Pure separation and sorting of microparticles from complex fluids are essential for biochemical analyses and clinical diagnostics. However, conventional techniques require highly complex and expensive labeling processes for high purity separation. In this study, we present a simple and label-free method for separating microparticles with high purity using the elasto-inertial characteristic of a non-Newtonian fluid in microchannel flow. At the inlet, particle-containing sample flow was pushed toward the side walls by introducing sheath fluid from the center inlet. Particles of 1 μm and 5 μm in diameter, which were suspended in viscoelastic fluid, were successfully separated in the outlet channels: larger particles were notably focused on the centerline of the channel at the outlet, while smaller particles continued flowing along the side walls with minimal lateral migration towards the centerline. The same technique was further applied to separate platelets from diluted whole blood. Through cytometric analysis, we obtained a purity of collected platelets of close to 99.9%. Conclusively, our microparticle separation technique using elasto-inertial forces in non-Newtonian fluid is an effective method for separating and collecting microparticles on the basis of size differences with high purity.

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Continuous Separation of Microparticles in a Microfluidic Channel via the Elasto-inertial Effect of Non-Newtonian Fluid

Inertial microfluidics has become a popular topic in microfluidics research for its good performance in particle manipulation and its advantages of simple structure, high throughput, and freedom from an external field. Compared with traditional microfluidic devices, the flow field in inertial microfluidics is between Stokes state and turbulence, whereas the flow is still regarded as laminar. However, many mechanical effects induced by the inertial effect are difficult to observe in traditional microfluidics, making particle motion analysis in inertial microfluidics more complicated. In recent years, the inertial migration effect in straight and curved channels has been explored theoretically and experimentally to realize on-chip manipulation with extensive applications from the ordinary manipulation of particles to biochemical analysis. In this review, the latest theoretical achievements and force analyses of inertial microfluidics and its development process are introduced, and its applications in circulating tumor cells, exosomes, DNA, and other biological particles are summarized. Finally, the future development of inertial microfluidics is discussed. Owing to its special advantages in particle manipulation, inertial microfluidics will play a more important role in integrated biochips and biomolecule analysis.

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Progress of Inertial Microfluidics in Principle and Application.

Modern microflow cytometers are sophisticated instruments capable of measuring multiple physical characteristics of a single cell simultaneously as the cells flow in suspension through a measuring device. Cytometers are the tool of choice for the high-speed acquisition and analysis of large single cell populations. However, traditional devices lack the ability to provide intracellular spatial information. In the past few decades, various flow cytometer systems with the ability to combine cell/particle detection and the acquisition of two or three-dimensional spatial information have been proposed. However, these devices suffer a number of drawbacks Accordingly, the problem of developing more sophisticated microflow cytometers based on electrical impedance, optical detection and image analysis methods has received significant attention in the literature. This review describes some of the major advances made in the microfluidic cytometry field over the past ten years. The review focuses specifically on recent proposals for enhanced microfluidic focusing techniques and detection/analysis methods, respectively. Overall, the review provides a useful insight into the microflow cytometer technology field for both new and existing users.

Review and perspectives on microfluidic flow cytometers

Inertial microfluidics has gained significant attention since first being proposed in 2007 owing to the advantages of simplicity, high throughput, precise manipulation, and freedom from an external field. Superior performance in particle focusing, filtering, concentrating, and separating has been demonstrated. As a passive technology, inertial microfluidics technology relies on the unconventional use of fluid inertia in an intermediate Reynolds number range to induce inertial migration and secondary flow, which depend directly on the channel structure, leading to particle migration to the lateral equilibrium position or trapping in a specific cavity. With the advances in micromachining technology, many channel structures have been designed and fabricated in the past decade to explore the fundamentals and applications of inertial microfluidics. However, the channel innovations for inertial microfluidics have not been discussed comprehensively. In this review, the inertial particle manipulations and underlying physics in conventional channels, including straight, spiral, sinusoidal, and expansion-contraction channels, are briefly described. Then, recent innovations in channel structure for inertial microfluidics, especially channel pattern modification and unconventional cross-sectional shape, are reviewed. Finally, the prospects for future channel innovations in inertial microfluidic chips are also discussed. The purpose of this review is to provide guidance for the continued study of innovative channel designs to improve further the accuracy and throughput of inertial microfluidics.

