Nanoporous micro-element arrays for particle interception in microfluidic cell separation

The ability to control cell-surface interactions in order to achieve binding of specific cell types is a major challenge for microfluidic immunoaffinity cell capture systems. In the majority of existing systems, the functionalized capture surface is constructed of solid materials, where flow stagnation at the solid-liquid interface is detrimental to the convection of cells to the surface. We study the use of ultra-high porosity (99%) nanoporous micro-posts in microfluidic channels for enhancing interception efficiency of particles in flow. We show using both modelling and experiment that nanoporous posts improve particle interception compared to solid posts through two distinct mechanisms: the increase of direct interception, and the reduction of near-surface hydrodynamic resistance. We provide initial validation that the improvement of interception efficiency also results in an increase in capture efficiency when comparing nanoporous vertically aligned carbon nanotube (VACNT) post arrays with solid PDMS post arrays of the same geometry. Using both bacteria (～1 μm) and cancer cell lines (～15 μm) as model systems, we found capture efficiency increases by 6-fold and 4-fold respectively. The combined model and experimental platform presents a new generation of nanoporous microfluidic devices for cell isolation.     

Controlled release of reagents in capillary-driven microfluidics using reagent integrators

The integration and release of reagents in microfluidics as used for point-of-care testing is essential for an easy and accurate operation of these promising diagnostic devices. Here, we present microfluidic functional structures, which we call reagent integrators (RIs), for integrating and releasing small amounts of dried reagents (ng quantities and less) into microlitres of sample in a capillary-driven microfluidic chip. Typically, a RI is less than 1 mm(2) in area and has an inlet splitting into a central reagent channel, in which reagents can be loaded using an inkjet spotter, and two diluter channels. During filling of the microfluidic chip, spotted reagents reconstitute and exit the RI with a dilution factor that relates to the relative hydraulic resistance of the channels forming the RI. We exemplify the working principle of RIs by (i) distributing ～100 pg of horseradish peroxidase (HRP) in different volume fractions of a 1 μL solution containing a fluorogenic substrate for HRP and (ii) performing an immunoassay for C-reactive protein (CRP) using 450 pg of fluorescently labeled detection antibodies (dAbs) that reconstitute in ～5 to 30% of a 1 μL sample of human serum. RIs preserve the conceptual simplicity of lateral flow assays while providing a great degree of control over the integration and release of reagents in a stream of sample. We believe RIs to be broadly applicable to microfluidic devices as used for biological assays.     

Multistep liquid-phase lithography for fast prototyping of microfluidic free-flow-electrophoresis chips

We present a fast and versatile method to produce functional micro free-flow electrophoresis chips. Microfluidic structures were generated between two glass slides applying multistep liquid-phase lithography, omitting troublesome bonding steps or cost-intensive master structures. Utilizing a novel spacer-less approach with the photodefinable polymer polyethyleneglycol dimethacrylate (PEG-DA), microfluidic devices with hydrophilic channels of only 25 μm in height were generated. The microfluidic chips feature ion-permeable segregation walls between the electrode channels and the separation bed and hydrophilic surfaces. The performance of the chip is demonstrated by free-flow electrophoretic separation of fluorescent xanthene dyes and fluorescently labeled amino acids.     

Facile creation of hierarchical PDMS microstructures with extreme underwater superoleophobicity for anti-oil application in microfluidic channels

Composition modification and surface microstructures have been widely utilized in interface science to improve the surface performance. In this paper, we observed a significant improvement of oil contact angle (CA) from 66 ± 2° to 120 ± 4° by introducing a radical silanol group on a flat PDMS surface through oxygen plasma pretreatment. By combining surface microstructures and plasma modification, we produced three kinds of superoleophobic surfaces: 20 μm pitch micropillar arrays, 2.5 μm pitch micropillar arrays and gecko foot-like hierarchical microstructures. Among them, the hierarchical surface with high surface roughness showed extreme underwater superoleophobicity, which featured ultrahigh CA (175 ± 3°) and ultrasmall sliding angle (<1°). Quantitative measurements demonstrated that these superoleophobic surfaces exhibited distinct adhesive behaviors, by which they were interpreted as Wenzel's, Cassie's and the Lotus state, respectively. A microfluidic channel with superoleophobic microstructures was further created by novel curve-assisted imprint lithography, and the characterization based on anti-oil contamination applications was carried out and discussed. We believe that the superoleophobic surfaces will power broad applications in oil microdroplet transportation, anti-oil channels and droplet microfluidic systems.     

