Magnetic microbead transport during resistive pulse sensing

Tunable resistive pulse sensing (TRPS) experiments have been used to quantitatively study the motion of 1 μm superparamagnetic beads in a variable magnetic field. Closed-form theory has been developed to interpret the experiments, incorporating six particle transport mechanisms which depend on particle position in and near a conical pore. For our experiments, calculations indicate that pressure-driven flow dominates electrophoresis and magnetism by a factor of ∼100 in the narrowest part of the pore, but that magnetic force should dominate further than ∼1 mm from the membrane. As expected, the observed resistive pulse rate falls as the magnet is moved closer to the pore, while the increase in pulse duration suggests that trajectories in the half space adjacent to the pore opening are important. Aggregation was not observed, consistent with the high hydrodynamic shear near the pore constriction and the high magnetization of aggregates. The theoretical approach is also used to calculate the relative importance of transport mechanisms over a range of geometries and experimental conditions extending well beyond our own experiments. TRPS is emerging as a versatile form of resistive pulse sensing, while magnetic beads are widely used in biotechnology and sensing applications.     

Co-ordinated detection of microparticles using tunable resistive pulse sensing and fluorescence spectroscopy

Tunable resistive pulse sensing (TRPS) has emerged as a useful tool for particle-by-particle detection and analysis of microparticles and nanoparticles as they pass through a pore in a thin stretchable membrane. We have adapted a TRPS device in order to conduct simultaneous optical measurements of particles passing through the pore. High-resolution fluorescence emission spectra have been recorded for individual 1.9 μm diameter particles at a sampling period of 4.3 ms. These spectra are time-correlated with RPS pulses in a current trace sampled every 20 μs. The flow rate through the pore, controlled by altering the hydrostatic pressure, determines the rate of particle detection. At pressures below 1 kPa, more than 90% of fluorescence and RPS events were matching. At higher pressures, some peaks were missed by the fluorescence technique due to the difference in sampling rates. This technique enhances the particle-by-particle specificity of conventional RPS measurements and could be useful for a range of particle characterization and bioanalysis applications.     

Field-free particle focusing in microfluidic plugs

Particle concentration is a key unit operation in biochemical assays. Although there are many techniques for particle concentration in continuous-phase microfluidics, relatively few are available in multiphase (plug-based) microfluidics. Existing approaches generally require external electric or magnetic fields together with charged or magnetized particles. This paper reports a passive technique for particle concentration in water-in-oil plugs which relies on the interaction between particle sedimentation and the recirculating vortices inherent to plug flow in a cylindrical capillary. This interaction can be quantified using the Shields parameter (θ), a dimensionless ratio of a particle’s drag force to its gravitational force, which scales with plug velocity. Three regimes of particle behavior are identified. When θ is less than the movement threshold (region I), particles sediment to the bottom of the plug where the internal vortices subsequently concentrate the particles at the rear of the plug. We demonstrate highly efficient concentration (∼100%) of 38 μm glass beads in 500 μm diameter plugs traveling at velocities up to 5 mm/s. As θ is increased beyond the movement threshold (region II), particles are suspended in well-defined circulation zones which begin at the rear of the plug. The length of the zone scales linearly with plug velocity, and at sufficiently large θ, it spans the length of the plug (region III). A second effect, attributed to the co-rotating vortices at the rear cap, causes particle aggregation in the cap, regardless of flow velocity. Region I is useful for concentrating/collecting particles, while the latter two are useful for mixing the beads with the solution. Therefore, the two key steps of a bead-based assay, concentration and resuspension, can be achieved simply by changing the plug velocity. By exploiting an interaction of sedimentation and recirculation unique to multiphase flow, this simple technique achieves particle concentration without on-chip components, and could therefore be applied to a range of heterogeneous screening assays in discrete nl plugs.     

Development of a magnetic immunosorbent for on-chip preconcentration of amyloid �� isoforms: Representatives of Alzheimer��s disease biomarkers

