A passive microfluidic device for continuous microparticle enrichment

A passive microfluidic device is reported for continuous microparticle enrichment. The microparticle is enriched based on the inertial effect in a microchannel with contracting‐expanding structures on one side where microparticles/cells are subjected to the inertial lift force and the momentum‐change‐induced inertial force induced by highly curved streamlines. Under the combined effect of the two forces, yeast cells and microparticles of different sizes were continuously focused in the present device over a range of Reynolds numbers from 16.7 to 125. ～68% of the particle‐free liquid was separated from the sample at Re = 66.7, and ～18 μL particle‐free liquid was fast obtained within 10 s. Results also showed that the geometry of the contracting‐expanding structure significantly influenced the lateral migration of the particle. Structures with a large angle induced strong inertial effect and weak disturbance effect of vortex on the particle, both of which enhanced the microparticle enrichment in microchannel. With simple structure, small footprint (18 × 0.35 mm), easy operation and cell‐friendly property, the present device has great potential in biomedical applications, such as the enrichment of cells and the fast extraction of plasma from blood for disease diagnose and therapy.     

A micropillar-integrated smart microfluidic device for specific capture and sorting of cells

An integrated smart microfluidic device consisting of nickel micropillars, microvalves, and microchannels was developed for specific capture and sorting of cells. A regular hexagonal array of nickel micropillars was integrated on the bottom of a microchannel by standard photolithography, which can generate strong induced magnetic field gradients under an external magnetic field to efficiently trap superparamagnetic beads (SPMBs) in a flowing stream, forming a bed with sufficient magnetic beads as a capture zone. Fluids could be manipulated by programmed controlling the integrated air-pressure-actuated microvalves, based on which in situ bio-functionalization of SPMBs trapped in the capture zone was realized by covalent attachment of specific proteins directly to their surface on the integrated microfluidic device. In this case, only small volumes of protein solutions (62.5 nL in the capture zone; 375 nL in total volume needed to fill the device from inlet A to the intersection of outlet channels F and G) can meet the need for protein! The newly designed microfluidic device reduced greatly chemical and biological reagent consumption and simplified drastically tedious manual handling. Based on the specific interaction between wheat germ agglutinin (WGA) and N-acetylglucosamine on the cell membrane, A549 cancer cells were effectively captured and sorted on the microfluidic device. Capture efficiency ranged from 62 to 74%. The integrated microfluidic device provides a reliable technique for cell sorting.     

A microfluidic device for continuous, real time blood plasma separation

A microfluidic device for continuous, real time blood plasma separation is introduced. The principle of the blood plasma separation from blood cells is supported by the Zweifach-Fung effect and was experimentally demonstrated using simple microchannels. The blood plasma separation device is composed of a blood inlet, a bifurcating region which leads to a purified plasma outlet, and a concentrated blood cell outlet. It was designed to separate blood plasma from an initial blood sample of up to 45% inlet hematocrit (volume percentage of cells). The microfluidic network was designed using an analogous electrical circuit, as well as analytical and numerical studies. The functionality of this device was demonstrated using defibrinated sheep blood. During 30 minutes of continuous blood infusion through the device, all the erythrocytes (red blood cells) traveled through the device toward the concentrated blood outlet while only the plasma was separated at the bifurcating regions and flowed towards the plasma outlet. The device has been operated continuously without any clogging or hemolysis of cells. The experimentally determined plasma selectivity with respect to blood hematocrit level was almost 100% regardless of the inlet hematocrit. The total plasma separation volume percent varied from 15% to 25% with increasing inlet hematocrit. Due to the device's simple structure and control mechanism, this microdevice is expected to be used for highly efficient continuous, real time cell-free blood plasma separation from blood samples for use in lab on a chip applications.     

