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1.
In this article, the authors present the fabrication of an enzyme-entrapped alginate hollow fiber using a microfluidic device. Further use of enzyme-entrapped alginate hollow fibers as a biocatalytic microchemical reactor for chemical synthesis is also deliberated in this article. To ensure that there is no enzyme leaching from the fiber, fiber surfaces were coated with chitosan. To confine the mobility of reactants and products within the porous hollow fibers the entire fibers were embedded into a transparent polydimethylsiloxane (PDMS) matrix which also works as a support matrix. A vanadium-containing bromoperoxidase enzyme isolated from Corallina confusa was used as a model enzyme to demonstrate the use of these alginate hollow-fiber reactors in bromo-oxidation of phenol red to bromophenol blue at different dye flow rates. Stability of the entrapped enzyme at different temperatures and the effect of the chitosan coating on the reaction conversion were also studied. It was observed that molecules as big as 27 kDa can be retained in the matrix after coating with chitosan while molecules with molecular-weight of around 378 Da can still diffuse in and out of the matrix. The kinetic conversion rate in this microfluidic bioreactor was more than 41-fold faster when compared with the standard test-tube procedure. 相似文献
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3.
Cellular transplantation is a promising technology with great clinical potential in regenerative medicine and disease management. However, effective control over patient immunological response is essential. The encapsulation of cells within hydrogel microspheres is an increasingly prevalent method for the protection of cellular grafts from immune rejection. Microfluidic “chip” reactors present elegant solutions to several capsule generation issues, including the requirement for intercapsule uniformity, high reproducibility, and sterile, good manufacturing practice compliance. This study presents a novel method for the on-chip production of stable, highly monodisperse alginate microspheres and demonstrates its utility in the encapsulation of an immortalized human-derived cell line. Four populations of immortalized human embryonic kidney cells (HEK293) were encapsulated on chip within monodisperse alginate capsules. Cell viability measurements were recorded for each of the four encapsulated populations for 90 days. 相似文献
4.
In this work, we demonstrate a robust and reliable approach to fabricate multi-compartment particles for cell co-culture studies. By taking advantage of the laminar flow within our microfluidic nozzle, multiple parallel streams of liquids flow towards the nozzle without significant mixing. Afterwards, the multiple parallel streams merge into a single stream, which is sprayed into air, forming monodisperse droplets under an electric field with a high field strength. The resultant multi-compartment droplets are subsequently cross-linked in a calcium chloride solution to form calcium alginate micro-particles with multiple compartments. Each compartment of the particles can be used for encapsulating different types of cells or biological cell factors. These hydrogel particles with cross-linked alginate chains show similarity in the physical and mechanical environment as the extracellular matrix of biological cells. Thus, the multi-compartment particles provide a promising platform for cell studies and co-culture of different cells. In our study, cells are encapsulated in the multi-compartment particles and the viability of cells is quantified using a fluorescence microscope after the cells are stained for a live/dead assay. The high cell viability after encapsulation indicates the cytocompatibility and feasibility of our technique. Our multi-compartment particles have great potential as a platform for studying cell-cell interactions as well as interactions of cells with extracellular factors. 相似文献
5.
This study presents a method for density-based separation of monodisperse encapsulated cells using a standing surface acoustic wave (SSAW) in a microchannel. Even though monodisperse polymer beads can be generated by the state-of-the-art technology in microfluidics, the quantity of encapsulated cells cannot be controlled precisely. In the present study, mono-disperse alginate beads in a laminar flow can be separated based on their density using acoustophoresis. A mixture of beads of equal sizes but dissimilar densities was hydrodynamically focused at the entrance and then actively driven toward the sidewalls by a SSAW. The lateral displacement of a bead is proportional to the density of the bead, i.e., the number of encapsulated cells in an alginate bead. Under optimized conditions, the recovery rate of a target bead group (large-cell-quantity alginate beads) reached up to 97% at a rate of 2300 beads per minute. A cell viability test also confirmed that the encapsulated cells were hardly damaged by the acoustic force. Moreover, cell-encapsulating beads that were cultured for 1 day were separated in a similar manner. In conclusion, this study demonstrated that a SSAW can successfully separate monodisperse particles by their density. With the present technique for separating cell-encapsulating beads, the current cell engineering technology can be significantly advanced. 相似文献
6.
