Hydrodynamic Focusing Microfluidic Device Research Articles

Synthesis of Poly(N-isopropylacrylamide) Janus Microhydrogels for Anisotropic Thermo-responsiveness and Organophilic/Hydrophilic Loading Capability.

Janus microparticles are compartmentalized particles with differing molecular structures and/or functionality on each of their two sides. Because of this unique property, Janus microparticles have been recognized as a new class of materials, thereby attracting a great deal of attention from various research fields. The versatility of these microparticles has been exemplified through their uses as building blocks for self-assembly, electrically responsive actuators, emulsifiers for painting and cosmetics, and carriers for drug delivery. This study introduces a detailed protocol that explicitly describes a synthetic method for designing novel Janus microhydrogels composed of a single base material, poly(N-isopropylacrylamide) (PNIPAAm). Janus microdroplets are firstly generated via a hydrodynamic focusing microfluidic device (HFMD) based on the separation of a supersaturated aqueous NIPAAm monomer solution and subsequently polymerized through exposure to UV irradiation. The resulting Janus microhydrogels were found to be entirely composed of the same base material, featured an easily identifiable compartmentalized morphology, and exhibited anisotropic thermo-responsiveness and organophilic/hydrophilic loading capability. We believe that the proposed method introduces a novel hydrogel platform with the potential for advanced synthesis of multi-functional Janus microhydrogels.

Synthesis of Poly(<em>N</em>-isopropylacrylamide) Janus Microhydrogels for Anisotropic Thermo-responsiveness and Organophilic/Hydrophilic Loading Capability

Janus microparticles are compartmentalized particles with differing molecular structures and/or functionality on each of their two sides. Because of this unique property, Janus microparticles have been recognized as a new class of materials, thereby attracting a great deal of attention from various research fields. The versatility of these microparticles has been exemplified through their uses as building blocks for self-assembly, electrically responsive actuators, emulsifiers for painting and cosmetics, and carriers for drug delivery. This study introduces a detailed protocol that explicitly describes a synthetic method for designing novel Janus microhydrogels composed of a single base material, poly(N-isopropylacrylamide) (PNIPAAm). Janus microdroplets are firstly generated via a hydrodynamic focusing microfluidic device (HFMD) based on the separation of a supersaturated aqueous NIPAAm monomer solution and subsequently polymerized through exposure to UV irradiation. The resulting Janus microhydrogels were found to be entirely composed of the same base material, featured an easily identifiable compartmentalized morphology, and exhibited anisotropic thermo-responsiveness and organophilic/hydrophilic loading capability. We believe that the proposed method introduces a novel hydrogel platform with the potential for advanced synthesis of multi-functional Janus microhydrogels.

A simple approach for the fabrication of 3D microelectrodes for impedimetric sensing

In this paper, we present a very simple method to fabricate three-dimensional (3D) microelectrodes integrated with microfluidic devices. We form the electrodes by etching a microwire placed across a microchannel. For precise control of the electrode spacing, we employ a hydrodynamic focusing microfluidic device and control the width of the etching solution stream. The focused widths of the etchant solution and the etching time determine the gap formed between the electrodes. Using the same microfluidic device, we can fabricate integrated 3D electrodes with different electrode gaps. We have demonstrated the functionality of these electrodes using an impedimetric particle counting setup. Using 3D microelectrodes with a diameter of 25 μm, we have detected 6 μm-diameter polystyrene beads in a buffer solution as well as erythrocytes in a PBS solution. We study the effect of electrode spacing on the signal-to-noise ratio of the impedance signal and we demonstrate that the smaller the electrode spacing the higher the signal obtained from a single microparticle. The sample stream is introduced to the system using the same hydrodynamic focusing device, which ensures the alignment of the sample in between the electrodes. Utilising a 3D hydrodynamic focusing approach, we force all the particles to go through the sensing region of the electrodes. This fabrication scheme not only provides a very low-cost and easy method for rapid prototyping, but which can also be used for applications requiring 3D electric field focused through a narrow section of the microchannel.

Microfluidic synthesis of thermo-responsive poly(N-isopropylacrylamide)–poly(ethylene glycol) diacrylate microhydrogels as chemo-embolic microspheres

In this paper, we have successfully synthesized and characterized poly(N-isopropylacrylamide) (PNIPAAm)–poly(ethylene glycol) diacrylate (PEGDA) microhydrogels. Various combinations of PNIPAAm-PEGDA microhydrogels were fabricated by the generation of monodisperse microdroplets whose sizes were comparable to a blood vessel of 260 and 320 µm with the help of a hydrodynamic focusing microfluidic device (HFMD), followed by synthesis of the microhydrogels through UV irradiation to the microdroplets. The thermo-responsive behaviors of the various microhydrogels were investigated by changing the PEGDA crosslinker concentration, which was found to be a dominant factor in tuning the shrinkage ratio in response to temperature change. As an in vitro embolization performance evaluation of the microhydrogels as chemo-embolic microspheres, the deliverability of the microhydrogels through a microcatheter was first confirmed and the compact occlusion of a channel was demonstrated based on a tapered microchannel in response to the temperature increase to physiological temperature of 36 °C. The controlled release behavior of the fluorescent dye from the microhydrogel was also investigated for chemotherapeutic purposes as a proof of concept study. The PNIPAAm-PEGDA microhydrogels could be used widely in embolization procedures based on the advantages of tunable thermo-responsive and controlled release behaviors.