A self-loading microfluidic device for determining the minimum inhibitory concentration of antibiotics

This article describes a portable microfluidic technology for determining the minimum inhibitory concentration (MIC) of antibiotics against bacteria. The microfluidic platform consists of a set of chambers molded in poly(dimethylsiloxane) (PDMS) that are preloaded with antibiotic, dried, and reversibly sealed to a second layer of PDMS containing channels that connect the chambers. The assembled device is degassed via vacuum prior to its use, and the absorption of gas by PDMS provides the mechanism for actuating and metering the flow of fluid in the microfluidic channels and chambers. During the operation of the device, degas driven flow introduces a suspension of bacterial cells, dissolves the antibiotic, and isolates cells in individual chambers without cross contamination. The growth of bacteria in the chambers in the presence of a pH indicator produces a colorimetric change that can be detected visually using ambient light. Using this device we measured the MIC of vancomycin, tetracycline, and kanamycin against Enterococcus faecalis 1131, Proteus mirabilis HI4320, Klebsiella pneumoniae, and Escherichia coli MG1655 and report values that are comparable to standard liquid broth dilution measurements. The device provides a simple method for MIC determination of individual antibiotics against human pathogens that will have applications for clinical and point-of-care medicine. Importantly, this device is designed around simplicity: it requires a single pipetting step to introduce the sample, no additional components or external equipment for its operation, and provides a straightforward visual measurement of cell growth. As the device introduces a novel approach for filling and isolating dead-end microfluidic chambers that does not require valves and actuators, this technology should find applications in other portable assays and devices.     

A simple, disposable microfluidic device for rapid protein concentration and purification via direct-printing

A facile and disposable microfluidic device for rapid protein concentration was fabricated by using a direct printing process. Two printed V-shaped microchannels in mirror image orientation were separated by a 100 mum wide toner gap. When a high electric field was applied across the two channels, nanofissures were formed by electric breakdown at the junction toner gap. This microfluidic device with nanofissures was used as a concentrator for protein. Negatively charged proteins were observed to concentrate at the anode side of the nanofissures upon application of an electric field across this junction. Using this device, about 10(3)-10(5)-fold protein concentration was achieved within 10 min. Systematic investigation showed that the concentration mechanism could be explained by the ion exclusion-enrichment effect of the nanofissures. In addition, the present microchip device integrated both functions of concentration and purification were confirmed. This simple on chip protein preconcentration and purification device could be a disposable sample preparation component in printed microfluidic systems used for practical biochemical assays.     

A microfluidic device for separating erythrocytes polluted by lead (II) from a continuous bloodstream flow

To sort and separate erythrocytes contaminated by lead (II) from whole bloodstream flow, the first step is to use a microchannel to transport the blood cells into a microdevice. Within the device, polluted erythrocytes can be separated from the bloodstream by applying local dielectrophoretic (DEP) forces. Exploiting the fact that Pb(2+) ions attach to the membranes of the erythrocytes, we utilize the microfluidic DEP device to perform property-based fractionation of the blood samples and to separate the polluted erythrocytes from the continuous bloodstream flow. Atomic absorption spectrometer analysis reveals that, to remove lead-polluted erythrocytes, the most effective driving velocity was less than 0.1 cm/s through our microfluidic DEP device, based on an applied power of 10 V(peak-peak) and a frequency of 15.5 MHz AC field. We were able to remove 80% of the polluted erythrocytes. Using gentle DEP manipulating techniques to efficiently sort unique cells within a complex biological sample may potentially allow biological sorting to be performed outside of hospitals, in facilities without biological analyzing equipment.     

Specific capture and temperature-mediated release of cells in an aptamer-based microfluidic device

Isolation of cells from heterogeneous mixtures is critically important in both basic cell biology studies and clinical diagnostics. Cell isolation can be realized based on physical properties such as size, density and electrical properties. Alternatively, affinity binding of target cells by surface-immobilized ligands, such as antibodies, can be used to achieve specific cell isolation. Microfluidics technology has recently been used in conjunction with antibody-based affinity isolation methods to capture, purify and isolate cells with higher yield rates, better efficiencies and lower costs. However, a method that allows easy release and collection of live cells from affinity surfaces for subsequent analysis and detection has yet to be developed. This paper presents a microfluidic device that not only achieves specific affinity capture and enrichment, but also enables non-destructive, temperature-mediated release and retrieval of cells. Specific cell capture is achieved using surface-immobilized aptamers in a microchamber. Release of the captured cells is realized by a moderate temperature change, effected via integrated heaters and a temperature sensor, to reversibly disrupt the cell-aptamer interaction. Experimental results with CCRF-CEM cells have demonstrated that the device is capable of specific capture and temperature-mediated release of cells, that the released cells remain viable and that the aptamer-functionalized surface is regenerable.     

