Quantitative Magnetic Separation of Particles and Cells Using Gradient Magnetic Ratcheting

Extraction of rare target cells from biosamples is enabling for life science research. Traditional rare cell separation techniques, such as magnetic activated cell sorting, are robust but perform coarse, qualitative separations based on surface antigen expression. A quantitative magnetic separation technology is reported using high‐force magnetic ratcheting over arrays of magnetically soft micropillars with gradient spacing, and the system is used to separate and concentrate magnetic beads based on iron oxide content (IOC) and cells based on surface expression. The system consists of a microchip of permalloy micropillar arrays with increasing lateral pitch and a mechatronic device to generate a cycling magnetic field. Particles with higher IOC separate and equilibrate along the miropillar array at larger pitches. A semi‐analytical model is developed that predicts behavior for particles and cells. Using the system, LNCaP cells are separated based on the bound quantity of 1 μm anti‐epithelial cell adhesion molecule (EpCAM) particles as a metric for expression. The ratcheting cytometry system is able to resolve a ±13 bound particle differential, successfully distinguishing LNCaP from PC3 populations based on EpCAM expression, correlating with flow cytometry analysis. As a proof‐of‐concept, EpCAM‐labeled cells from patient blood are isolated with 74% purity, demonstrating potential toward a quantitative magnetic separation instrument.     

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.     

High‐Density Microfluidic Particle‐Cluster‐Array Device for Parallel and Dynamic Study of Interaction between Engineered Particles

A high‐density and high‐performance microfluidic particle‐cluster‐array device utilizing a novel hydrodynamically tunable pneumatic valve (HTPV) is reported for parallel and dynamic monitoring of the interactions taking place in particle clusters. The key concept involves passive operation of the HTPV through elastic deformation of a thin membrane using only the hydrodynamic force inherent in microchannel flows. This unique feature allows the discrete and high‐density (≈30 HTPVs mm^?2) arrangement of numerous HTPVs in a microfluidic channel without any pneumatic connection. In addition, the HTPV achieves high‐performance clustering (≈92%) of three different particles in an array format through the optimization of key design and operating parameters. Finally, a contamination‐free, parallel, and dynamic biochemical analysis strategy is proposed, which employs a simple one‐inlet–one‐outlet device operated by the effective combination of several techniques, including particle clustering, the interactions between engineered particles, two‐phase partitioning and dehydration control of aqueous plugs, and shape/color‐based particle identification.     

Continuous Flow Deformability‐Based Separation of Circulating Tumor Cells Using Microfluidic Ratchets

Circulating tumor cells (CTCs) offer tremendous potential for the detection and characterization of cancer. A key challenge for their isolation and subsequent analysis is the extreme rarity of these cells in circulation. Here, a novel label‐free method is described to enrich viable CTCs directly from whole blood based on their distinct deformability relative to hematological cells. This mechanism leverages the deformation of single cells through tapered micrometer scale constrictions using oscillatory flow in order to generate a ratcheting effect that produces distinct flow paths for CTCs, leukocytes, and erythrocytes. A label‐free separation of circulating tumor cells from whole blood is demonstrated, where target cells can be separated from background cells based on deformability despite their nearly identical size. In doping experiments, this microfluidic device is able to capture >90% of cancer cells from unprocessed whole blood to achieve 10⁴‐fold enrichment of target cells relative to leukocytes. In patients with metastatic castration‐resistant prostate cancer, where CTCs are not significantly larger than leukocytes, CTCs can be captured based on deformability at 25× greater yield than with the conventional CellSearch system. Finally, the CTCs separated using this approach are collected in suspension and are available for downstream molecular characterization.     

Magnetophoretic continuous purification of single-walled carbon nanotubes from catalytic impurities in a microfluidic device

A magnetophoretic continuous purification method is presented of single-walled carbon nanotubes (SWCNTs) from the superparamagnetic iron-catalyst impurities in a microfluidic device without any influence on inherent SWCNT properties. By employing microfluidics and a magnetic-field-induced saw-tooth nickel microstructure, a highly enhanced magnetic force in adjoining microchannels is exploited. The iron impurities of SWCNTs are attracted towards areas of higher magnetic-flux density in the microchannels where magnetic field was asymmetrically generated perpendicularly to the streamline. We obtained highly purified SWCNTs at a rate of 0.36 mg h(-1) and that are estimated to be about 99% purity.     