Thrombin generation and fibrin formation under flow on biomimetic tissue factor‐rich surfaces

Blood flow regulates coagulation and fibrin assembly by controlling the rate of transport of zymogens, enzymes and plasma proteins to and from the site of an injury. The objective of this work was to define the hemodynamic conditions under which fibrin can form under flow on tissue factor (TF)-rich substrates. TF-coated silica beads (~800nm) were patterned into 18-85-μm spots. Normal pooled plasma and factors VIII, IX and XI deficient plasmas were perfused over the beads coated with 0.08, 0.8 and 8 molecules-TFμm(-2) at shear rates of 50-1000s(-1) . Fibrin deposition and thrombin generation were measured by fluorescence microscopy in a hydrodynamic focusing microfluidic device. Fibrin deposition was supported on patterned bead spots, but not planar TF substrates at the same surface TF concentration. There was a threshold spot size and a shear rate dependent TF concentration that was necessary to support fibrin polymerization. FVIII and FIX had minor effects on fibrin dynamics at 8 molecules-TFμm(-2) , but were essential at 0.8 molecules-TFμm(-2) . The absence of FXI influenced thrombin generation and fibrin deposition at both 0.8 and 8 molecules-TFμm(-2) . These results show that fibrin deposition requires perturbations in the flow field that protect reactions from dilution by flow under venous and arterial conditions. FVIII and FIX have a modest effect on fibrin deposition at high TF concentrations, but are necessary for fibrin deposition at low TF concentrations. FXI amplifies thrombin generation under flow at both low and high TF concentrations.

Adhesive Transfer Soft Lithography: Low-Cost and Flexible Rapid Prototyping of Microfluidic Devices

A simple and low-cost approach was proposed for prototyping PDMS based microfluidic devices by transferringadhesive film microstructures onto a flexible substrate as a mould for PDMS replicas. The microstructures were engravedon an adhesive coated film using a commercial cutting plotter and then transferred (or laminated) onto a flexiblesubstrate, allowing for engraved isolated patterns. The proposed technique was demonstrated by a hydrodynamic focusingmicrofluidic device, having splitting and re-combining sheath channels. The whole processing could be finished within 1h in a normal laboratory environment. This approach offers an easy, flexible and rapid prototyping of microfluidic andlab-on-a-chip devices to users without expertise in microfabrication. In addition, by minimizing the use of chemicals, theprocess becomes more environmentally friendly than conventional photolithography based micro-fabrication techniques.

Analysis of intracellular reducing levels in human hepatocytes on three‐dimensional focusing microchip

A novel three-dimensional hydrodynamic focusing microfluidic device integrated with high-throughput cell sampling and detection of intracellular contents is presented. It has a pivotal role in maintaining the reducing environment in cells. Intracellular reducing species such as vitamin C and glutathione in normal and tumor cells were labeled by a newly synthesized 2,2,6,6-tetramethyl-piperidine-1-oxyl-based fluorescent probe. Hepatocytes are adherent cells, which are prone to attaching to the channel surface. To avoid the attachment of cells on the channel surface, a single channel microchip with three sheath-flow channels located on both sides of and below the sampling channel was developed. Hydrostatic pressure generated by emptying the sample waste reservoir was used as driving force of fluid on the microchip. Owing to the difference between the liquid levels of the reservoirs, the labeled cells were three-dimensional hydrodynamically focused and transported from the sample reservoir to the sample waste reservoir. Hydrostatic pressure takes advantage of its ease of generation on a microfluidic chip without any external pressure pump, which drives three sheath-flow streams to constrain a sample flow stream into a narrow stream to avoid blockage of the sampling channel by adhered cells. The intracellular reducing levels of HepG2 cells and L02 cells were detected by home-built laser-induced fluorescence detector. The analysis throughput achieved in this microfluidic system was about 59-68 cells/min.