Novel microfluidic device for the continuous separation of cancer cells using dielectrophoresis

We describe the design, microfabrication, and testing of a microfluidic device for the separation of cancer cells based on dielectrophoresis. Cancer cells, specifically green fluorescent protein‐labeled MDA‐MB‐231, are successfully separated from a heterogeneous mixture of the same and normal blood cells. MDA‐MB‐231 cancer cells are separated with an accuracy that enables precise detection and counting of circulating tumor cells present among normal blood cells. The separation is performed using a set of planar interdigitated transducer electrodes that are deposited on the surface of a glass wafer and slightly protrude into the separation microchannel at one side. The device includes two parts, namely, a glass wafer and polydimethylsiloxane element. The device is fabricated using standard microfabrication techniques. All experiments are conducted with low conductivity sucrose‐dextrose isotonic medium. The variation in response between MDA‐MB‐231 cancer cells and normal cells to a certain band of alternating‐current frequencies is used for continuous separation of cells. The fabrication of the microfluidic device, preparation of cells and medium, and flow conditions are detailed. The proposed microdevice can be used to detect and separate malignant cells from heterogeneous mixture of cells for the purpose of early screening for cancer.     

Simple host-guest chemistry to modulate the process of concentration and crystallization of membrane proteins by detergent capture in a microfluidic device

This paper utilizes cyclodextrin-based host-guest chemistry in a microfluidic device to modulate the crystallization of membrane proteins and the process of concentration of membrane protein samples. Methyl-beta-cyclodextrin (MBCD) can efficiently capture a wide variety of detergents commonly used for the stabilization of membrane proteins by sequestering detergent monomers. Reaction Center (RC) from Blastochloris viridis was used here as a model system. In the process of concentrating membrane protein samples, MBCD was shown to break up free detergent micelles and prevent them from being concentrated. The addition of an optimal amount of MBCD to the RC sample captured loosely bound detergent from the protein-detergent complex and improved sample homogeneity, as characterized by dynamic light scattering. Using plug-based microfluidics, RC crystals were grown in the presence of MBCD, giving a different morphology and space group than crystals grown without MBCD. The crystal structure of RC crystallized in the presence of MBCD was consistent with the changes in packing and crystal contacts hypothesized for removal of loosely bound detergent. The incorporation of MBCD into a plug-based microfluidic crystallization method allows efficient use of limited membrane protein sample by reducing the amount of protein required and combining sparse matrix screening and optimization in one experiment. The use of MBCD for detergent capture can be expanded to develop cyclodextrin-derived molecules for fine-tuned detergent capture and thus modulate membrane protein crystallization in an even more controllable way.     

A polymer-based microfluidic device for immunosensing biochips

This paper describes the design, fabrication, and test of a PDMS/PMMA-laminated microfluidic device for an immunosensing biochip. A poly(dimethyl siloxane)(PDMS) top substrate molded by polymer casting and a poly(methyl methacrylate)(PMMA) bottom substrate fabricated by hot embossing are bonded with pressure and hermetically sealed. Two inlet ports and an air vent are opened through the PDMS top substrate, while gold electrodes for electrochemical biosensing are patterned onto the PMMA bottom substrate. The analyte sample is loaded from the sample inlet port to the detection chamber by capillary force, without any external intervening forces. For this and to control the time duration of sample fluid in each compartment of the device, including the inlet port, diffusion barrier, reaction chamber, flow-delay neck, and detection chamber, the fluid conduit has been designed with various geometries of channel width, depth, and shape. Especially, the fluid path has been designed so that the sample flow naturally stops after filling the detection chamber to allow sufficient time for biochemical reaction and subsequent washing steps. As model immunosensing tests for the microfluidic device, functionalizations of ferritin and biotin to the sensing surfaces on gold electrodes and their biospecific interactions with antiferritin antiserum and streptavidin have been investigated. An electrochemical detection method for immunosensing by biocatalyzed precipitation has been developed and applied for signal registration. With the biochip, the whole immunosensing processes could be completed within 30 min.     