Selection of particles or cells of specific shapes from a complex mixture is an essential procedure for various biological and industrial applications, including synchronization of the cell cycle, classification of environmental bacteria, and elimination of aggregates from synthesized particles. Here, we investigate the separation behaviors of nonspherical and spherical particles∕cells in the hydrodynamic filtration (HDF) scheme, which was previously developed for continuous size-dependent particle∕cell separation. Nonspherical particle models were prepared by coating the hemisphere of spherical polymer particles with a thin Au layer and by bonding the Janus particles to form twins and triplets resembling dividing and aggregating cells, respectively. High-speed imaging revealed a difference in the separation behaviors of spherical and nonspherical particles at a branch point; nonspherical particles showed rotation behavior and did not enter the branch channel even when their minor axis was smaller than the virtual width of the flow region entering the branch channel, w(1). The confocal-laser high-speed particle intensity velocimetry system visualized the flow profile inside the HDF microchannel, demonstrating that the steep flow-velocity distribution at the branch point is the main factor causing the rotation behavior of nonspherical particles. As applications, we successfully separated spherical and nonspherical particles with various major∕minor lengths and also demonstrated the selection of budding∕single cells from a yeast cell mixture. We therefore conclude that the HDF scheme can be used for continuous shape-based particle∕cell separation. 相似文献
7.
Flow cytometry is a standard analytical method in cell biology and clinical diagnostics and is widely distributed for the experimental investigation of microparticle characteristics. In this work, the design, realization, and measurement results of a novel planar optofluidic flow cytometric device with an integrated three-dimensional (3D) adjustable optofluidic lens system for forward-scattering∕extinction-based biochemical analysis fabricated by silicon micromachining are presented. To our knowledge, this is the first planar cytometric system with the ability to focus light three-dimensionally on cells∕particles by the application of fluidic lenses. The single layer microfluidic platform enables versatile 3D hydrodynamic sample focusing to an arbitrary position in the channel and incorporates integrated fiber grooves for the insertion of glass fibers. To confirm the fluid dynamics and raytracing simulations and to characterize the sensor, different cell lines and sets of microparticles were investigated by detecting the extinction (axial light loss) signal, demonstrating the high sensitivity and sample discrimination capability of this analysis system. The unique features of this planar microdevice enable new biotechnological analysis techniques due to the highly increased sensitivity. 相似文献
8.
Ramesh Ramji Ming Wang Ali Asgar S. Bhagat Daniel Tan Shao Weng Nitish V. Thakor Chwee Teck Lim Chia-Hung Chen 《Biomicrofluidics》2014,8(3)
Droplet-based microfluidics has shown potential in high throughput single cell assays by encapsulating individual cells in water-in-oil emulsions. Ordering cells in a micro-channel is necessary to encapsulate individual cells into droplets further enhancing the assay efficiency. This is typically limited due to the difficulty of preparing high-density cell solutions and maintaining them without cell aggregation in long channels (>5 cm). In this study, we developed a short pinched flow channel (5 mm) to separate cell aggregates and to form a uniform cell distribution in a droplet-generating platform that encapsulated single cells with >55% encapsulation efficiency beating Poisson encapsulation statistics. Using this platform and commercially available Sox substrates (8-hydroxy-5-(N,N-dimethylsulfonamido)-2-methylquinoline), we have demonstrated a high throughput dynamic single cell signaling assay to measure the activity of receptor tyrosine kinases (RTKs) in lung cancer cells triggered by cell surface ligand binding. The phosphorylation of the substrates resulted in fluorescent emission, showing a sigmoidal increase over a 12 h period. The result exhibited a heterogeneous signaling rate in individual cells and showed various levels of drug resistance when treated with the tyrosine kinase inhibitor, gefitinib. 相似文献
9.