Deterministic Migration‐Based Separation of White Blood Cells

Functional and phenotypic analyses of peripheral white blood cells provide useful clinical information. However, separation of white blood cells from peripheral blood requires a time‐consuming, inconvenient process and thus analyses of separated white blood cells are limited in clinical settings. To overcome this limitation, a microfluidic separation platform is developed to enable deterministic migration of white blood cells, directing the cells into designated positions according to a ridge pattern. The platform uses slant ridge structures on the channel top to induce the deterministic migration, which allows efficient and high‐throughput separation of white blood cells from unprocessed whole blood. The extent of the deterministic migration under various rheological conditions is explored, enabling highly efficient migration of white blood cells in whole blood and achieving high‐throughput separation of the cells (processing 1 mL of whole blood less than 7 min). In the separated cell population, the composition of lymphocyte subpopulations is well preserved, and T cells secrete cytokines without any functional impairment. On the basis of the results, this microfluidic platform is a promising tool for the rapid enrichment of white blood cells, and it is useful for functional and phenotypic analyses of peripheral white blood cells.     

Microfluidic characterization and continuous separation of cells and particles using conducting poly(dimethyl siloxane) electrode induced alternating current-dielectrophoresis

This paper presents a poly(dimethyl siloxane) (PDMS) polymer microfluidic device using alternating current (ac) dielectrophoresis (DEP) for separating live cells from interfering particles of similar sizes by their polarizabilities under continuous flow and for characterizing DEP behaviors of cells in stagnant flow. The ac-DEP force is generated by three-dimensional (3D) conducting PDMS composite electrodes fabricated on a sidewall of the device main channel. Such 3D PDMS composite electrodes are made by dispersing microsized silver (Ag) fillers into PDMS gel. The sidewall AgPDMS electrodes can generate a 3D electric field that uniformly distributes throughout the channel height and varies along the channel lateral direction, thereby producing stronger lateral DEP effects over the entire channel. This allows not only easy observation of cell/particle lateral motion but also using the lateral DEP force for manipulation of cells/particles. The former feature is used to characterize the frequency-dependent DEP behaviors of Saccharomyces cerevisiae (yeast) and Escherichia coli (bacteria). The latter is utilized for continuous separation of live yeast and bacterial cells from similar-size latex particles as well as live yeast cells from dead yeast cells. The separation efficiency of 97% is achieved in all cases. The demonstration of these functions shows promising applications of the microfluidic device.     

Anticipating Cutoff Diameters in Deterministic Lateral Displacement (DLD) Microfluidic Devices for an Optimized Particle Separation

Deterministic lateral displacement (DLD) devices enable to separate nanometer to micrometer‐sized particles around a cutoff diameter, during their transport through a microfluidic channel with slanted rows of pillars. In order to design appropriate DLD geometries for specific separation sizes, robust models are required to anticipate the value of the cutoff diameter. So far, the proposed models result in a single cutoff diameter for a given DLD geometry. This paper shows that the cutoff diameter actually varies along the DLD channel, especially in narrow pillar arrays. Experimental and numerical results reveal that the variation of the cutoff diameter is induced by boundary effects at the channel side walls, called the wall effect. The wall effect generates unexpected particle trajectories that may compromise the separation efficiency. In order to anticipate the wall effect when designing DLD devices, a predictive model is proposed in this work and has been validated experimentally. In addition to the usual geometrical parameters, a new parameter, the number of pillars in the channel cross dimension, is considered in this model to investigate its influence on the particle trajectories.     