Probing liquid surfaces under vacuum using SEM and ToF-SIMS

We report a newly developed self-contained interface for high-vapor pressure liquid surfaces to vacuum-based analytical instruments. It requires no wires or tubing connections to the outside of the instrument and uses a microfluidic channel with a 3 μm diameter window into the flowing fluid beneath it. This window supports the liquid against the vacuum by the liquid's surface tension and limits the high-density vapor region traversed by the probe beams to only a few microns. We demonstrate this microfluidic interface for in situ liquid surfaces in a time-of-flight secondary ion mass spectrometer (ToF-SIMS) and a scanning electron microscope (SEM) with chemical analysis.     

High-volume production of single and compound emulsions in a microfluidic parallelization arrangement coupled with coaxial annular world-to-chip interfaces

This study describes a microfluidic platform with coaxial annular world-to-chip interfaces for high-throughput production of single and compound emulsion droplets, having controlled sizes and internal compositions. The production module consists of two distinct elements: a planar square chip on which many copies of a microfluidic droplet generator (MFDG) are arranged circularly, and a cubic supporting module with coaxial annular channels for supplying fluids evenly to the inlets of the mounted chip, assembled from blocks with cylinders and holes. Three-dimensional flow was simulated to evaluate the distribution of flow velocity in the coaxial multiple annular channels. By coupling a 1.5 cm × 1.5 cm microfluidic chip with parallelized 144 MFDGs and a supporting module with two annular channels, for example, we could produce simple oil-in-water (O/W) emulsion droplets having a mean diameter of 90.7 μm and a coefficient of variation (CV) of 2.2% at a throughput of 180.0 mL h(-1). Furthermore, we successfully demonstrated high-throughput production of Janus droplets, double emulsions and triple emulsions, by coupling 1.5 cm × 1.5 cm - 4.5 cm × 4.5 cm microfluidic chips with parallelized 32-128 MFDGs of various geometries and supporting modules with 3-4 annular channels.     

SERS decoding of micro gold shells moving in microfluidic systems

In this study, in situ surface‐enhanced Raman scattering (SERS) decoding was demonstrated in microfluidic chips using novel thin micro gold shells modified with Raman tags. The micro gold shells were fabricated using electroless gold plating on PMMA beads with diameter of 15 μm. These shells were sophisticatedly optimized to produce the maximum SERS intensity, which minimized the exposure time for quick and safe decoding. The shell surfaces produced well‐defined SERS spectra even at an extremely short exposure time, 1 ms, for a single micro gold shell combined with Raman tags such as 2‐naphthalenethiol and benzenethiol. The consecutive SERS spectra from a variety of combinations of Raman tags were successfully acquired from the micro gold shells moving in 25 μm deep and 75 μm wide channels on a glass microfluidic chip. The proposed functionalized micro gold shells exhibited the potential of an on‐chip microfluidic SERS decoding strategy for micro suspension array.     

Soft lithographic patterning of supported lipid bilayers onto a surface and inside microfluidic channels

We present simple soft lithographic methods for patterning supported lipid bilayer (SLB) membranes onto a surface and inside microfluidic channels. Micropatterns of polyethylene glycol (PEG)-based polymers were fabricated on glass substrates by microcontact printing or capillary moulding. The patterned PEG surfaces have shown 97 +/- 0.5% reduction in lipid adsorption onto two dimensional surfaces and 95 +/- 1.2% reduction inside microfluidic channels in comparison to glass control. Atomic force microscopy measurements indicated that the deposition of lipid vesicles led to the formation of SLB membranes by vesicle fusion due to hydrophilic interactions with the exposed substrate. Furthermore, the functionality of the patterned SLBs was tested by measuring the binding interactions between biotin (ligand)-labeled lipid bilayer and streptavidin (receptor). SLB arrays were fabricated with spatial resolution down to approximately 500 nm on flat substrate and approximately 1 microm inside microfluidic channels, respectively.     

Micro-immunohistochemistry using a microfluidic probe

A flexible method to extract more high-quality information from tissue sections is critically needed for both drug discovery and clinical pathology. Here, we present micro-immunohistochemistry (μIHC), a method for staining tissue sections at the micrometre scale. Nanolitres of antibody solutions are confined over micrometre-sized areas of tissue sections using a vertical microfluidic probe (vMFP) for their incubation with primary antibodies, the key step in conventional IHC. The vMFP operates several micrometres above the tissue section, can be interactively positioned on it, and even enables the staining of individual cores of tissue microarrays with multiple antigens. μIHC using such a microfluidic probe is preservative of tissue samples and reagents, alleviates antibody cross-reactivity issues, and allows a wide range of staining conditions to be applied on a single tissue section. This method may therefore find broad use in tissue-based diagnostics and in research.     