Determination of amyloid β (Aβ) isoforms and in particular the proportion of the Aβ 1-42 isoform in cerebrospinal fluid (CSF) of patients suspected of Alzheimer’s disease might help in early diagnosis and treatment of that illness. Due to the low concentration of Aβ peptides in biological fluids, a preconcentration step prior to the detection step is often necessary. This study utilized on-chip immunoprecipitation, known as micro-immunoprecipitation (μIP). The technique uses an immunosorbent (IS) consisting of magnetic beads coated with specific anti-Aβ antibodies organized into an affinity microcolumn by a magnetic field. Our goal was to thoroughly describe the critical steps in developing the IS, such as selecting the proper beads and anti-Aβ antibodies, as well as optimizing the immobilization technique and μIP protocol. The latter includes selecting optimal elution conditions. Furthermore, we demonstrate the efficiency of anti-Aβ IS for μIP and specific capture of 5 Aβ peptides under optimized conditions using various subsequent analytical methods, including matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), capillary electrophoresis, microchip electrophoresis, and immunoblotting. Synthetic Aβ peptides samples prepared in buffer and spiked in human CSF were analyzed. Finally, on-chip immunoprecipitation of Aβ peptides in human CSF sample was performed.     

Transport and separation of micron sized particles at isotachophoretic transition zones

Conventionally, isotachophoresis (ITP) is used for separation of ionic samples according to their electrophoretic mobilities. We demonstrate that the scope of ITP applications may be extended toward particle concentration and separation. Owing to the distributions of electrolyte concentration and electric field inside a transition zone between two electrolytes, a number of different forces act on a small particle. As far as possible, we provide estimates for the order of magnitude of these forces and analyze their scaling with the particle size and the electric-field strength. Furthermore, we experimentally demonstrate that polymer beads of 5 μm diameter dispersed in a high mobility “leading” electrolyte are picked up and carried along by an ITP transition zone which is formed with a low mobility “trailing” electrolyte. By studying the particle positions and trajectories, we show that impurities in the electrolytes play a significant role in the experiments. Additionally, it is experimentally shown that different types of beads can be separated at an ITP transition zone. In particular, beads of 1 μm diameter are not carried along with the transition zone, in contrast to the 5 μm beads. The presented technique thus adds to the portfolio of electrokinetic transport, concentration, and separation methods in microfluidics.     

Curvature-induced dielectrophoresis for continuous separation of particles by charge in spiral microchannels

The separation of particles from a heterogeneous mixture is critical in chemical and biological analyses. Many methods have been developed to separate particles in microfluidic devices. However, the majority of these separations have been limited to be size based and binary. We demonstrate herein a continuous dc electric field driven separation of carboxyl-coated and noncoated 10 μm polystyrene beads by charge in a double-spiral microchannel. This method exploits the inherent electric field gradients formed within the channel turns to manipulate particles by dielectrophoresis and is thus termed curvature-induced dielectrophoresis. The spiral microchannel is also demonstrated to continuously sort noncoated 5 μm beads, noncoated 10 μm beads, and carboxyl-coated 10 μm beads into different collecting wells by charge and size simultaneously. The observed particle separation processes in different situations are all predicted with reasonable agreements by a numerical model. This curvature-induced dielectrophoresis technique eliminates the in-channel microelectrodes and obstacles that are required in traditional electrode- and insulator-based dielectrophoresis devices. It may potentially be used to separate multiple particle targets by intrinsic properties for lab-on-a-chip applications.     

Geometrical optimization of microstripe arrays for microbead magnetophoresis

Manipulation of magnetic beads plays an increasingly important role in molecular diagnostics. Magnetophoresis is a promising technique for selective transportation of magnetic beads in lab-on-a-chip systems. We investigate periodic arrays of exchange-biased permalloy microstripes fabricated using a single lithography step. Magnetic beads can be continuously moved across such arrays by combining the spatially periodic magnetic field from microstripes with a rotating external magnetic field. By measuring and modeling the magnetophoresis properties of thirteen different stripe designs, we study the effect of the stripe geometry on the magnetophoretic transport properties of the magnetic microbeads between the stripes. We show that a symmetric geometry with equal width of and spacing between the microstripes facilitates faster transportation and that the optimal period of the periodic stripe array is approximately three times the height of the bead center over the microstripes.     

Density-dependent separation of encapsulated cells in a microfluidic channel by using a standing surface acoustic wave

This study presents a method for density-based separation of monodisperse encapsulated cells using a standing surface acoustic wave (SSAW) in a microchannel. Even though monodisperse polymer beads can be generated by the state-of-the-art technology in microfluidics, the quantity of encapsulated cells cannot be controlled precisely. In the present study, mono-disperse alginate beads in a laminar flow can be separated based on their density using acoustophoresis. A mixture of beads of equal sizes but dissimilar densities was hydrodynamically focused at the entrance and then actively driven toward the sidewalls by a SSAW. The lateral displacement of a bead is proportional to the density of the bead, i.e., the number of encapsulated cells in an alginate bead. Under optimized conditions, the recovery rate of a target bead group (large-cell-quantity alginate beads) reached up to 97% at a rate of 2300 beads per minute. A cell viability test also confirmed that the encapsulated cells were hardly damaged by the acoustic force. Moreover, cell-encapsulating beads that were cultured for 1 day were separated in a similar manner. In conclusion, this study demonstrated that a SSAW can successfully separate monodisperse particles by their density. With the present technique for separating cell-encapsulating beads, the current cell engineering technology can be significantly advanced.     

