An equilibrium method for continuous-flow cell sorting using dielectrophoresis

Separations represent a fundamental unit operation in biology and biotechnology. Commensurate with their importance is the diversity of methods that have been developed for performing them. One important class of separations are equilibrium gradient methods, wherein a medium with some type of spatial nonuniformity is combined with a force field to focus particles to equilibrium positions related to those particles' intrinsic properties. A second class of techniques that is nonequilibrium exploits labels to sort particles based upon their extrinsic properties. While equilibrium techniques such as iso-electric focusing (IEF) have become instrumental within analytical chemistry and proteomics, cell separations predominantly rely upon the second, label-based class of techniques, exemplified by fluorescence-activated cell sorting (FACS) and magnetic-activated cell sorting (MACS). To extend the equilibrium techniques available for separating cells, we demonstrate the first implementation of a new microfluidic equilibrium separation method, which we call isodielectric separation (IDS), for sorting cells based upon electrically distinguishable phenotypes. IDS is analogous to isoelectric focusing, except instead of separating amphoteric molecules in a pH gradient using electrophoresis, we separate cells and particles in an electrical conductivity gradient using dielectrophoresis. IDS leverages many of the advantages of microfluidics and equilibrium gradient separation methods to create a device that is continuous-flow, capable of parallel separations of multiple (>2) subpopulations from a heterogeneous background, and label-free. We demonstrate the separation of polystyrene beads based upon surface conductance as well as sorting nonviable from viable cells of the budding yeast Saccharomyces cerevisiae.     

Separation of microparticles and biological cells inside an evaporating droplet using dielectrophoresis

Microparticles or biological cells mixed in water were separated using the combination of an electrical force due to dielectrophoresis and a mechanical one generated in an evaporating droplet. Micropatterned electrodes of Au were fabricated on the silicon dioxide layer and were used to generate dielectrophoresis. Polystyrene particles, red blood cells. and E. coli were used as separating objects. Microparticles and biological cells were separated by adjusting the amplitude and frequency of the applied voltage. Although the mechanical force was enough to transport the particles to the boundary of the droplet, nevertheless, it could not detach the particles trapped at the electrode. Based on this work, the microparticles and biological cells can be separated, controlled, and sensed without using a liquid pumping unit.     

Continuous dielectrophoretic size-based particle sorting

Continuous-flow dielectrophoretic (DEP) particle separation based on size is demonstrated in a microfluidic device. Polystyrene microspheres suspended in a neutrally buoyant aqueous solution are used as model particles to study DEP induced by an array of slanted, planar, interdigitated electrodes inside of a soft-lithography microchannel. The E-field gradients from the slanted electrodes impart a net transverse force component on the particles that causes them to "ratchet" across the channel. Over the length of the device, larger particles are deflected more than smaller particles according to the balance of hydrodynamic drag and DEP forces. Consequently, a flow-focused particle suspension containing different-sized particles is fractionated as the beads flow and separate down the length of the device. The flow behavior of spherical particles is modeled, and the total transverse particle displacement in the microfluidic device predicts fourth-order size and voltage and second-order inverse flow rate dependences. The model is verified experimentally for a range of flow rates, particle sizes, and E-field strengths.     

Wafer-level assembly of carbon nanotube networks using dielectrophoresis

We use dielectrophoresis (DEP) to controllably and simultaneously assemble multiple carbon nanotube (CNT) networks at the wafer level. By an appropriate choice of electrode dimensions and geometry, an electric field is generated that captures CNTs from a sizable volume of suspension, resulting in good CNT network uniformity and alignment. During the DEP process, the electrical characteristics of the CNT network are measured and correlated with the network morphology. These experiments give novel insight into the physics of DEP assembly of CNT networks, and demonstrate the scalability of DEP for future device applications.     

Three-dimensional assembly of single-walled carbon nanotube interconnects using dielectrophoresis

We present a hybrid approach that combines top-down fabrication with bottom-up directed assembly for making single-walled carbon nanotube (SWNT) based three-dimensional interconnects. The SWNTs are assembled using dielectrophoresis at room temperature on a microfabricated 3D platform. The two-terminal resistance of the assembled SWNTs at 10?Vpp assembly voltage is approximately 545?Ω. Simulation of the dielectrophoretic assembly is carried out to understand the behavior of the SWNTs during assembly. Encapsulation of these devices using a conformal pinhole-free parylene layer resulted in a decrease of the total resistance.     