Microfluidic‐Based Approaches in Targeted Cell/Particle Separation Based on Physical Properties: Fundamentals and Applications

Cell separation is a key step in many biomedical research areas including biotechnology, cancer research, regenerative medicine, and drug discovery. While conventional cell sorting approaches have led to high‐efficiency sorting by exploiting the cell's specific properties, microfluidics has shown great promise in cell separation by exploiting different physical principles and using different properties of the cells. In particular, label‐free cell separation techniques are highly recommended to minimize cell damage and avoid costly and labor‐intensive steps of labeling molecular signatures of cells. In general, microfluidic‐based cell sorting approaches can separate cells using “intrinsic” (e.g., fluid dynamic forces) versus “extrinsic” external forces (e.g., magnetic, electric field, etc.) and by using different properties of cells including size, density, deformability, shape, as well as electrical, magnetic, and compressibility/acoustic properties to select target cells from a heterogeneous cell population. In this work, principles and applications of the most commonly used label‐free microfluidic‐based cell separation methods are described. In particular, applications of microfluidic methods for the separation of circulating tumor cells, blood cells, immune cells, stem cells, and other biological cells are summarized. Computational approaches complementing such microfluidic methods are also explained. Finally, challenges and perspectives to further develop microfluidic‐based cell separation methods are discussed.     

High‐Speed Colloidal Quantum Dot Photodiodes via Accelerating Charge Separation at Metal–Oxide Interface

With ever‐growing technological demands in the imaging sensor industry for autonomous driving and augmented reality, developing sensors that can satisfy not only image resolution but also the response speed becomes more challenging. Herein, the focus is on developing a high‐speed photosensor capable of obtaining high‐resolution, high‐speed imaging with colloidal quantum dots (QDs) as the photosensitive material. In detail, high‐speed QD photodiodes are demonstrated with rising and falling times of τ_r = 28.8 ± 8.34 ns and τ_f = 40 ± 9.81 ns, respectively, realized by fast separation of electron–hole pairs due to the action of internal electric field at the QD interface, mainly by the interaction between metal oxide and the QD's ligands. Such energy transfer relations are analyzed and interpreted with time‐resolved photoluminescence measurements, providing physical understanding of the device and working principles.     

Perfusion in microfluidic cross-flow: separation of white blood cells from whole blood and exchange of medium in a continuous flow

We describe a microfluidic technique for separation of particles and cells and a device that employs this technique to separate white blood cells (WBC) from whole human blood. The separation is performed in cross-flow in an array of microchannels with a deep main channel and large number of orthogonal, shallow side channels. As a suspension of particles advances through the main channel, a perfusion flow through the side channels gradually exchanges the medium of the suspension and washes away particles that are sufficiently small to enter the shallow side channels. The microfluidic device is tested with a suspension of polystyrene beads and is shown to efficaciously exchange the carrier medium while retaining all beads. In tests with whole human blood, the device is shown to reduce the content of red blood cells (RBC) by a factor of approximately 4000 with retention of 98% of WBCs. The ratio between WBCs and RBCs reached at an outlet of the device is 2.4 on average. The device is made of a single cast of poly(dimethylsiloxane) sealed with a cover glass and is simple to fabricate. The proposed technique of separation by perfusion in continuous cross-flow could be used to enrich rare populations of cells based on differences in size, shape, and deformability.     

A ferrofluid guided system for the rapid separation of the non-magnetic particles in a microfluidic device

A microfluidic device was fabricated via UV lithography technique to separate non-magnetic fluoresbrite carboxy microspheres (approximately 4.5 microm) in the pH 7 ferrofluids made of magnetite nanoparticles (approximately 10 nm). A mixture of microspheres and ferrofluid was injected to a lithographically developed Y shape microfluidic device, and then by applying the external magnet fields (0.45 T), the microspheres were clearly separated into different channels because of the magnetic force acting on those non-magnetic particles. During this study, various pumping speeds and particle concentrations associated with the various distances between the magnet and the microfluidic device were investigated for an efficient separation. This study may be useful for the separation of biological particles, which are very sensitive to pH value of the solutions.     

