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At least 415 records · Page 23Linked to original sources

PubMed2006

Phase-changing sacrificial materials for interfacing microfluidics with ion-permeable membranes to create on-chip preconcentrators and electric field gradient focusing microchips.

We have developed a novel approach for interfacing ionically conductive membranes with microfluidic systems using phase-changing sacrificial layers. Imprinted microchannels in a polymer substrate are filled with a heated liquid that solidifies at room temperature, a monomer solution is placed over the protected channels and polymerized to form a rigid semipermeable copolymer, and then the protective layer is melted and removed, leaving an open microchannel interfaced with a polymer membrane. We have applied this method in miniaturizing electric field gradient focusing (EFGF) and carrying out on-chip protein preconcentration. A semipermeable copolymer in the EFGF microchips fills a region of changing cross-sectional area, which allows a gradient in electric field to be established when an electrical potential is applied. Our technique provides microchip EFGF devices that offer 3-fold improved resolution in protein focusing compared with capillary-based systems. In addition, these EFGF microchips can separate peptide samples with resolution similar to what is obtained in capillary electrophoresis microdevices, and the micro-EFGF systems enrich analytes by a factor of >150. Finally, we have fabricated membrane-integrated microfluidic devices that can concentrate protein samples (R-phycoerythrin) over 10 000-fold to facilitate microchip capillary electrophoresis. Interfacing microchannels with ion-permeable membranes has great potential to enhance microchip analysis of biomolecules.

Electric Conductivity↗

PubMed2006

Continuous-flow pI-based sorting of proteins and peptides in a microfluidic chip using diffusion potential.

Efficient sample preparation tools are the key to measuring molecular signals in a complex biological system. A novel continuous-flow isoelectric point (pI)-based sorting technique has been developed for proteins and peptides in a microfluidic chip format. It can sort biomolecules at a relatively high flow rate of up to 10 microL/min and does not require carrier ampholytes, which can create molecular backgrounds for subsequent analysis. Furthermore, the electrophoretic field required to run the pI-based sorting is generated by the diffusion of buffer ions in situ, at the liquid junction between two laminar flows within the microfluidic channel. Utilizing the diffusion potential in combination with a pH difference between the buffers, we demonstrated a separation of binary mixtures of pI markers and proteins without applying any external field. The sorting resolution and its efficiency are sufficiently high for sample preparation and could be further improved by optimizing buffers or with an additional transverse electric field. Once fully developed, it can potentially be a pI-based sample fractionation tool for proteomic analysis of complex biomolecule samples.

Diffusion↗

PubMed2006

Construction of a biomimetic surface on microfluidic chips for biofouling resistance.

A biomimetic surface has been formed on the poly(methyl methacrylate) (PMMA) microfluidic chips for biofouling resistance on the basis of a simple modification. Accordingly, an amphiphilic phospholipid copolymer of 2-methacryloyloxyethyl phosphorylcholine and n-butyl methacrylate (PMB) was developed to introduce the phosphorylcholine functional groups onto the PMMA surface via the anchoring of hydrophobic n-butyl methacrylate units. The 2-methacryloyloxyethyl phosphorylcholine segments could form hydrophilic domains, considered to be located on the surface, to provide a biocompatible surface. X-ray photoelectron spectroscopy and Fourier transform infrared spectra confirmed the success of surface functionalization. The PMB-modified microchips containing phosphorylcholine moieties exhibited more stable electroosmotic mobility compared with the untreated one. In addition to being characterized for minimized nonspecific adhesion of serum proteins and plasma platelets, the PMB-functionalized microchannels have been exemplified by electrophoresis of proteins. This one-step procedure offers an effective approach for a biomimetic surface design on microfluidic chips, which is promising in high-throughput and complex biological analysis.

Biomimetics↗

PubMed2005

Surface-reactive acrylic copolymer for fabrication of microfluidic devices.

A surface-reactive acrylic polymer, poly(glycidyl methacrylate-co-methyl methacrylate) (PGMAMMA), was synthesized and evaluated for suitability as a substrate for fabrication of microfluidic devices for chemical analysis. This polymer has good thermal and optical properties and is mechanically robust for cutting and hot embossing. A key advantage of this polymeric material is that the surface can be easily modified to control inertness and electroosmotic flow using a variety of chemical procedures. In this work, the procedures for aminolysis, photografting of linear polyacrylamide, and atom-transfer radical polymerization on microchannel surfaces in PGMAMMA substrates were developed, and the performance of resultant microfluidic electrophoresis devices was demonstrated for the separation of amino acids, peptides, and proteins. Separation efficiencies as high as 4.6 x 10(4) plates for a 3.5-cm-long separation channel were obtained. The results indicate that PGMAMMA is an excellent substrate for microfabricated fluidic devices, and a broad range of applications should be possible.