Separation of two phenotypically similar cell types via a single common marker in microfluidic channels

To isolate clinically and biologically relevant cell types from a heterogeneous population, fluorescent or magnetic tagging together with knowledge of surface biomarker profiles represents the state of the art. To date, it remains exceedingly difficult to separate phenotypically and physically similar cell types from a mixed population. We report a microfluidic platform engineered to separate two highly similar cell types using a single antibody by taking advantage of subtle variations in surface receptor density and cell size. This platform utilizes antibody-conjugated surfaces in microfluidic channels together with precise modulation of fluid shear stresses to accomplish selective fractionation in a continuous flow process. Antibody conjugation density variation on the adhesive surfaces is achieved by covalently immobilizing an antibody in the presence of poly(ethylene glycol). This platform is used to demonstrate separation of two CD31 positive cell types, human umbilical vein endothelial cells and human micro vascular endothelial cells.     

Modification of the glass surface property in PDMS-glass hybrid microfluidic devices

This paper presents a simple method to change the hydrophilic nature of the glass surface in a poly(dimethylsiloxane) (PDMS)-glass hybrid microfluidic device to hydrophobic by an extra-heating step during the fabrication process. Glass substrates bonded to a native or oxygen plasma-treated PDMS chip having microchambers (12.5 mm diameter, 110 μm height) were heated at 200°C for 3 h, and then the hydrophobicity of the glass surfaces on the substrate was evaluated by measuring the contact angle of water. By the extra-heating process, the glass surfaces became hydrophobic, and its contact angle was around 109°, which is nearly the same as native PDMS surfaces. To demonstrate the usefulness of this surface modification method, a PDMS-glass hybrid microfluidic device equipped with microcapillary vent structures for pneumatic manipulation of droplets was fabricated. The feasibility of the microcapillary vent structures on the device with the hydrophobic glass surfaces are confirmed in practical use through leakage tests of the vent structures and liquid handling for the electrophoretic separation of DNA molecules.     

流动聚焦型微流控体系中的液滴的图案化排列和转变模式

流体在微流通道中形成剪切流场（低雷诺数）．不同于宏观体系,由于剪切力和表面张力的竞争作用,产生的液滴在微尺度下的微流通道中形成特殊的排列现象---周期性类似“晶格”排列现象．设计了新型流动聚焦型微流控芯片,分析研究在微流体系中液滴周期性图案化排列和转变机理性,液滴排列模式受两方面因素影响：水油两相的流速比值和微通道尺寸．当微通道宽度为250或300 μm时,液滴形成单层分散,双层和单层挤压排列．当微通道宽度为350 μm 时,液滴会形成单层分散到三层排列到双层挤压最后到单层挤压排列．当出口通道宽度增加到400 μm时,甚至出现了液滴四层排列的现象．同时研究了各个液滴排列模式的“转变点”.     

Laser-induced thermal bubbles for microfluidic applications

We present a unique bubble generation technique in microfluidic chips using continuous-wave laser-induced heat and demonstrate its application by creating micro-valves and micro-pumps. In this work, efficient generation of thermal bubbles of controllable sizes has been achieved using different geometries of chromium pads immersed in various types of fluid. Effective blocking of microfluidic channels (cross-section 500 × 40 μm(2)) and direct pumping of fluid at a flow rate of 7.2-28.8 μl h(-1) with selectable direction have also been demonstrated. A particular advantage of this technique is that it allows the generation of bubbles at almost any location in the microchannel and thus enables microfluidic control at any point of interest. It can be readily integrated into lab-on-a-chip systems to improve functionality.     

A continuous-flow,microfluidic fraction collection device

A microfluidic device is presented that performs electrophoretic separation coupled with fraction collection. Effluent from the 3.5 cm separation channel was focused via two sheath flow channels into one of seven collection channels. By holding the collection channels at ground potential and varying the voltage ratio at the two sheath flow channels, the separation effluent was directed to either specific collection channels, or could be swept past all channels in a defined time period. As the sum of the voltages applied to the two sheath flow channels was constant, the electric field remained at 275 V/cm during the separation regardless of the collection channel used. The constant potential in the separation channel allowed uninterrupted separation for late-migrating peaks while early-migrating peaks were being collected. To minimize the potential for carryover between fractions, the device geometry was optimized using a three-level factorial model. The optimum conditions were a 22.5° angle between the sheath flow channels and the separation channel, and a 350 μm length of channel between the separation outlet and the fraction channels. Using these optimized dimensions, the device performance was evaluated by separation and fraction collection of a fluorescently labeled amino acid mixture. The ability to fraction collect on a microfluidic platform will be especially useful during automated or continuous operation of these devices or to collect precious samples.     

