Sheathless Focusing Research Articles

Bipolar Electrode-based Sheath-Less Focusing and Continuous Acoustic Sorting of Particles and Cells in an Integrated Microfluidic Device.

Sheath-less focusing and sorting of cells or particles is an important preprocessing step in a variety of biochemical applications. Most of the previous sorting methods depend on the use of sheath flows to realize efficient cell focusing. The sheath flow dilutes the sample and requires precise flow control via additional channels. We, for the first time, reported a method of bipolar electrode (BPE)-based sheath-less focusing, switching, and tilted-angle standing surface acoustic wave-based sorting of cells and particles in continuous flow. The device consists of a piezoelectric substrate with a pair of BPEs for focusing and switching, and a pair of interdigitated transducers for cell sorting. Smaller cells experience a weak acoustic force and reach the lower outlet, whereas larger cells are subjected to a strong acoustic force such that they are propelled toward the upper outlet. We first validate the device functionality by sorting 5 and 8 μm PS beads with a high sorting efficiency. The working and deflection regions were increased by propelling the particle beam toward the bottom edge of BPE via changing the applied voltage of BPE, further improving the sorting performance with high efficiency (94%) and purity (92%). We then conducted a verification for sorting THP-1 and yeast cells, and the efficiency and purity reached 90.7 and 91.5%, respectively. This integrated device eliminates the requirement of balancing the flow of several sheath inlets and provides a robust and unique approach for cell sorting applications, showing immense promise in various applications, such as medical diagnosis, drug delivery, and personalized medicine.

Geometry-Dependent Efficiency of Dean-Flow Affected Lateral Particle Focusing and Separation in Periodically Inhomogeneous Microfluidic Channels.

In this study, inertial focusing phenomenon was investigated, which can be used as a passive method for sample preparation and target manipulation in case of particulate suspensions. Asymmetric channel geometry was designed to apply additional inertial forces besides lift forces to promote laterally ordered particles to achieve sheathless focusing or size-dependent sorting. The evolving hydrodynamic forces were tailored with altered channel parameters (width and height), and different flow rates, to get a better understanding of smaller beads’ lateral migration. Fluorescent beads (with the diameter of 4.8 µm and 15.8 µm) were used to distinguish the focusing position in continuous flow, and experimental results were compared to in silico models for particle movement prediction, made in COMSOL Multiphysics. The focusing behaviour of the applied microfluidic system was mainly characterised for particle size in the range close to blood cells and bacteria.

A sheathless inertial focusing technique for optofluidic devices

Here, we demonstrate single-position, three-dimensional (3D) focusing of cells or micron-sized particles in the range 0.175 < a/w < 0.9 (a, cell or particle diameter; w, width of the microchannel), on a single-layer, single-channel microfluidic chip, which totally alleviates the need of using any sheath flow or external force, making the microfluidic chip standalone operational. The focusing is a result of inertial microfluidic hydrodynamic forces such as inertial lift forces and Dean drag forces, which are determined by the geometry of microchannel. With the microfluidic channel comprising a series of radially increasing, uniform semi-arcs interleaved by linear sections, sheathless focusing at flow rates up to 700 μL/min is achieved in our study. The result can be well explained by a developed empirical model relating the ratio of inertial lift forces and Dean drag forces, and the geometrical parameters of the microchannel. Following this approach, we illustrate experimental characterization of micron-sized sample focusing using fluorescent microparticles, pancreatic cancer cells, and macrophages under Reynolds number flows ranging between 20 and 153. We foresee the single-position focusing outcome of the proposed sheathless chip design in developing portable microfluidic and optofluidic devices for in vitro theranostics.

Tunable, Sheathless Focusing of Diamagnetic Particles in Ferrofluid Microflows with a Single Set of Overhead Permanent Magnets.

There has been increasing interest in the use of magnetic fluids to manipulate diamagnetic particles in microfluidic devices. Current methods for diamagnetic-particle focusing in magnetic fluids require either a pair of repulsive magnets or a diamagnetic sheath flow. We demonstrate herein a tunable, sheathless focusing of diamagnetic particles in a microchannel ferrofluid flow with a single set of overhead permanent magnets. Particles are focused into a single stream near the bottom wall of a straight rectangular microchannel, where a magnetic-field minimum is formed as a result of the magnetization of the ferrofluid. This focusing can be readily switched off and on by removing and replacing the permanent magnets. More importantly, the particle-focusing position can be tuned by shifting the magnets with respect to the microchannel. We perform a systematic experimental study of the parametric effects of the fluid-particle-channel system on diamagnetic-particle focusing in terms of a defined particle-focusing effectiveness.

Sheathless Focusing and Separation of Microparticles Using Tilted-Angle Traveling Surface Acoustic Waves.