Multi-breath dry powder inhaler for delivery of cohesive powders in the treatment of bronchiectasis

A series of co-engineered macrolide–mannitol particles were successfully prepared using azithromycin (AZ) as a model drug. The formulation was designed to target local inflammation and bacterial colonization, via the macrolide component, while the mannitol acted as mucolytic and taste-masking agent. The engineered particles were evaluated in terms of their physico-chemical properties and aerosol performance when delivered via a novel high-payload dry powder Orbital^? inhaler device that operates via multiple inhalation manoeuvres. All formulations prepared were of suitable size for inhalation drug delivery and contained a mixture of amorphous AZ with crystalline mannitol. A co-spray dried formulation containing 200?mg of 50:50?w/w AZ: mannitol had 57.6%?±?7.6% delivery efficiency with a fine particle fraction (≤6.8?µm) of the emitted aerosol cloud being 80.4%?±?1.1%, with minimal throat deposition (5.3?±?0.9%). Subsequently, it can be concluded that the use of this device in combination with the co-engineered macrolide–mannitol therapy may provide a means of treating bronchiectasis.     

MOFBOTS: Metal–Organic‐Framework‐Based Biomedical Microrobots

Motile metal?organic frameworks (MOFs) are potential candidates to serve as small‐scale robotic platforms for applications in environmental remediation, targeted drug delivery, or nanosurgery. Here, magnetic helical microstructures coated with a kind of zinc‐based MOF, zeolitic imidazole framework‐8 (ZIF‐8), with biocompatibility characteristics and pH‐responsive features, are successfully fabricated. Moreover, it is shown that this highly integrated multifunctional device can swim along predesigned tracks under the control of weak rotational magnetic fields. The proposed systems can achieve single‐cell targeting in a cell culture media and a controlled delivery of cargo payloads inside a complex microfluidic channel network. This new approach toward the fabrication of integrated multifunctional systems will open new avenues in soft microrobotics beyond current applications.     

Energetics and Escape of Interchain‐Delocalized Ion Pairs in Nonpolar Media

The electron acceptor F4TCNQ p‐dopes aggregates “nanowires” of poly(3‐hexylthiophene) in nonpolar solvents but does not dope unaggregated chains. The standard free energy change for the charge transfer to form an ion pair is ΔG°_et = ‐0.21 eV. The dissociation constant to produce free ions in toluene by DC conductivity is K°_d = 1 × 10^‐8 ± 50% (ΔG°_d = 0.48 ± 0.05 eV). This remarkably large K°_d, for ions in such a low polarity medium, may reflect interchain delocalization of the hole. The particular characteristics of this material system enables determination of both ΔG°_et and ΔG°_d, to find the overall free energy change from the two neutral species to completely separated ions in nonpolar media. It is endergonic by +0.27 ± 0.05 eV in contrast to ‐0.6 eV estimated from reported HOMO LUMO differences, illustrating the challenges that persist in determining such energetics. Steady state microwave conductivity experiments on doped aggregates confirm that holes in the aggregates cannot easily escape their dopant counterion, but at higher dopant concentrations, holes become mobile. These results provide insight into the mechanisms of charge separation involving intermolecularly delocalized charges in nonpolar media, an integral process in organic photovoltaic devices and doped molecular films.     

Rapid Production of Cell‐Laden Microspheres Using a Flexible Microfluidic Encapsulation Platform

This study establishes a novel microfluidic platform for rapid encapsulation of cells at high densities in photocrosslinkable microspherical hydrogels including poly(ethylene glycol)‐diacrylate, poly(ethylene glycol)‐fibrinogen, and gelatin methacrylate. Cell‐laden hydrogel microspheres are advantageous for many applications from drug screening to regenerative medicine. Employing microfluidic systems is considered the most efficient method for scale‐up production of uniform microspheres. However, existing platforms have been constrained by traditional microfabrication techniques for device fabrication, restricting microsphere diameter to below 200 µm and making iterative design changes time‐consuming and costly. Using a new molding technique, the microfluidic device employs a modified T‐junction design with readily adjustable channel sizes, enabling production of highly uniform microspheres with cell densities (10–60 million cells mL^?1) and a wide range of diameters (300–1100 µm), which are critical for realizing downstream applications, through rapid photocrosslinking (≈1 s per microsphere). Multiple cell types are encapsulated at rates of up to 1 million cells per min, are evenly distributed throughout the microspheres, and maintain high viability and appropriate cellular activities in long‐term culture. This microfluidic encapsulation platform is a valuable and readily adoptable tool for numerous applications, including supporting injectable cell therapy, bioreactor‐based cell expansion and differentiation, and high throughput tissue sphere‐based drug testing assays.     