A novel crossed microfluidic device for the precise positioning of proteins and vesicles

Herein we present a novel way to create arrays of different proteins or lipid vesicles using a crossed microfluidic device. The concept relies on the combination of I) a designated two-step surface chemistry, which allows activation for subsequent binding events, and II) crossing microfluidic channels for the local functionalization by separated laminar streams. Besides its simplicity and cost efficiency, this concept has the advantage that it keeps the proteins in a hydrated environment throughout the experiment. We have demonstrated the feasibility of such a device to create a chessboard pattern of different fluorescently labeled lipid vesicles, which offers the possibility to contain biomolecules, drugs or membrane proteins.     

Micro-impedance cytometry for detection and analysis of micron-sized particles and bacteria

The sensitivity of a microfluidic impedance flow cytometer is governed by the dimensions of the sample analysis volume. A small volume gives a high sensitivity, but this can lead to practical problems including fabrication and clogging of the device. We describe a microfluidic impedance cytometer which uses an insulating fluid to hydrodynamically focus a sample stream of particles suspended in electrolyte, through a large sensing volume. The detection region consists of two pairs of electrodes fabricated within a channel 200 μm wide and 30 μm high. The focussing technique increases the sensitivity of the system without reducing the dimensions of the microfluidic channel. We demonstrate detection and discrimination of 1 μm and 2 μm diameter polystyrene beads and also Escherichia coli. Impedance data from single particles are correlated with fluorescence emission measured simultaneously. Data are also compared with conventional flow cytometry and dynamic light scattering: the coefficient of variation (CV) of size is found to be comparable between the systems.     

Polymer adsorption onto a micro-sphere from optical tweezers electrophoresis

We explore the design and operation of an optical-tweezers electrophoresis apparatus to resolve polymer adsorption dynamics onto a single micro-sphere in a micro-fluidic environment. Our model system represents a broader class of micro-fluidic electrophoresis experiments for biosensing and fundamental colloid and surface science diagnostics. We track the adsorption of 100 kDa poly(ethylene oxide) homopolymer onto a colloidal silica sphere that is optically trapped in a crossed parallel-plate micro-channel. The adsorption dynamics are probed on the ～1 μm particle length scale with ～1 s temporal resolution. Because the particle electrophoretic mobility and channel electro-osmotic flow are exquisitely sensitive to the polymer layer hydrodynamic thickness, particle dynamics can be complicated by polymer adsorption onto the micro-channel walls. Nevertheless, using experiments and a theoretical model of electro-osmotic flow in channels with non-uniform wall ζ-potentials, we show that such influences can be mitigated by adopting a symmetrical flow configuration. The equilibrium hydrodynamic layer thickness of 100 kDa poly(ethylene oxide) on colloidal silica is ～10 nm at polymer concentrations ?10 ppm (weight percent), with the dynamics reflecting polymer solution concentration, flow rate, and polydispersity.     

Integrated three-dimensional filter separates nanoscale from microscale elements in a microfluidic chip

We report on the integration of a size-based three-dimensional filter, with micrometre-sized pores, in a commercial microfluidic chip. The filter is fabricated inside an already sealed microfluidic channel using the unique capabilities of two-photon polymerization. This direct-write technique enables integration of the filter by post-processing in a chip that has been fabricated by standard technologies. The filter is located at the intersection of two channels in order to control the amount of flow passing through the filter. Tests with a suspension of 3 μm polystyrene spheres in a Rhodamine 6G solution show that 100% of the spheres are stopped, while the fluorescent molecules are transmitted through the filter. We demonstrate operation up to a period of 25 minutes without any evidence of clogging. Preliminary validation of the device for plasma separation from whole blood is shown. Moreover, the filter can be cleaned and reused by reversing the flow.     

Fabrication of circular microfluidic channels by combining mechanical micromilling and soft lithography

The fabrication of microfluidic channels with complex three-dimensional (3D) geometries presents a major challenge to the field of microfluidics, because conventional lithography methods are mainly limited to rectangular cross-sections. In this paper, we demonstrate the use of mechanical micromachining to fabricate microfluidic channels with complex cross-sectional geometries. Micro-scale milling tools are first used to fabricate semi-circular patterns on planar metallic surfaces to create a master mold. The micromilled pattern is then transferred to polydimethylsiloxane (PDMS) through a two-step reverse molding process. Using these semi-circular PDMS channels, circular cross-sectioned microchannels are created by aligning and adhering two channels face-to-face. Straight and serpentine-shaped microchannels were fabricated, and the channel geometry and precision of the metallic master and PDMS molds were assessed through scanning electron microscopy and non-contact profilometry. Channel functionality was tested by perfusion of liquid through the channels. This work demonstrates that micromachining enabled soft lithography is capable of fabricating non-rectangular cross-section channels for microfluidic applications. We believe that this approach will be important for many fields from biomimetics and vascular engineering to microfabrication and microreactor technologies.