Droplet-based microfluidic washing module for magnetic particle-based assays

In this paper, we propose a continuous flow droplet-based microfluidic platform for magnetic particle-based assays by employing in-droplet washing. The droplet-based washing was implemented by traversing functionalized magnetic particles across a laterally merged droplet from one side (containing sample and reagent) to the other (containing buffer) by an external magnetic field. Consequently, the magnetic particles were extracted to a parallel-synchronized train of washing buffer droplets, and unbound reagents were left in an original train of sample droplets. To realize the droplet-based washing function, the following four procedures were sequentially carried in a droplet-based microfluidic device: parallel synchronization of two trains of droplets by using a ladder-like channel network; lateral electrocoalescence by an electric field; magnetic particle manipulation by a magnetic field; and asymmetrical splitting of merged droplets. For the stable droplet synchronization and electrocoalescence, we optimized droplet generation conditions by varying the flow rate ratio (or droplet size). Image analysis was carried out to determine the fluorescent intensity of reagents before and after the washing step. As a result, the unbound reagents in sample droplets were significantly removed by more than a factor of 25 in the single washing step, while the magnetic particles were successfully extracted into washing buffer droplets. As a proof-of-principle, we demonstrate a magnetic particle-based immunoassay with streptavidin-coated magnetic particles and fluorescently labelled biotin in the proposed continuous flow droplet-based microfluidic platform.     

An automated microfluidic system for single-stranded DNA preparation and magnetic bead-based microarray analysis

We present an integrated microfluidic device capable of performing single-stranded DNA (ssDNA) preparation and magnetic bead-based microarray analysis with a white-light detection for detecting mutations that account for hereditary hearing loss. The entire operation process, which includes loading of streptavidin-coated magnetic beads (MBs) and biotin-labeled polymerase chain reaction products, active dispersion of the MBs with DNA for binding, alkaline denaturation of DNA, dynamic hybridization of the bead-labeled ssDNA to a tag array, and white-light detection, can all be automatically accomplished in a single chamber of the microchip, which was operated on a self-contained instrument with all the necessary components for thermal control, fluidic control, and detection. Two novel mixing valves with embedded polydimethylsiloxane membranes, which can alternately generate a 3-μl pulse flow at a peak rate of around 160 mm/s, were integrated into the chip for thoroughly dispersing magnetic beads in 2 min. The binding efficiency of biotinylated oligonucleotides to beads was measured to be 80.6% of that obtained in a tube with the conventional method. To critically test the performance of this automated microsystem, we employed a commercial microarray-based detection kit for detecting nine mutation loci that account for hereditary hearing loss. The limit of detection of the microsystem was determined as 2.5 ng of input K562 standard genomic DNA using this kit. In addition, four blood samples obtained from persons with mutations were all correctly typed by our system in less than 45 min per run. The fully automated, “amplicon-in-answer-out” operation, together with the white-light detection, makes our system an excellent platform for low-cost, rapid genotyping in clinical diagnosis.     

Three Dimensional Electro-magnetic Field Diffusion in Multilayer Cylindrical Structures

The problem of the magnetic field diffusion through cylindrical structures is considered. An analytic solution is given for the problem of diffusion, through any number of resistive coaxial layers, of the three components of a quite general input vector magnetic field assigned on a boundary cylindrical surface. The limitations on the class of input functions (or more generally distributions) are the existence of the double space Fourier transform and the time Laplace transform; this makes the solution suitable for studying the transient-state behaviour in cylindrical structures of finite or infinite length.     