Dynamic cell fractionation and transportation using moving dielectrophoresis

This study presents a new cell manipulation method using a moving dielectrophoretic force to transport or fractionate cells along a microfluidic channel. The proposed moving dielectrophoresis (mDEP) is generated by sequentially energizing a single electrode or an array of electrodes to form an electric field that moves cells continuously along the microchannel. Cell fractionation is controlled by the applied electrical frequency, and cell transportation is controlled by the interelectrode activation time. The applicability of this method was demonstrated to simultaneously fractionate and transport Saccharomyces cerevisiae yeast cells, both viable and nonviable, by operating at conditions under which the cells were subjected to positive and negative dielectrophoresis, respectively. Compared to the conventional dielectrophoresis (cDEP and traveling wave dielectrophoresis (twDEP), moving dielectrophoresis allows cells to be separated on the basis of the real part of the Clausius-Mossotti factor, as in cDEP, but yet allows the direct transportation of separated cells without using fluid flow, as in twDEP. This dielectrophoresis technique provides a new way to manipulate cells and can be readily implemented on programmable multielectrode devices.     

Intuitive, image-based cell sorting using optofluidic cell sorting

We present a microfluidic cell-sorting device which augments microscopy with the capability to perform facile image-based cell sorting. This combination enables intuitive, complex phenotype sorting based on spatio-temporal fluorescence or cell morphology. The microfluidic device contains a microwell array that can be passively loaded with mammalian cells via sedimentation and can be subsequently inspected with microscopy. After inspection, we use the scattering force from a focused infrared laser to levitate cells of interest from their wells into a flow field for collection. First, we demonstrate image-based sorting predicated on whole-cell fluorescence, which could enable sorting based on temporal whole-cell fluorescence behavior. Second, we demonstrate image-based sorting predicated on fluorescence localization (nuclear vs whole-cell fluorescence), highlighting the capability of our approach to sort based on imaged subcellular events, such as localized protein expression or translocation events. We achieve postsort purities up to 89% and up to 155-fold enrichment of target cells. Optical manipulation literature and a direct cell viability assay suggest that cells remain viable after using our technique. The architecture is highly scalable and supports over 10 000 individually addressable trap sites. Our approach enables sorting of significant populations based on subcellular spatio-temporal information, which is difficult or impossible with existing widespread sorting technologies.     

纳米结构在微纳传感器应用中的介电泳操控研究

要将纳米结构作为功能元件应用在微纳传感器等电学系统中，首先要解决的问题是材料的定位操作问题。介电泳技术是一种很有潜力的纳米操控技术。在自行设计的几种电极的基础上，采用介电泳技术对微纳米材料进行操控，可以实现微纳米材料沿电场方向的排布并口电极间的跨接。初步测试了采用介电泳技术操控自行生长的纳米结构制作而成的湿度传感器的基本特性，结果表明介电泳技术可以很好地实现纳米材料在传感器领域中的应用。     

Separation of latex spheres using dielectrophoresis and fluid flow

The authors present a method for separation of two latex spheres populations using dielectrophoresis (DEP) and the fluid drag force. Microelectrodes of a suitable layout are used to trap one population of spheres, while the other one is dragged away from the electrodes by the generated fluid flow. The finite difference method is implemented in C++ to calculate the potential distribution by solving Laplace's equation. From the potential distribution, the DEP force on particles is calculated. The drag force on particles due to the liquid motion is calculated from the observed fluid velocity. The experimental results are shown to be in good agreement with the numerical solution.     

Manipulation of bioparticles using traveling wave dielectrophoresis: numerical approach

A typical microelectrode structure, interdigitated bar electrodes, has been developed for the selective manipulation and separation of bioparticles using traveling field dielectrophoresis effects under AC electrokinetics. This paper presents meshless numerical solutions of traveling wave dielectrophoresis for such interdigitated electrode array energized with 4-phase signal. This meshless method is based on a truncated 2D Taylor series expansion with the first three orders together with a weighted least-square approximation procedure. Dielectrophoretic forces and traveling wave forces are calculated and their applications for bioparticle manipulation and separation are discussed.     

Trapping heavy metals by using calcium hydroxyapatite and dielectrophoresis

We propose a novel technique for the removal of heavy metal waste from contaminated water. Our method consists in using dielectrophoresis (DEP) to trap hydroxyapatite (HAP) particles of 1 microm size in water after they have adsorbed heavy metal (Pb, Zn, Cu, Co and Cr). Although HAP can adsorb heavy metals in water and as such offers great promise as a waste-cleaning tool , one of the current challenges is the efficient removal of the HAP particles once they have adsorbed the heavy metals. We show in this paper that DEP can be used to concentrate such particles in certain regions, thus rendering the rest of the solution volume nearly free of contaminated particles. We present here both experimental and numerical results for suspensions at low concentrations.     