Channel innovations for inertial microfluidics

The manipulation of cells and particles suspended in viscoelastic fluids in microchannels has drawn increasing attention, in part due to the ability for single-stream three-dimensional focusing in simple channel geometries. Improvement in the understanding of non-Newtonian effects on particle dynamics has led to expanding exploration of focusing and sorting particles and cells using viscoelastic microfluidics. Multiple factors, such as the driving forces arising from fluid elasticity and inertia, the effect of fluid rheology, the physical properties of particles and cells, and channel geometry, actively interact and compete together to govern the intricate migration behavior of particles and cells in microchannels. Here, we review the viscoelastic fluid physics and the hydrodynamic forces in such flows and identify three pairs of competing forces/effects that collectively govern viscoelastic migration. We discuss migration dynamics, focusing positions, numerical simulations, and recent progress in viscoelastic microfluidic applications as well as the remaining challenges. Finally, we hope that an improved understanding of viscoelastic flows in microfluidics can lead to increased sophistication of microfluidic platforms in clinical diagnostics and biomedical research. Insights into the dynamic behavior of biological fluids on the microscale are enabling more efficient analysis of cells, bacteria, and other small biological particles for research and clinical diagnostics. Blood, saliva and other biofluids have viscoelastic properties, which means that they exhibit both viscous and elastic behaviors depending on the forces to which they are subjected. These properties shape the migration behaviors of suspended bioparticles in unique ways. Jian Zhou and Ian Papautsky of the University of Illinois at Chicago have reviewed current progress in understanding the migration behaviors in such fluids within microfluidic devices. The authors discuss design principles that have enabled the development of microfluidic systems capable of separating and purifying cells, bacteria, and small vesicles from highly heterogeneous biological specimens, and highlight future challenges that need to be addressed.

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Viscoelastic microfluidics: progress and challenges.

Massive growth of the microfluidics field has triggered numerous advances in focusing, separating, ordering, concentrating, and mixing of microparticles. Microfluidic systems capable of performing these functions are rapidly finding applications in industrial, environmental, and biomedical fields. Passive and label-free methods are one of the major categories of such systems that have received enormous attention owing to device operational simplicity and low costs. With new platforms continuously being proposed, our aim here is to provide an updated overview of the state of the art for passive label-free microparticle separation, with emphasis on performance and operational conditions. In addition to the now common separation approaches using Newtonian flows, such as deterministic lateral displacement, pinched flow fractionation, cross-flow filtration, hydrodynamic filtration, and inertial microfluidics, we also discuss separation approaches using non-Newtonian, viscoelastic flow. We then highlight the newly emerging approach based on shear-induced diffusion, which enables direct processing of complex samples such as untreated whole blood. Finally, we hope that an improved understanding of label-free passive sorting approaches can lead to sophisticated and useful platforms toward automation in industrial, environmental, and biomedical fields.

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Label-free microfluidic sorting of microparticles.

Fabrication of microfluidic devices by soft lithography is by far the most popular approach due to simplicity and low cost. In this approach PDMS (polydimethylsiloxane) is cast on a photoresist master to generate replicas that are then sealed against glass slides using oxygen plasma. In this work, we demonstrated fabrication of soft photolithography masters using lamination of ADEX dry film as an alternative to the now classic SU-8 resist masters formed by spin coating. Advantages of using ADEX dry film include the easily-achievable uniform thickness without edge bead; simplicity of the process with significant time savings due to non-sticky nature of the film; and fewer health concerns due to less toxic developing solution and antimony-free composition. As we demonstrate, the process can be performed in a low-cost improvised fabrication room in ambient light, in place of a conventional yellow-light cleanroom environment. We believe this approach holds the promise of delivering state-of-the-art microfluidic techniques to the broad field of biomedical and pharmaceutical research.

https://www.mdpi.com/2072-666X/10/3/192/pdf

Rapid Prototyping of Soft Lithography Masters for Microfluidic Devices Using Dry Film Photoresist in a Non-Cleanroom Setting.