微波和脉冲磁场复合作用下鲜切哈密瓜杀菌实验研究

本文进行了微波和脉冲磁场复合作用下鲜切哈密瓜的杀菌实验研究,考察了微波预处理,磁场强度,脉冲次数及初始茵落总数对杀菌效果的影响。研究结果表明：微波和脉冲磁场的复合处理可以有效杀死鲜切哈密瓜中的细菌,茵落总数的最大杀菌率可以达到99．9％;12s的微波预处理可以起到物料升温与一定的杀菌作用,为脉冲磁场下鲜切哈密瓜的杀菌创造了有利条件;当磁场强度为0．6T时,随着脉冲作用次数的增加,菌落总数先下降,后有所增加,其转折点为20;当脉冲数为10时,随着磁场强度的增加,茵落总数总体上呈下降的趋势;初始菌落总数的增加有利于微波杀菌效果的提高,但对脉冲磁场的杀菌效果影响较小。     

Construction and operation of a microrobot based on magnetotactic bacteria in a microfluidic chip

Magnetotactic bacteria (MTB) are capable of swimming along magnetic field lines. This unique feature renders them suitable in the development of magnetic-guided, auto-propelled microrobots to serve in target molecule separation and detection, drug delivery, or target cell screening in a microfluidic chip. The biotechnology to couple these bacteria with functional loads to form microrobots is the critical point in its application. Although an immunoreaction approach to attach functional loads to intact MTB was suggested, details on its realization were hardly mentioned. In the current paper, MTB-microrobots were constructed by attaching 2 μm diameter microbeads to marine magnetotactic ovoid MO-1 cells through immunoreactions. These microrobots were controlled using a special control and tracking system. Experimental results prove that the attachment efficiency can be improved to ∼30% via an immunoreaction. The motility of the bacteria attached with different number of loads was also assessed. The results show that MTB can transport one load at a velocity of ∼21 μm/s and still move and survive for over 30 min. The control and tracking system is fully capable of directing and monitoring the movement of the MTB-microrobots. The rotating magnetic fields can stop the microrobots by trapping them as they swim within a circular field with a controllable size. The system has potential use in chemical analyses and medical diagnoses using biochips as well as in nano/microscale transport.     

Negative dielectrophoretic capture of bacterial spores in food matrices

A microfluidic device with planar square electrodes is developed for capturing particles from high conductivity media using negative dielectrophoresis (n-DEP). Specifically, Bacillus subtilis and Clostridium sporogenes spores, and polystyrene particles are tested in NaCl solution (0.05 and 0.225 S∕m), apple juice (0.225 S∕m), and milk (0.525 S∕m). Depending on the conductivity of the medium, the Joule heating produces electrothermal flow (ETF), which continuously circulates and transports the particles to the DEP capture sites. Combination of the ETF and n-DEP results in different particle capture efficiencies as a function of the conductivity. Utilizing 20 μm height DEP chambers, “almost complete” and rapid particle capture from lower conductivity (0.05 S∕m) medium is observed. Using DEP chambers above 150 μm in height, the onset of a global fluid motion for high conductivity media is observed. This motion enhances particle capture on the electrodes at the center of the DEP chamber. The n-DEP electrodes are designed to have well defined electric field minima, enabling sample concentration at 1000 distinct locations within the chip. The electrode design also facilitates integration of immunoassay and other surface sensors onto the particle capture sites for rapid detection of target micro-organisms in the future.     

Dielectrophoretic manipulation of particles in a modified microfluidic H filter with multi-insulating blocks

The conventional microfluidic H filter is modified with multi-insulating blocks to achieve a flow-through manipulation and separation of microparticles. The device transports particles by exploiting electro-osmosis and electrophoresis, and manipulates particles by utilizing dielectrophoresis (DEP). Polydimethylsiloxane (PDMS) blocks fabricated in the main channel of the PDMS H filter induce a nonuniform electric field, which exerts a negative DEP force on the particles. The use of multi-insulating blocks not only enhances the DEP force generated, but it also increases the controllability of the motion of the particles, facilitating their manipulation and separation. Experiments were conducted to demonstrate the controlled flow direction of particles by adjusting the applied voltages and the separation of particles by size under two different input conditions, namely (i) a dc electric field mode and (ii) a combined ac and dc field mode. Numerical simulations elucidate the electrokinetic and hydrodynamic forces acting on a particle, with theoretically predicted particle trajectories in good agreement with those observed experimentally. In addition, the flow field was obtained experimentally with fluorescent tracer particles using the microparticle image velocimetry (μ-PIV) technique.     

