trapping and separation of live cells from a live and dead yeast mixture (0.1282 S/m) suspended in DIW at a constant voltage of 20 Vpp: (a) Isolation efficiency of live cells from a mixture using IDEs; (b–e) Micrograph examples where the percentage of isolated live cells increase by increasing frequency (b) 0%; (c) 22.22%; (d) 77.35%; (e) 80.19%; (f) 91.66%; and (g) 100%. White cells are Live, and blue-dyed cells are dead cells. The live and dead cells mixture was 1:1 in the suspension prior to separation.

Figure 11

Differential separation of the live yeasts from live and dead (1:1) cell mixtures by DEP at 20 V_pp, 20 MHz while dead cells were removed by nDEP: (a) cell mixture in KCL; (b) cell mixture in D-PBS.

Figure 12

Separation estimation of live/dead yeasts in a mixture. The live and dead cells mixture was 3:4 in the suspension before separation: (a) Viability estimation before (Pre-DEP) and after measurement (Post-DEP); (b) Percentage of trapped and released live and dead cell densities after DEP. The separation condition was: 20 Vpp, 1MHz, and 1 µm·s⁻¹ flow rate.

Figure 13

Cell density assay before and after DEP. The cells were immobilized using pDEP at 1 MHz, 20 V_pp: (a) Approximate cell density before (Pre-DEP) and after measurement (Post-DEP); (b) Percentage of trapped and released cell density after DEP.