Sheathless focusing and separation of microparticles is an important preprocessing step in various biochemical assays in which enriched sample isolation is critical. Most of the previous microfluidic particle separation techniques have used sheath flows to achieve efficient sample focusing. The sheath flow dilutes the analyte and requires additional microchannels and accurate flow control. We demonstrated a tilted-angle traveling surface acoustic wave (taTSAW)-based sheathless focusing and separation of particles in a continuous flow. The proposed device consists of a piezoelectric substrate with a pair of interdigitated transducers (IDTs) deposited at two different angles relative to the flow direction. A Y-shaped polydimethylsiloxane (PDMS) microchannel having one inlet and two outlet ports was positioned on top of the IDTs such that the acoustic energy coupling into the fluid was maximized and wave attenuation by the PDMS walls was minimized. The two IDTs independently produced high-frequency taTSAWs, which propagated at ±30° with respect to the flow direction and imparted a direct acoustic radiation force onto the target particles. A sample mixture of 4.8 and 3.2 μm particles was focused and then separated by the actuation of the IDTs at 194 and 136 MHz frequencies, respectively, without using an additional sheath flow. The proposed taTSAW-based particle separation device offered a high purity >99% at the both outlets over a wide range of flow speeds (up to 83.3 mm/s).

Cellular immunity monitoring in long-duration spaceflights based on an automatic miniature flow cytometer

Monitoring cellular immunity is essential for long-duration spaceflight missions. To address this need, we developed an automatic miniature flow cytometer that is targeted to monitor the cellular immune function of astronauts or passengers during long-duration spaceflights. Automatic sample preparation, a disposable sheathless focusing chip, optical self-alignment, and side scatter (SSC) detection capability were developed to meet the strict requirements of spaceflight applications. SSC and four colors of fluorescence are measured to realize lymphocyte subsets counted from whole blood. This technology was used in four-subject 180-day controlled ecological life support system (CELSS) experiments to monitor the cellular immunity of four crewmembers by counting the lymphocyte subsets (total CD3+, CD3+CD4+, and CD3+CD8+ cells). The microgravity compatibility was successfully verified on a parabolic flight. Thus, the presented system was proven a feasible solution for health care applications during long-duration manned spaceflight.

Sheathless Focusing and Separation of Diverse Nanoparticles in Viscoelastic Solutions with Minimized Shear Thinning.

Viscoelastic microfluidics becomes an efficient and label-free hydrodynamic technology to enrich and separate micrometer-scale particles, including blood cells, circulating tumor cells, and bacteria. However, the manipulation of nanoscale particles by viscoelastic microfluidics remains a major challenge, because the viscoelastic force acting on the smaller particle decreases dramatically. In contrast to the commonly used polymer solutions of high molecular weight, herein we utilize the aqueous solutions of poly(ethylene oxide) (PEO) of low molecular weight with minimized shear thinning but sufficient elastic force for high-quality focusing and separation of various nanoparticles. The focusing efficiencies of 100 nm polystyrene (PS) nanoparticles and λ-DNA molecules are 84% and 85%, respectively, in a double spiral microchannel, without the aid of sheath flows. Furthermore, we demonstrate the size-based viscoelastic separation of two sets of binary mixtures-100/2000 nm PS particles and λ-DNA molecules/blood platelets-all achieving separation efficiencies of >95% in the same device. Our proposal technique would be a promising approach for enrichment/separation of the nanoparticles encountered in applications of analytical chemistry and nanotechnology.

Sheathless and high throughput sorting of paramagnetic microparticles in a magneto-hydrodynamic microfluidic device.

Sorting of microorganisms and particles from a mixture is critical for numerous biotechnological and medical applications. Several sorting methods such as pinched flow fractionation (PFF), optical sorting, dielectrophoresis, acoustic separation, magnetophoresis and deterministic lateral displacement (DLD) have been reported in literature. But most of these methods lack ideal characteristics of a sorter such as ability to process at high throughput, simple design, non-complicated fabrication method, sheathless operation and high purity in separation. In this paper, we have introduced a novel sorting technique by integrating focusing of magnetic particles in a narrow microchannel with their hydrodynamic separation at a downstream expansion channel which meets majority of the aforementioned characteristics. To achieve this, the sheathless focusing of paramagnetic microparticles in the narrow microchannel and their deflection at the expansion channel were first studied at various flow rates (0.5-5 ml h-1). Then, a mixture of 5 and 11 μm paramagnetic particles was introduced into the device and their separation was examined quantitatively. It was found that the magnetic particles were focused along the wall of channel, however their centers were positioned on two distinct streamlines owing to difference in their sizes. Hence, these two particles were found separated from each other as they flew into the expansion region. This technique of size based separation of paramagnetic particles works at a high throughput of 107 particles per hour and offers more than 98% purity in sorting.

Fabrication and Operation of Acoustofluidic Devices Supporting Bulk Acoustic Standing Waves for Sheathless Focusing of Particles.