Inducing self-rotation of cells with natural and artificial melanin in a linearly polarized alternating current electric field

The phenomenon of self-rotation observed in naturally and artificially pigmented cells under an applied linearly polarized alternating current (non-rotating) electrical field has been investigated. The repeatable and controllable rotation speeds of the cells were quantified and their dependence on dielectrophoretic parameters such as frequency, voltage, and waveform was studied. Moreover, the rotation behavior of the pigmented cells with different melanin content was compared to quantify the correlation between self-rotation and the presence of melanin. Most importantly, macrophages, which did not originally rotate in the applied non-rotating electric field, began to exhibit self-rotation that was very similar to that of the pigmented cells, after ingesting foreign particles (e.g., synthetic melanin or latex beads). We envision the discovery presented in this paper will enable the development of a rapid, non-intrusive, and automated process to obtain the electrical conductivities and permittivities of cellular membrane and cytoplasm in the near future.     

Rapid magnetic heating treatment by highly charged maghemite nanoparticles on Wistar rats exocranial glioma tumors at microliter volume

One of the most significant challenges implementing colloidal magnetic nanoparticles in medicine is the efficient heating of microliter quantities by applying a low frequency alternating magnetic field. The ultimate goal is to accomplish nonsurgically the treatment of millimeter size tumors. Here, we demonstrate the synthesis, characterization, and the in vitro as well as in vivo efficiency of a dextran coated maghemite (γ-Fe₂O₃) ferrofluid with an exceptional response to magnetic heating. The difference to previous synthetic attempts is the high charge of the dextran coating, which according to our study maintains the colloidal stability and good dispersion of the ferrofluid during the magnetic heating stage. Specifically, in vitro 2 μl of the ferrofluid gives an outstanding temperature rise of 33 °C within 10 min, while in vivo treatment, by infusing 150 μl of the ferrofluid in animal model (rat) glioma tumors, causes an impressive cancer tissue dissolution.     

Magnetophoretic manipulation in microsystem using carbonyl iron-polydimethylsiloxane microstructures

This paper reports the use of a recent composite material, noted hereafter i-PDMS, made of carbonyl iron microparticles mixed in a PolyDiMethylSiloxane (PDMS) matrix, for magnetophoretic functions such as capture and separation of magnetic species. We demonstrated that this composite which combine the advantages of both components, can locally generate high gradients of magnetic field when placed between two permanent magnets. After evaluating the magnetic susceptibility of the material as a function of the doping ratio, we investigated the molding resolution offered by i-PDMS to obtain microstructures of various sizes and shapes. Then, we implemented 500 μm i-PDMS microstructures in a microfluidic channel and studied the influence of flow rate on the deviation and trapping of superparamagnetic beads flowing at the neighborhood of the composite material. We characterized the attraction of the magnetic composite by measuring the distance from the i-PDMS microstructure, at which the beads are either deviated or captured. Finally, we demonstrated the interest of i-PDMS to perform magnetophoretic functions in microsystems for biological applications by performing capture of magnetically labeled cells.     

An integrated actuating and sensing system for light-addressable potentiometric sensor (LAPS) and light-actuated AC electroosmosis (LACE) operation

To develop a lab on a chip (LOC) integrated with both sensor and actuator functions, a novel two-in-one system based on optical-driven manipulation and sensing in a microfluidics setup based on a hydrogenated amorphous silicon (a-Si:H) layer on an indium tin oxide/glass is first realized. A high-intensity discharge xenon lamp functioned as the light source, a chopper functioned as the modulated illumination for a certain frequency, and a self-designed optical path projected on the digital micromirror device controlled by the digital light processing module was established as the illumination input signal with the ability of dynamic movement of projected patterns. For light-addressable potentiometric sensor (LAPS) operation, alternating current (AC)-modulated illumination with a frequency of 800 Hz can be generated by the rotation speed of the chopper for photocurrent vs bias voltage characterization. The pH sensitivity, drift coefficient, and hysteresis width of the Si₃N₄ LAPS are 52.8 mV/pH, −3.2 mV/h, and 10.5 mV, respectively, which are comparable to the results from the conventional setup. With an identical two-in-one system, direct current illumination without chopper rotation and an AC bias voltage can be provided to an a-Si:H chip with a manipulation speed of 20 μm/s for magnetic beads with a diameter of 1 μm. The collection of magnetic beads by this light-actuated AC electroosmosis (LACE) operation at a frequency of 10 kHz can be easily realized. A fully customized design of an illumination path with less decay can be suggested to obtain a high efficiency of manipulation and a high signal-to-noise ratio of sensing. With this proposed setup, a potential LOC system based on LACE and LAPS is verified with the integration of a sensor and an actuator in a microfluidics setup for future point-of-care testing applications.