Interfacing cell-based assays in environmental scanning electron microscopy using dielectrophoresis

Development of the dielectrophoretic (DEP) live cell trapping technology and its interfacing with the environmental scanning electron microscopy (ESEM) is described. DEP microelectrode arrays were fabricated on glass substrate using photolithography and lift-off. Chip-based arrays were applied for ESEM analysis of DEP-trapped human leukemic cells. This work provides proof-of-concept interfacing of the DEP cell retention and trapping technology with ESEM to provide a high-resolution analysis of individual nonadherent cells.     

Active control of silver nanoparticles spacing using dielectrophoresis for surface-enhanced Raman scattering

We demonstrate an active microfluidic platform that integrates dielectrophoresis for the control of silver nanoparticles spacing, as they flow in a liquid channel. By careful control of the nanoparticles spacing, we can effectively increase the surface-enhanced Raman scattering (SERS) signal intensity based on augmenting the number of SERS-active hot-spots, while avoiding irreversible aggregation of the particles. The system is benchmarked using dipicolinate (2,6-pyridinedicarboxylic acid) (DPA), which is a biomarker of Bacillus anthracis. The validity of the results is discussed using several complementing characterization scenarios.     

Bitonic sorting using an optoelectronic recirculating architecture

An optoelectronic bitonic sorter based on a recirculating architecture is presented. The data are input inword parallel-bit parallel fashion and processed by two smart pixel arrays made up of bitwise compare-and-exchange modules. Along with the logic design, the control and synchronization of the bitwise compare-and-exchange modules are discussed. Finally, the capacity, hardware requirements, response time, and throughput of the recirculating bitonic sorter are compared with a pipeline implementation. The proposed recirculating architecture is shown to require less hardware than the pipelined systems. However, the decrease in hardware results in a decrease in system throughput.     

Manipulation of microparticles using Bessel beams from semiconductor lasers

Optical manipulation of microscopic objects (including living cells) using Bessel beams from semiconductor lasers has been demonstrated for the first time. In addition, it has been found in the experiments that a Bessel beam of sufficient power from a semiconductor laser makes it possible to manipulate simultaneously several microscopic objects captured into its central lobe and the first ring.     

Size sorting of protein assemblies using polymeric gradient surfaces

We report on a novel approach for the size-dependent fractionation of protein assemblies on polymeric surfaces. Using a simple temperature gradient method to generate one-dimensional gradients of grafted poly(ethylene glycol), we fabricated silicon-oxide chips with a gradually changing surface density of kinesin motor molecules. We demonstrate that such a bioactive surface can be used to sort gliding microtubules according to their length. To our knowledge, this is the first example of the self-organized sorting of protein assemblies on surfaces.     

Preparation of cross-linked sodium alginate microparticles using glutaraldehyde in methanol

Polymeric sodium alginate microparticles were prepared by precipitating sodium alginate in methanol, followed by cross-linking with glutaraldehyde. The extent of cross-linking was controlled by the time of exposure to glutaraldehyde. The topology of microparticles was characterized by scanning electron microscopy (SEM), which indicated smooth surfaces. The equilibrium swelling experiments were carried out in water to observe the effect of cross-linking and drug loading for better utility of microparticles. It was found that swelling decreased, but drug loading increased, with an increase in cross-linking of the matrix.     

Preparation and characterization of spironolactone-loaded gelucire microparticles using spray-drying technique

The basic objectives of this study were to prepare and characterize solid dispersions of poorly soluble drug spironolactone (SP) using gelucire carriers by spray-drying technique. The properties of the microparticles produced were studied by differential scanning calorimetry (DSC), scanning electron microscopy, saturation solubility, encapsulation efficiency, and dissolution studies. The absence of SP peaks in DSC profiles of microparticles suggests the transformation of crystalline SP into an amorphous form. The in vitro dissolution test showed a significant increase in the dissolution rate of microparticles as compared with pure SP and physical mixtures (PMs) of drug with gelucire carriers. Therefore, the dissolution rate of poorly water-soluble drug SP can be significantly enhanced by the preparation of solid dispersion using spray-drying technique.