(240g) A New Strategy for Sample Concentration and Enrichment: Contactless Dielectrophoresis

The polarizable cells/micro particles exposed in non-uniform electric field can be manipulated based on their electrical signatures and size using dielectrophoresis (DEP) [1]. DEP has been used effectively to isolate target cells including the separation of human leukemia cells from red blood cells in an isotonic solution[2], entrapment of human breast cancer cells from blood[3], separation of human monocytic from peripheral blood mononuclear cells (PBMC)[4], separation of bacteria from blood[5], cancer cells from CD34+ hematopoietic stem cells[6], and isolate mammalian cells based on their cell-cycle phase[7, 8]. However, direct contact between the electrodes and the biological sample in the traditional DEP causes unpredictable chemical reactions, fouling, contamination, bubble formation near integrated electrodes, and an expensive and complicated fabrication process [9, 10]. We have developed an alternative method to provide the spatially non-uniform electric field required for DEP in which electrode sample direct contact is avoided[11]. In this method, electrodes are inserted into side channels which are separated from main channel by thin insulating PDMS barriers. The side channels are filled with high conductive solution such as PBS, and the biological sample is injected into the main channel. The insulating barriers exhibit a capacitive behavior and therefore an electric field can be produced in the main channel by applying an AC field across the barriers (Fig.1a). The absence of contact between electrodes and the sample fluid inside the channel prevents bubble formation and avoids any contaminating effects the electrodes may have on the sample. We have designed and fabricated a microfluidic device (Fig.1a) based on this new technique and we have observed DEP responses in human leukemia cancer cells. These cells were observed to be trapped due to positive DEP force at 150kHz and 215Vrms applied voltage across the side channels 1, 2 and 3 (Fig.1b-g). Particles parallel to the electric field attract each other because of this dipole-dipole force, resulting in pearl-chaining of the trapped cells in the direction of the electric field in the microfluidic channel [1, 12-15]. We have also observed cell chain formation in our experiments with an applied AC electric filed (Fig.1d and 1g). The microfluidic device fabrication process can be broken down into 4 steps: 1) Microfluidic designs patterned on a silicon wafer using Deep Reactive Ion Etching (DRIE). 2) PDMS device created from the wafer stamp. 3) Fluidic connections punched through PDMS device with blunt needles. 4) Glass slides bonded were to the PDMS device using plasma cleaner. The THP-1 human Leukemia monocyte cells were washed twice and resuspended in our prepared DEP buffer (8.5% sucrose [wt/vol], 0.3% glucose [wt/vol], and 0.725% [vol/vol] RPMI). The electrical conductivity of the buffer was measured with a Mettler Toledo SevenGo pro conductivity meter (Mettler-Toledo, Inc., Columbus, OH) to ensure that its conductivity was 100μS/cm. These cells were observed to be spherical when in suspension. The average diameter of these cells is 15.4±2 μm (n=30).The cell were observed to be spherical in suspension. The microfluidic device was modeled numerically in Comsol multi-physics 3.4 using the AC/DC module (Comsol Inc., Burlington, MA, USA). Because DEP depends on the gradient of the electric field ∇E=-∇(∇∅) , the first step in modeling is to determine the electric field distribution within a channel's geometry. This is done by solving for the potential distribution, , using the Laplace equation, ∇^2∅=0. The boundary conditions used are prescribed uniform potentials at the inlet or outlet of the side channels. The numerical and experimental results for the boundary conditions V1=215Vrms at 150kHz, V3=Ground, and V4=open are given in Figures 1b-d. Figures 1b and c show the surface and line plot of the gradient of the electric field along the centerline the main microfluidic channel. There is a high gradient of electric field inside the main channel and between the side channels 1 and 3. The THP-1 cells were observed to be trapped at this area because of positive DEP (Fig.1d). Figure 1e-g show the numerical and experimental results for the boundary conditions V1=V2=215Vrms at 150kHz, V3=Ground, and V4=open. The line plot of the gradient of the electric field shows that there is higher gradient of the electric field compare to the other boundary conditions set up. More cells were observed to be trapped experiencing positive DEP with the second boundary conditions (Fig.1g) The major advantages of contactless dielectrophoresis over other methods of cell sorting include a lack of extensive sample preparation (no antibody labeling, one needs only prepare a single cell preparation) and the speed of isolation (minutes versus hours from the time of sample acquisition). This rapid turnaround is extremely important when evaluating gene expression. We believe this new technique has many attractive features when compared to traditional DEP and insulator based dielectrophoresis (iDEP). Reduced joule heating, no direct electrode contact resulting in less potential for contamination (important for animal studies), simplified inexpensive fabrication process are the other noticeable advantageous of this new techniqueBecause the induced electric field is not as intense as comparable methods and is focused just at the trapping zones, we theorize the joule heating within the main microfluidic channel is insignificant. This could mitigate the stability and robustness issues encountered with conventional iDEP, due to the conductivity distribution's strong dependence on temperature. Furthermore, challenges associated with cell lysing due to high temperatures or irreversible electroporation due to high field strengths are overcome with our new design approach under these conditions. Figure Caption: Figure 1: (a) Schematic of the microfluidic device, (b-d) Numerical and experimental results for V1=215Vrms at 150kHz and V3=0, V2= V4=open, (b) Surface plot of the gradient of the field (kg2mC-2S-4) within the main microchannel, (c) Line plot of the gradient (kg2mC-2S-4) along the line of the main channel , (d) THP-1 cells are trapped due to positive DEP, (e-g) Numerical and experimental results for V1=V2=215Vrms at 150kHz and V3=0,and V4=open, (e) Surface plot of the gradient of the field (kg2mC-2S-4) within the main microchannel (f) Line plot of the gradient (kg2mC-2S-4) along the line of the main channel (g) THP-1 cells are trapped due to positive DEP. References: 1. Pohl, H., Dielectrophoresis. 1978, Cambridge: Cambridge University Press. 2. Becker, F., et al., The removal of human leukaemia cells from blood using interdigitated microelectrodes. J. Phys. D: Appl. Phys, 1994. 27: p. 2659-2662. 3. 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