High‐throughput dielectrophoretic separator based on printed circuit boards

A review is presented of the present status of the theory, the developed technology and the current applications of dielectrophoresis (DEP). Over the past 10 years around 2000 publications have addressed these three aspects, and current trends suggest that the theory and technology have matured sufficiently for most effort to now be directed towards applying DEP to unmet needs in such areas as biosensors, cell therapeutics, drug discovery, medical diagnostics, microfluidics, nanoassembly, and particle filtration. The dipole approximation to describe the DEP force acting on a particle subjected to a nonuniform electric field has evolved to include multipole contributions, the perturbing effects arising from interactions with other cells and boundary surfaces, and the influence of electrical double-layer polarizations that must be considered for nanoparticles. Theoretical modelling of the electric field gradients generated by different electrode designs has also reached an advanced state. Advances in the technology include the development of sophisticated electrode designs, along with the introduction of new materials (e.g., silicone polymers, dry film resist) and methods for fabricating the electrodes and microfluidics of DEP devices (photo and electron beam lithography, laser ablation, thin film techniques, CMOS technology). Around three-quarters of the 300 or so scientific publications now being published each year on DEP are directed towards practical applications, and this is matched with an increasing number of patent applications. A summary of the US patents granted since January 2005 is given, along with an outline of the small number of perceived industrial applications (e.g., mineral separation, micropolishing, manipulation and dispensing of fluid droplets, manipulation and assembly of micro components). The technology has also advanced sufficiently for DEP to be used as a tool to manipulate nanoparticles (e.g., carbon nanotubes, nano wires, gold and metal oxide nanoparticles) for the fabrication of devices and sensors. Most efforts are now being directed towards biomedical applications, such as the spatial manipulation and selective separationenrichment of target cells or bacteria, high-throughput molecular screening, biosensors, immunoassays, and the artificial engineering of three-dimensional cell constructs. DEP is able to manipulate and sort cells without the need for biochemical labels or other bioengineered tags, and without contact to any surfaces. This opens up potentially important applications of DEP as a tool to address an unmet need in stem cell research and therapy.

Isolation and enumeration of circulating tumor cells (CTCs) are used to monitor metastatic disease progression and guide cancer therapy. However, currently available technologies are limited to cells expressing specific cell surface markers, such as epithelial cell adhesion molecule (EpCAM) or have limited specificity because they are based on cell size alone. We developed a device, ApoStream(™) that overcomes these limitations by exploiting differences in the biophysical characteristics between cancer cells and normal, healthy blood cells to capture CTCs using dielectrophoretic technology in a microfluidic flow chamber. Further, the system overcomes throughput limitations by operating in continuous mode for efficient isolation and enrichment of CTCs from blood. The performance of the device was optimized using a design of experiment approach for key operating parameters such as frequency, voltage and flow rates, and buffer formulations. Cell spiking studies were conducted using SKOV3 or MDA-MB-231 cell lines that have a high and low expression level of EpCAM, respectively, to demonstrate linearity and precision of recovery independent of EpCAM receptor levels. The average recovery of SKOV3 and MDA-MB-231 cancer cells spiked into approximately 12 × 10(6) peripheral blood mononuclear cells obtained from 7.5 ml normal human donor blood was 75.4% ± 3.1% (n = 12) and 71.2% ± 1.6% (n = 6), respectively. The intra-day and inter-day precision coefficients of variation of the device were both less than 3%. Linear regression analysis yielded a correlation coefficient (R(2)) of more than 0.99 for a spiking range of 4-2600 cells. The viability of MDA-MB-231 cancer cells captured with ApoStream was greater than 97.1% and there was no difference in cell growth up to 7 days in culture compared to controls. The ApoStream device demonstrated high precision and linearity of recovery of viable cancer cells independent of their EpCAM expression level. Isolation and enrichment of viable cancer cells from ApoStream enables molecular characterization of CTCs from a wide range of cancer types.