Amines↗

PubMed2006

Microfluidic isolation of leukocytes from whole blood for phenotype and gene expression analysis.

Technologies that enable the isolation of cell subtypes from small samples of complex populations will greatly facilitate the implementation of proteomics and genomics to human diseases. Transcriptome analysis of blood requires the depletion of contaminating erythrocytes. We report an automated microfluidic device to rapidly deplete erythrocytes from whole blood via deionized water lysis and to collect enriched leukocytes for phenotype and genomic analyses. Starting with blood from healthy subjects, we demonstrate the utility of this microfluidic cassette and lysis protocol to prepare unstimulated leukocytes, and leukocytes stimulated ex vivo with Staphylococcal enterotoxin B, which mimics some of the cellular effects seen in patients with severe bacterial infections. Microarrays are used to assess the global gene expression response to enterotoxin B. The results demonstrate that this system can isolate unactivated leukocytes from small blood samples without any significant loss, which permits more information to be obtained from subsequent analysis, and will be readily applicable to clinical settings.

Cell Separation↗

PubMed2006

Size exclusion chromatography of microliter volumes for on-line use in low-pressure microfluidic systems.

We have developed a new cartridge format for on-line size exclusion processing in low-pressure, portable microfluidic devices. The described system allows size exclusion chromatography of microliter volumes (termed muL-SEC) to be performed with ready integration in complex protocols for continuous-flow sample processing and analysis. The refillable cartridge format was employed for the preparation of Bacillus subtilus spores lysed in the presence of a strong reducing agent. While the reducing agent is known to interfere with subsequent fluorescent labeling of the solubilized proteins, the described continuous-flow size exclusion processing allowed complete isolation of interferences from a 10-muL sample in 70 s. Following efficient labeling, the protein sample was injected and separated on-chip using gel electrophoresis. To increase the resolution, speed, and sample capacity of buffer exchange under low-pressure operation (40 psi), parameters such as the size exclusion resin, load volumes, flow rates, buffer composition, and cartridge geometry were optimized and are presented here. The muL-SEC analysis is compatible with automated sample preparation for microfluidic systems and has resulted in significantly increased analysis speed and throughput over benchtop methods. The presented technique has the potential to improve capabilities such as buffer exchange, size fractionation, and high-abundance protein removal-steps that are frequently required prior to on-chip, point-of-care, and mass spectrometric analyses.

Bacillus subtilis↗

PubMed2006

Solution-phase surface modification in intact poly(dimethylsiloxane) microfluidic channels.

An improved approach composed of an oxidation reaction in acidic H2O2 solution and a sequential silanization reaction using neat silane reagents for surface modification of poly(dimethylsiloxane) (PDMS) substrates was developed. This solution-phase approach is simple and convenient for some routine analytical applications in chemistry and biology laboratories and is designed for intact PDMS-based microfluidic devices, with no device postassembly required. Using this improved approach, two different functional groups, poly(ethylene glycol) (PEG) and amine (NH2), were introduced onto PDMS surfaces for passivation of nonspecific protein absorption and attachment of biomolecules, respectively. X-ray electron spectroscopy and temporal contact angle experiments were employed to monitor functional group transformation and dynamic characteristics of the PEG-grafted PDMS substrates; fluorescent protein solutions were introduced into the PEG-grafted PDMS microchannels to test their protein repelling characteristics. These analytical data indicate that the PEG-grafted PDMS surfaces exhibit improved short-term surface dynamics and robust long-term stability. The amino-grafted PDMS microchannels are also relatively stable and can be further activated for modifications with peptide, DNA, and protein on the surfaces of microfluidic channels. The resulting biomolecule-grafted PDMS microchannels can be utilized for cell immobilization and incubation, semiquantitative DNA hybridization, and immunoassay.

Amines↗

PubMed2006

Lipid bilayer formation by contacting monolayers in a microfluidic device for membrane protein analysis.

Artificial planar lipid bilayers are a powerful tool for the functional study of membrane proteins, yet they have not been widely used due to their low stability and reproducibility. This paper describes an accessible method to form a planar lipid bilayer, simply by contacting two monolayers assembled at the interface between water and organic solvent in a microfluidic chip. The membrane of an organic solvent containing phospholipids at the interface was confirmed to be a bilayer by the capacitance measurement and by measuring the ion channel signal from reconstituted antibiotic peptides. We present two different designs for bilayer formation. One equips two circular wells connected, in which the water/solvent/water interface was formed by simply injecting a water droplet into each well. Another equips the cross-shaped microfluidic channel. In the latter design, formation of the interface at the sectional area was controlled by external syringe pumps. Both methods are extremely simple and reproducible, especially in microdevices, and will lead to automation and multiple bilayer formation for the high-throughput screening of membrane transport in physiological and pharmaceutical studies.