A microfluidic electroporation device for cell lysis

We demonstrate a micro-electroporation device for cell lysis prior to subcellular analysis. Simple circuit models show that electrical lysis method is advantageous because it is selective towards plasma membrane while leaving organelle membrane undamaged. In addition, miniaturization of this concept leads to negligible heat generation and bubble formation. The designed microdevices were fabricated using a combination of photolithography, metal-film deposition, and electroplating. We demonstrate the electro-lysis of human carcinoma cells in these devices to release the subcellular materials.     

A software-programmable microfluidic device for automated biology

Specific-purpose microfluidic devices have had considerable impact on the biological and chemical sciences, yet their use has largely remained limited to specialized laboratories. Here we present a general-purpose software-programmable microfluidic device which is capable of performing a multitude of low- and high-level functions without requiring any hardware modifications. To demonstrate the applicability and modularity of the device we implemented a variety of applications such as a microfluidic display, fluid metering and active mixing, surface immunoassays, and cell culture. We believe that analogously to personal computers, programmable, general-purpose devices will increase the accessibility and advance the pervasiveness of microfluidic technology.     

A microfluidic device for performing pressure-driven separations

Microchannels in microfluidic devices are frequently chemically modified to introduce specific functional elements or operational modalities. In this work, we describe a miniaturized hydraulic pump created by coating selective channels in a glass microfluidic manifold with a polyelectrolyte multilayer (PEM) that alters the surface charge of the substrate. Pressure-driven flow is generated due to a mismatch in the electroosmotic flow (EOF) rates induced upon the application of an electric field to a tee channel junction that has one arm coated with a positively charged PEM and the other arm left uncoated in its native state. In this design, the channels that generate the hydraulic pressure are interconnected via the third arm of the tee to a field-free analysis channel for performing pressure-driven separations. We have also shown that modifications in the cross-sectional area of the channels in the pumping unit can enhance the hydrodynamic flow through the separation section of the manifold. The integrated device has been demonstrated by separating Coumarin dyes in the field-free analysis channel using open-channel liquid chromatography under pressure-driven flow conditions.     

Characterization of particle capture in a sawtooth patterned insulating electrokinetic microfluidic device

Here we present a scheme to separate particles according to their characteristic physical properties, including size, charge, polarizability, deformability, surface charge mobility, dielectric features, and local capacitance. Separation is accomplished using a microdevice based on direct current insulator gradient dielectrophoresis that can isolate and concentrate multiple analytes simultaneously at different positions. The device is dependent upon dielectrophoretic and electrokinetic forces incorporating a global longitudinal direct current field as well as using shaped insulating features within the channel to induce local gradients. This design allows for the production of strong local field gradients along a global field causing particles to enter, initially transported through the channel by electrophoresis and electroosmosis (electrokinetics), and to be isolated via repulsive dielectrophoretic forces that are proportional to an exponent of the field gradient. Sulfate-capped polystyrene nano and microparticles (20, 200 nm, and 1 μm) were used as probes to demonstrate the influence of channel geometry and applied longitudinal field on separation behavior. These results are consistent with models using similar channel geometry and indicate that specific particulate species can be isolated within a distinct portion of the device, whereas concentrating particles by factors from 10(3) to 10(6).     

A plug and play microfluidic device

Chip-to-world interface is a major issue in the field of microfluidics and its applications. We developed a plug and play microfluidic device composed of a fluid driving unit and a polymer chip containing microfluidic channels and reservoirs. The one and only connection of the device to the external world is a set of electric control lines for the driving unit. Just putting the reagents and samples onto the reservoirs, the chip can be operated for chemical or biochemical reaction and analysis. We demonstrate here that silicon-based micropumps embedded in the present device allow us to achieve flexible fluidic manipulations with minimum time delay and dead volume.     

A microfluidic device for self-synchronised production of droplets

The primary requirement for a mixing operation in droplet-based microfluidic devices is an accurate pairing of droplets of reaction fluids over an extended period of time. In this paper, a novel device for self-synchronous production of droplets has been demonstrated. The device uses a change in impedance across a pair of electrodes introduced due to the passage of a pre-formed droplet to generate a second droplet at a second pair of electrodes. The device was characterised using image analysis. Droplets with a volume of ~23.5 ± 3.1 nl (i.e.~93% of the volume of pre-formed droplets) were produced on applying a voltage of 500 V. The synchronisation efficiency of the device was 83%. As the device enables self-synchronised production of droplets, it has a potential to increase the reliability and robustness of mixing operations in droplet-based microfluidic devices.     