Glass capillary tubes have been widely used in microfluidics for generating microdroplets and microfibers. Here, we report on the application of glass capillary to inertial focusing of microparticles and cells for high-throughput flow cytometry. Our device uses a commercially available capillary tube with a square cross-section. Wrapping the tube into a helical shape induces the Dean vortices that aid focusing of cells or microbeads into a single position. We investigated the inertial focusing of microbeads in the device at various Re and concentrations and demonstrated 3D focusing with ∼100% efficiency for a wide range of microparticle diameters. We integrated the device with a laser counting system and demonstrated continuous counting of 10 μm microbeads with a high throughput of 13 000 beads/s as well as counting of fluorescently labeled white blood cells in the diluted whole blood. The helical capillary device offers a number of key advantages, including rapid and ultra-low-cost plug-and-play fabrication, optical transparency, and full compatibility with bright field or fluorescent imaging, easy re-configurability of the device radius for tuning focusing behavior, and ability to rotate for easy side-wall observation. With precise and consistent 3D focusing of microbeads and cells with a wide range of sizes at high throughput and without the use of sheath flows, we envision that this simple capillary-based inertial microfluidic device will create new opportunities for this technique to be widely adopted in the laboratory research.

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A low-cost, plug-and-play inertial microfluidic helical capillary device for high-throughput flow cytometry.

Fabrication of microfluidic devices by soft lithography is by far the most popular approach due to its simplicity and low cost. The approach relies on casting of elastomers, such as polydimethylsiloxane (PDMS), on masters fabricated from photoresists on silicon substrates. These masters, however, can be expensive, complicated to fabricate, and fragile. Here we describe an optimized replica molding approach to preserve the original masters by heat molding of polycarbonate (PC) sheets on PDMS molds. The process is faster and simpler than previously reported methods and does not result in a loss of resolution or aspect ratio for the features. The generated PC masters were used to successfully replicate a wide range of microfluidic devices, including rectangular channels with aspect ratios from 0.025 to 7.3, large area spiral channels, and micropost arrays with 5 µm spacing. Moreover, fabrication of rounded features, such as semi-spherical microwells, was possible and easy. Quantitative analysis of the replicated features showed variability of <2%. The approach is low cost, does not require cleanroom setting or hazardous chemicals, and is rapid and simple. The fabricated masters are rigid and survive numerous replication cycles. Moreover, damaged or missing masters can be easily replaced by reproduction from previously cast PDMS replicas. All of these advantages make the PC masters highly desirable for long-term preservation of soft lithography masters for microfluidic devices.

Polycarbonate Masters for Soft Lithography.

Abstract Particle migration dynamics in viscoelastic fluids in spiral channels have attracted interest in recent years due to potential applications in the 3D focusing and label-free sorting of particles and cells. Despite a number of recent studies, the underlying mechanism of Dean-coupled elasto-inertial migration in spiral microchannels is not fully understood. In this work, for the first time, we experimentally demonstrate the evolution of particle focusing behavior along a channel downstream length at a high blockage ratio. We found that flow rate, device curvature, and medium viscosity play important roles in particle lateral migration. Our results illustrate the full focusing pattern along the downstream channel length, with side-view imaging yielding observations on the vertical migration of focused streams. Ultimately, we anticipate that these results will offer a useful guide for elasto-inertial microfluidics device design to improve the efficiency of 3D focusing in cell sorting and cytometry applications. 

/pdf/evolution-of-focused-streams-for-viscoelastic-flow-in-spiral-1b1og9d3.pdf

Hua Gao

Papers

Label-free microfluidic sorting of microparticles.

Rapid Prototyping of Soft Lithography Masters for Microfluidic Devices Using Dry Film Photoresist in a Non-Cleanroom Setting.

A low-cost, plug-and-play inertial microfluidic helical capillary device for high-throughput flow cytometry.

Polycarbonate Masters for Soft Lithography.

Evolution of focused streams for viscoelastic flow in spiral microchannels