Acoustophoresis refers to the displacement of suspended objects in response to directional forces from sound energy. Given that the suspended objects must be smaller than the incident wavelength of sound and the width of the fluidic channels are typically tens to hundreds of micrometers across, acoustofluidic devices typically use ultrasonic waves generated from a piezoelectric transducer pulsating at high frequencies (in the megahertz range). At characteristic frequencies that depend on the geometry of the device, it is possible to induce the formation of standing waves that can focus particles along desired fluidic streamlines within a bulk flow. Here, we describe a method for the fabrication of acoustophoretic devices from common materials and clean room equipment. We show representative results for the focusing of particles with positive or negative acoustic contrast factors, which move towards the pressure nodes or antinodes of the standing waves, respectively. These devices offer enormous practical utility for precisely positioning large numbers of microscopic entities (e.g., cells) in stationary or flowing fluids for applications ranging from cytometry to assembly.

Fabrication and Operation of Acoustofluidic Devices Supporting Bulk Acoustic Standing Waves for Sheathless Focusing of Particles

Acoustophoresis refers to the displacement of suspended objects in response to directional forces from sound energy. Given that the suspended objects must be smaller than the incident wavelength of sound and the width of the fluidic channels are typically tens to hundreds of micrometers across, acoustofluidic devices typically use ultrasonic waves generated from a piezoelectric transducer pulsating at high frequencies (in the megahertz range). At characteristic frequencies that depend on the geometry of the device, it is possible to induce the formation of standing waves that can focus particles along desired fluidic streamlines within a bulk flow. Here, we describe a method for the fabrication of acoustophoretic devices from common materials and clean room equipment. We show representative results for the focusing of particles with positive or negative acoustic contrast factors, which move towards the pressure nodes or antinodes of the standing waves, respectively. These devices offer enormous practical utility for precisely positioning large numbers of microscopic entities (e.g., cells) in stationary or flowing fluids for applications ranging from cytometry to assembly.

Inertial modulation of hydrophoretic cell sorting and focusing

This paper demonstrates a method of modulating hydrophoretic focusing and sorting by inertial forces in a microfluidic device. At low Reynolds number (Re), hydrophoresis enables size-based sorting by generating specific equilibrium positions for differently sized particles. As increasing Re, hydrophoresis becomes coupled with inertial effects that bias the equilibrium positions to create a narrow focusing stream. Using this method, we achieved both size-based sorting and sheathless focusing in a device.

Particle focusing in microfluidic devices

Focusing particles (both biological and synthetic) into a tight stream is usually a necessary step prior to counting, detecting, and sorting them. The various particle focusing approaches in microfluidic devices may be conveniently classified as sheath flow focusing and sheathless focusing. Sheath flow focusers use one or more sheath fluids to pinch the particle suspension and thus focus the suspended particles. Sheathless focusers typically rely on a force to manipulate particles laterally to their equilibrium positions. This force can be either externally applied or internally induced by channel topology. Therefore, the sheathless particle focusing methods may be further classified as active or passive by the nature of the forces involved. The aim of this article is to introduce and discuss the recent developments in both sheath flow and sheathless particle focusing approaches in microfluidic devices.

Sheathless Focusing of Microbeads and Blood Cells Based on Hydrophoresis

This paper presents a microfluidic device for sheathless focusing of microbeads and blood cells based on a hydrophoretic platform comprising a V-shaped obstacle array (VOA). The VOA generates lateral pressure gradients that induce helical recirculations. Following the focusing flow particles passing through the VOA are focused in the center of the channel. In the device, the focusing pattern can be modulated by varying the gap height of the VOA. To achieve complete focusing within 4.4% coefficient of variation, the relative size differences between the gap and the particle were 3 and 4 microm for 10 and 15 microm beads, respectively. Red blood cells were used to study the hydrophoretic focusing pattern of biconcave, disk-shaped particles.

Ultrasonic particle‐concentration for sheathless focusing of particles for analysis in a flow cytometer

The development of inexpensive small flow cytometers is recognized as an important goal for many applications ranging from medical uses in developing countries for disease diagnosis to use as an analytical platform in support of homeland defense. Although hydrodynamic focusing is highly effective at particle positioning, the use of sheath fluid increases assay cost and reduces instrument utility for field and autonomous remote operations. This work presents the creation of a novel flow cell that uses ultrasonic acoustic energy to focus small particles to the center of a flowing stream for analysis by flow cytometry. Experiments using this flow cell are described wherein its efficacy is evaluated under flow cytometric conditions with fluorescent microspheres. Preliminary laboratory experiments demonstrate acoustic focusing of flowing 10-microm latex particles into a tight sample stream that is approximately 40 microm in diameter. Prototype flow cytometer measurements using an acoustic-focusing flow chamber demonstrated focusing of a microsphere sample to a central stream approximately 40 microm in diameter, yielding a definite fluorescence peak for the microspheres as compared with a broad distribution for unfocused microspheres. The flow cell developed here uses acoustic focusing, which inherently concentrates the sample particles to the center of the sample stream. This method could eliminate the need for sheath fluid, and will enable increased interrogation times for enhanced sensitivity, while maintaining high particle-analysis rates. The concentration effect will also enable the analysis of extremely dilute samples on the order of several particles per liter, at analysis rates of a few particles per second. Such features offer the possibility of a truly versatile low-cost portable flow cytometer for field applications.