Automation↗

PubMed2005

Miniaturized lead sensor based on lead-specific DNAzyme in a nanocapillary interconnected microfluidic device.

A miniaturized lead sensor has been developed by combining a lead-specific DNAzyme with a microfabricated device containing a network of microfluidic channels that are fluidically coupled via a nanocapillary array interconnect. A DNAzyme construct, selective for cleavage in the presence of Pb2+ and derivatized with fluorophore (quencher) at the 5' (3') end of the substrate and enzyme strands, respectively, forms a molecular beacon that is used as the recognition element. The nanocapillary array membrane interconnect is used to manipulate fluid flows and deliver the small-volume sample to the beacon in a spatially confined detection window where the DNAzyme is interrogated using laser-induced fluorescence detection. A transformed log plot of the fluorescent signal exhibits a linear response (r2 = 0.982) over a Pb2+ concentration range of 0.1 - 100 microM, and a detection limit of 11 nM. The sensor has been applied to the determination of Pb2+ in an electroplating sludge reference material, the result agreeing with the certified value within 4.9%. Quantitative measurement of Pb2+ in this complex sample demonstrates the selectivity of this sensor scheme and points favorably to the application of such technologies to analysis of environmental samples. The unique combination of a DNAzyme with a microfluidic-nanofluidic hybrid device makes it possible to change the DNAzyme to select for other compounds of interest, and to incorporate multiple sensing systems within a single device for greater flexibility. This work represents the initial steps toward creation of a robust field sensor for lead in groundwater or drinking water.

Biosensing Techniques↗

PubMed2004

Controlled vesicle self-assembly in microfluidic channels with hydrodynamic focusing.

Traditional liposome preparation methods are based on mixing of bulk phases, leading to inhomogeneous chemical and/or mechanical conditions during formation; hence liposomes are often polydisperse in size and lamellarity. Here we show the formation of liposomes that encapsulate reagents in a continuous two-phase flow microfluidic network with precision control of size from 100 to 300 nm by manipulation of liquid flow rates. We demonstrate that by creating a solvent-aqueous interfacial region in a microfluidic format that is homogeneous and controllable on the length scale of a liposome, we can facilitate the fine control of liposome size and polydispersity.

Fluoresceins↗

PubMed2005

A microfluidic biomaterial.

We report on the incorporation of microfluidic structure within a high-water-content hydrogel [4% (w/v) calcium alginate]. We used the microfluidic network to control the chemical environment within the hydrogel and demonstrated higher rates of delivery and extraction of solutes than was achievable by diffusion alone.

Alginates↗

PubMed2006

Controlled microfluidic encapsulation of cells, proteins, and microbeads in lipid vesicles.

Cells have been encapsulated inside lipid vesicles by using a new microfluidic lipid vesicle formulation technique. Lipid vesicles are formulated within minutes without using toxic lipid solvents. The encapsulation efficiency inside the vesicles is controlled by the microfluidic flows. Green fluorescent proteins (GFP), carcinoma cells, and bead encapsulated vesicles have mean diameters of 27.2 mum, 62.4 mum, and 55.9 mum, respectively. The variations of vesicle sizes are approximately 20% for the GFP and cell encapsulated vesicles and approximately 10% for the bead encapsulated vesicles.

HeLa Cells↗

PubMed2006

Interfacial thermocapillary vortical flow for microfluidic mixing.

We present a method for the mixing of fluids in a quasi two-dimensional system with low Reynolds number by means of generating a vortical flow. A two-dimensional cavitation bubble is induced in liquid-expanded phase by locally heating a Langmuir monolayer at the air/liquid interface with an IR laser. The laser-induced cavitation bubble works as a microfluidic pump and generates a thermocapillary flow around the pump. As a result, the surrounding liquid-expanded phase flows in one direction. Perturbing the thermocapillary flow with solid folds that are created by compression and reexpansion of the monolayer induces the vortical flow behind the folds. Applying the equation of creeping flow, we find a torque halfway from the center causing the vortical flow. The vorticity created in this way stretches the liquid-expanded and gaseous phase in the azimuthal direction and at the same time thins both phases in the radial direction. If the vortical flow could be maintained long enough to reach a radial thinning that would allow the interdiffusion of surfactants at the surface, then this technique would open a route for the effective two-dimensional microfluidic mixing at low Reynolds numbers.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2006

Generation of complex, static solution gradients in microfluidic channels.