Effect of Flow Rates on Generation of Monodisperse Clay–Poly(N-isopropylacrylamide) Embolic Microspheres Using Hydrodynamic Focusing Microfluidic Device

Vascular embolization is a minimally invasive nonsurgical technique obstructing a blood vessel by lodgment of embolic materials to treat cancers and vascular lesions. In this paper, we have carried out a parametric study of generation of monodisperse clay–poly(N-isopropylacrylamide) (clay–PNIPAAm) embolic microspheres of which size is comparable to a blood vessel (about 400 µm). To achieve monodisperse water-phase clay/NIPAAm microdroplets, we have designed and fabricated a poly(dimethylsiloxane) (PDMS) hydrodynamic focusing microfluidic device (HFMD) for the generation of microdroplets with the affinity of continuous oil-phase fluid to the hydrophobic PDMS taken into account. We have investigated the influence of process-related flow conditions on the microdroplet generation to determine a proper processing window for obtaining monodisperse microdroplets with the fabricated HFMD. A parametric study of generation of monodisperse microdroplets was carried out by changing volumetric flow rates of two immiscible fluids within the determined processing window. For the suggested condition, the fabricated clay–PNIPAAm microspheres of about 400 µm in diameter showed an extremely narrow size distribution with a coefficient of variation of 0.41%. We have also showed the floatability of the fabricated clay–PNIPAAm microspheres in saline and the smooth passage of the microspheres through a commercially available microcatheter as in vitro characterization for embolization.

Three-dimensional (3D) hydrodynamic focusing for continuous sampling and analysis of adherent cells

A simple three-dimensional (3D) hydrodynamic focusing microfluidic device integrated with continuous sampling, rapid dynamic lysis, capillary electrophoretic (CE) separation and detection of intracellular content is presented. One of the major difficulties in microfluidic cell analysis for adherent cells is that the cells are prone to attaching to the channel surface. To solve this problem, a cross microfluidic chip with three sheath-flow channels located on both sides of and below the sampling channel was developed. With the three sheath flows around the sample solution-containing cells, the formed soft fluid wall prevents the cells from adhering to the channel surface. Labeled cells were 3D hydrodynamically focused by the sheath-flow streams and smoothly introduced into the cross-section one by one. The introduction of sheath-flow streams not only ensured single-cell sampling but avoided blockage of the sampling channel by adherent cells as well. The maximum rate for introduction of individual cells into the separation channel was about 151 cells min(-1). With electric field applied on the separation channel, the aligned cells were driven into the separation channel and rapidly lysed within 400 ms at the entry of the channel by sodium dodecylsulfate (SDS) added in the sheath-flow solution. The microfluidic system was evaluated by analysis of reduced glutathione (GSH) and reactive oxygen species (ROS) in single HepG2 cells. The average analysis throughput of ROS and GSH in single cells was 16-18 cells min(-1).

An efficient 3-dimensional hydrodynamic focusing microfluidic device by means of locally increased aspect ratio

In this paper, we have designed, fabricated and characterized an efficient 3- dimensional hydrodynamic focusing microfluidic device (3D-HFMD) in numerical and experimental ways. The 3D-HFMD achieved vertical focusing of core sample flow effectively in the pressure driven flow with the help of locally increased aspect ratio of thickness to width without any horizontal separation wall, thereby enabling us to realize 3D focusing after simple horizontal focusing. Extensive numerical analysis shows that the 3D-HFMD enhances the performance of vertical focusing compared with the previous designs over a wide range of the Reynolds numbers, even in the case of a the global aspect ratio less than 0.5. Experimental results from the fabricated 3D-HFMD confirmed that 3D hydrodynamic focusing could be successfully achieved, even when the dimension of the channel was kept consistent with that of general microfluidic channels. The 3D-HFMD suggested in this study could be applied to various integrated biological or chemical microfluidic devices.

Three-dimensional hydrodynamic focusing in two-layer polydimethylsiloxane (PDMS) microchannels

In this work, we designed and fabricated a three-dimensional hydrodynamic focusing microfluidic device. The device comprises a two-layer PDMS microchannel structure. There are four inlet ports and one outlet port. The fluids are all injected by syringe pumps. A sample flow stream was first vertically constrained into a narrow stream, and then horizontally focused on one small core region from a cross-section perspective, which is useful for cell/particle counting. We showed the numerical and experimental images of the focused stream shape from a cross-section perspective; experimental images were captured using a confocal fluorescence microscope. We also investigated the effect of channel aspect ratio on the vertical focusing effect using CFD simulations. The results showed that the sample flow can be focused successfully in the lower aspect ratio of the main channel (slightly greater than 0.5) in our design. Furthermore, the effect of the Reynolds number on the vertical focusing effect was also investigated. The numerical results showed that the rectangular-like shape of the focused stream from the cross-section perspective was deformed as the Reynolds number was high due to stronger secondary flows produced in the vertical focusing unit. This phenomenon was also demonstrated experimentally. The device only works well at low Reynolds numbers (approximately less than 5). The device can be integrated into an on-chip flow cytometer.