A microfluidic device for accurate detection of hs-cTnI

This device is aimed at ensuring that the sample is uniformly and equivalently reacted with the antibody on the NC membrane in each test when the microfluidic liquid system is introduced to the chip. In this study, the developed microfluidic chip can avoid the presence of the sample and conjugate pads in the chip, while the precision of the chromatography system can be greatly improved using the same particles, NC membrane and antibody alongside the traditional strip. The results, taking the detection of cTnI as an example, revealed that the coefficient of variation (CV) is controlled within 4%, while the maximum record of the contrast chromatographic reagent strip can reach 15%. Additionally, the detection sensitivity can maintain the same order of magnitudes with that of the traditional chromatographic strip. With the results, the determination correlation of the developed microfluidic chip has been greatly improved. In addition, the CV of the chip in this study is greatly improved in comparison with that of the traditional strip. The biggest improvement lies in the mixing between the sample and the microspheres, indicating that this is a new approach to improve the CV of the traditional strip.     

A magnetically active microfluidic device for chemiluminescence bioassays

Highly active horseradish peroxidase functionalized magnetic nanoparticles were prepared and packed into a microfluidic channel, producing an in-line bioreactor that enabled a sensitive chemiluminescence assay of H(2)O(2). The proposed magnetically active microfluidic device proved useful for chemiluminescence assays of biomedically interesting compounds.     

Locally enhanced concentration and detection of oligonucleotides in a plug-based microfluidic device

We propose a novel technique that allows oligonucleotides with specific end-modification within a plug in a plug-based microfluidic device to undergo a locally enhanced concentration at the rear of the plug as the plug moves downstream. DNA was enriched and detected in situ upon exploiting a combined effect underlain by an entropic force induced through fluid shear (i.e. a hydrodynamic-repellent effect) and the interfacial adsorption (aqueous/oil interface) attributed to affinity. Flow fields within a plug were visualized quantitatively using micro-particle image velocimetry (micro-PIV); the distribution of the fluid shear strain rate explains how the hydrodynamic-repellent effect engenders a dumbbell-like region with an increased concentration of DNA. The concentration of FAM (6-carboxy-fluorescein)-labeled DNA (FC-DNA) and of TAMRA (tetramethyl-6-carboxyrhodamine)-labeled DNA (TC-DNA), respectively, and the hybridization of probe DNA (modified with FAM) with target DNA (modified with TAMRA) were investigated in devices; a confocal fluorescence microscope (CFM) was utilized to monitor the processes and to resolve the corresponding 2D patterns and 3D reconstruction of the DNA distribution in a plug. TC-DNA, but not FC-DNA, concentrating within a plug was affected by the combined effect so as to achieve a concentration factor (C(r)) twice that of FC-DNA because of the lipophilicity of TAMRA. Using fluorescence resonance-energy transfer (FRET), we characterized the hybridization of the DNA in a plug; the detection limit of a system, improved by virtue of the proposed technique (the locally enhanced concentration), for DNA detection was estimated to be 20-50 nM. This technique enables DNA to concentrate locally in a nL-pL free-solution plug, the locally enhanced concentration to profit the hybridization efficiency and the detection of DNA, prospectively serving as a versatile means to accomplish a rapid DNA detection in a small volume for a Lab-on-a-Chip (LOC) system.     

A linear dilution microfluidic device for cytotoxicity assays

A two-layer polymer microfluidic device is presented which creates nine linear dilutions from two input fluid streams mixed in varying volumetric proportions. The linearity of the nine dilutions is conserved when the flow rate is held constant at 1.0 microl min(-1) (R(2) = 0.9995) and when it is varied from 0.5-16 microl min(-1) (R(2) = 0.9998). An analytical expression is presented for designing microfluidic devices with arbitrary numbers of linear dilutions. To demonstrate the efficacy of this device, primary human epidermal keratinocytes (HEK) were stained with nine dilutions of calcein, resulting in a linear spread of fluorescent intensities (R(2) = 0.94). The operating principles of the device can be scaled up to incorporate any number of linear dilutions. This scalability, coupled with an intrinsic ability to create linear dilutions under a variety of operating conditions, makes the device applicable to high throughput screening applications such as combinatorial chemistry or cytotoxicity assays.