A microfluidic device is used to generate a complex gradient of diffusible molecules in a static solution. The gradient is precise and steady both in space and in time. This device, made from poly(dimethylsiloxane), consists of three layers. The molecules in reservoirs on the top layer diffuse through the flat middle layer of hydrogel and reach an equilibrium distribution. Microfluidic channels on the bottom layer that are in close contact with the hydrogel contain free solution that has concentration gradients based on the gradient in the gel. The gradient profile in the channel can be designed to have an arbitrary form (within the range of the existing gradient in the hydrogel) by controlling the local direction of the channel at each point.

Dimethylpolysiloxanes↗

PubMed2025

Unlocking Zeptomolar Single-Molecule Detection by Synergizing Digital Microfluidics and Digital CRISPR.

Accurate diagnosis relies on the highly sensitive and quantitative detection of multiple immune-related biomarkers. However, current detection methods still face significant limitations in sensitivity, specificity, and background signal control. Here, we introduce DDA (Dual-Digital immunoAssay), a fully automated, universal immunoassay platform that synergizes digital microfluidics with digital Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-based amplification. This "dual-digital" strategy pushes the detection limit into the zeptomolar (zM) regime, enabling unprecedented sensitivity for single-molecule analysis. The DDA platform is built upon a digital microfluidic microwell array chip, integrating magnetic bead-based immunocapture with RNA-guided CRISPR/Cas13a signal amplification. This system enables a fully automated, "sample-in, answer-out" workflow. By systematically optimizing the entire process, DDA significantly reduces background noise and enhances detection sensitivity, achieving a limit of detection (LOD) down to 100 zM for key protein biomarkers. This represents a >100-fold improvement over leading commercial ultrasensitive assays. With single-molecule resolution and full automation, DDA provides a robust solution for the precise quantification of low-abundance immune biomarkers. As a proof-of-concept, we demonstrate its ability to accurately quantify key heart-failure-associated biomarkers, including NT-proBNP (LOD: 1 aM), IL-6 (LOD: 1.5 aM), and TNF-α (LOD: 2.5 aM), directly in complex serum samples. This platform holds great promise for automated multibiomarker screening and risk assessment, showcasing its powerful potential for the early diagnosis of major diseases such as cardiovascular diseases, cancers, neurodegenerative disorders, and infectious diseases.

Humans↗

PubMed2005

Directed growth of pure phosphatidylcholine nanotubes in microfluidic channels.

The morphology of self-assembled phospholipid membranes (e.g., micelles, vesicles, rods, tubes, etc.) depends on the method of formation, secondary manipulation, temperature, and storage conditions. In this contribution, microfluidic systems are used to create pure phosphatidylcholine (PC) micro- and nanotubes with unprecedented lengths. Tubes up to several centimeters in length and aligned with the long axis of the microchannel were created from spots of dry films of 1,2-dilauroyl-sn-glycero-3-phosphocholine (DLPC), 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). These high aspect ratio structures, which, to our knowledge, represent the first examples of extended tubes formed from pure PC lipids, were examined by fluorescence microscopy, electron and optical microscopy, and optical manipulation tools (i.e., a laser trap and laser scalpel) to characterize structure and stability. In particular, the tubular structure was confirmed by observation of fluorescent dyes that were sequestered within the aqueous cavity or within the phospholipid tube. Compared to other phospholipid tubes, the tubes formed from PC lipids in microfluidic channels show high mechanical stability and rigidity that depend on tube size, age, and storage conditions.

Microfluidics↗

PubMed2006

Controllable preparation of monodisperse O/W and W/O emulsions in the same microfluidic device.

This letter presents a simple way to prepare monodisperse O/W and W/O emulsions in the same T-junction microfluidic device just by changing the wetting properties of the microchannel wall with different surfactants. Highly uniform droplets ranging from 50 to 400 mum with a polydispersity index (sigma) value of less than 2% were successfully prepared. With the change in surfactants and surfactant concentrations, the interfacial tension and the wetting properties varied, and disordered or ordered two-phase flow patterns could be controllable. Monodisperse O/W and W/O emulsions were prepared under the action of a cross-flowing shear force or a perpendicular shear force by using an oil solution with 0.1-2.0 wt % Span 80 and an aqueous solution with 0.1-2.0 wt % Tween 20 as a continuous-phase flow, respectively. It gives a controllable method of preparing O/W and W/O emulsions in the same microfluidic device.

Emulsions↗

PubMed2006

Hybridization of DNA to bead-immobilized probes confined within a microfluidic channel.

We report the factors influencing the capture of DNA by DNA-modified microbeads confined within a microfluidic channel. Quantitative correlation of target capture efficiency to probe surface concentration, solution flow rate, and target concentration are discussed. The results indicate that the microfluidic system exhibits a limit of detection of approximately 10(-10) M (approximately 10(-16) mol) DNA and a selectivity factor of approximately 8 x 10(3). Typical hybridization times are on the order of minutes.

Biotinylation↗