A Radial Flow Microfluidic Device for Ultra‐High‐Throughput Affinity‐Based Isolation of Circulating Tumor Cells

Circulating tumor cells (CTCs) are believed to play an important role in metastasis, a process responsible for the majority of cancer‐related deaths. But their rarity in the bloodstream makes microfluidic isolation complex and time‐consuming. Additionally the low processing speeds can be a hindrance to obtaining higher yields of CTCs, limiting their potential use as biomarkers for early diagnosis. Here, a high throughput microfluidic technology, the OncoBean Chip, is reported. It employs radial flow that introduces a varying shear profile across the device, enabling efficient cell capture by affinity at high flow rates. The recovery from whole blood is validated with cancer cell lines H1650 and MCF7, achieving a mean efficiency >80% at a throughput of 10 mL h^?1 in contrast to a flow rate of 1 mL h^?1 standardly reported with other microfluidic devices. Cells are recovered with a viability rate of 93% at these high speeds, increasing the ability to use captured CTCs for downstream analysis. Broad clinical application is demonstrated using comparable flow rates from blood specimens obtained from breast, pancreatic, and lung cancer patients. Comparable CTC numbers are recovered in all the samples at the two flow rates, demonstrating the ability of the technology to perform at high throughputs.     

Study of microscale hydraulic jump phenomenon for hydrodynamic trap-and-release of microparticles

Easy trap-and-release of microparticles is necessary to study biological cellular behavior. The hydraulic jump phenomenon inspired us to conceive a microfluidic device for the hydrodynamic trap-and-release of microparticles. A sudden height increase in a microfluidic channel leads to a dramatic decrease in flow velocity, allowing effective trapping of the microparticles by energy conversion. The trapped particles can be released by stronger inertial force based on simply increasing the flow velocity. We present a systematic, numerical study of trap-and-release of the microparticles using multiphase Navier-Stokes equations. Effect of geometry flow velocity, particle diameter, and adhesion force on trap-and-release was studied.     

On-chip high-speed sorting of micron-sized particles for high-throughput analysis

A new design of particle sorting chip is presented. The device employs a dielectrophoretic gate that deflects particles into one of two microfluidic channels at high speed. The device operates by focussing particles into the central streamline of the main flow channel using dielectrophoretic focussing. At the sorting junction (T- or Y-junction) two sets of electrodes produce a small dielectrophoretic force that pushes the particle into one or other of the outlet channels, where they are carried under the pressure-driven fluid flow to the outlet. For a 40 microm wide and high channel, it is shown that 6 microm diameter particles can be deflected at a rate of 300/s. The principle of a fully automated sorting device is demonstrated by separating fluorescent from non-fluorescent latex beads.     

Sheathless hydrophoretic particle focusing in a microchannel with exponentially increasing obstacle arrays

We present a novel microfluidic device with exponentially increasing obstacle arrays to enable sheathless particle focusing. The anisotropic fluidic resistance of slant obstacles generates transverse flows, along which particles are focused to one sidewall. In the successive channel with exponentially increasing widths, bent obstacles extended from the slant obstacles increase the focusing efficiency of the particles. With the device, we achieved the focusing efficiency of 76%, 94%, and 98% for 6, 10, and 15 microm beads, respectively. The focusing efficiency of the particles can be further improved in the devices with more extension steps. In addition, using the microfluidic devices with the symmetric structure of the slant and bent obstacles, we achieved complete focusing of biological cells to the centerline of a channel within 1.7% coefficient of variation. The results demonstrated the sheathless hydrophoretic focusing of microparticles and cells with the advantages of a sheathless method, passive operation, single channel, and flow rate independence.