Cell isolation in designated regions or from heterogeneous samples is often required for many microfluidic cell-based assays. However, current techniques have either limited throughput or are incapable of viable off-chip collection. We present an innovative approach, allowing high-throughput and label-free cell isolation and enrichment from heterogeneous solution using cell size as a biomarker. The approach utilizes the irreversible migration of particles into microscale vortices, developed in parallel expansion-contraction trapping reservoirs, as the cell isolation mechanism. We empirically determined the critical particle∕cell diameter D(crt) and the operational flow rate above which trapping of cells∕particles in microvortices is initiated. Using this approach we successfully separated larger cancer cells spiked in blood from the smaller blood cells with processing rates as high as 7.5×10(6) cells∕s. Viable long-term culture was established using cells collected off-chip, suggesting that the proposed technique would be useful for clinical and research applications in which in vitro culture is often desired. The presented technology improves on current technology by enriching cells based on size without clogging mechanical filters, employing only a simple single-layered microfluidic device and processing cell solutions at the ml∕min scale. 相似文献
10.
We present facile strategies for the fabrication of two types of microfluidic devices made of hydrogels using the natural biopolymers, alginate, and gelatin as substrates. The processes presented include the molding-based preparation of hydrogel plates and their chemical bonding. To prepare calcium-alginate hydrogel microdevices, we suppressed the volume shrinkage of the alginate solution during gelation using propylene glycol alginate in the precursor solution along with sodium alginate. In addition, a chemical bonding method was developed using a polyelectrolyte membrane of poly-L-lysine as the electrostatic glue. To prepare gelatin-based microdevices, we used microbial transglutaminase to bond hydrogel plates chemically and to cross-link and stabilize the hydrogel matrix. As an application, mammalian cells (fibroblasts and vascular endothelial cells) were cultivated on the microchannel surface to form three-dimensional capillary-embedding tissue models for biological research and tissue engineering. 相似文献
11.
The process of blood vessel formation is accompanied by very minimal flow in the beginning, followed by increased flow rates once the vessel develops sufficiently. Many studies have been performed for endothelial cells at shear stress levels of 0.1-60 dyn∕cm(2); however, little is known about the effect of extremely slow flows (shear stress levels of 10(-4)-10(-2) dyn∕cm(2)) that endothelial cells may experience during early blood vessel formation where flow-sensing by indirect mass transport sensing rather than through mechanoreceptor sensing mechanisms would become more important. Here, we show that extremely low flows enhance proliferation, adherens junction protein localization, and nitric oxide secretion of endothelial cells, but do not induce actin filament reorganization. The responses of endothelial cells in different flow microenvironments need more attention because increasing evidence shows that endothelial cell behaviors at the extremely slow flow regimes cannot be linearly extrapolated from observations at faster flow rates. The devices and methods described here provide a useful platform for such studies. 相似文献
12.
Cell encapsulation technology is a promising strategy applicable to tissue engineering and cell therapy. Many advanced microencapsulation chips that function via multiple syringe pumps have been developed to generate mono-disperse hydrogel beads encapsulating cells. However, their operation is difficult and only trained microfluidic engineers can use them with dexterity. Hence, we propose a microfluidic manifold system, driven by a single syringe pump, which can enable the setup of automated flow sequences and generate highly mono-disperse alginate beads by minimizing disturbances to the pump pressure. The encapsulation of P19 mouse embryonic carcinoma cells and embryonic body formation are demonstrated to prove the efficiency of the proposed system. 相似文献
13.
Vascular function, homeostasis, and pathological development are regulated by the endothelial cells that line blood vessels. Endothelial function is influenced by the integrated effects of multiple factors, including hemodynamic conditions, soluble and insoluble biochemical signals, and interactions with other cell types. Here, we present a membrane microfluidic device that recapitulates key components of the vascular microenvironment, including hemodynamic shear stress, circulating cytokines, extracellular matrix proteins, and multiple interacting cells. The utility of the device was demonstrated by measuring monocyte adhesion to and transmigration through a porcine aortic endothelial cell monolayer. Endothelial cells grown in the membrane microchannels and subjected to 20 dynes∕cm(2) shear stress remained viable, attached, and confluent for several days. Consistent with the data from macroscale systems, 25 ng∕ml tumor necrosis factor (TNF)-α significantly increased RAW264.7 monocyte adhesion. Preconditioning endothelial cells for 24 h under static or 20 dynes∕cm(2) shear stress conditions did not influence TNF-α-induced monocyte attachment. In contrast, simultaneous application of TNF-α and 20 dynes∕cm(2) shear stress caused increased monocyte adhesion compared with endothelial cells treated with TNF-α under static conditions. THP-1 monocytic cells migrated across an activated endothelium, with increased diapedesis in response to monocyte chemoattractant protein (MCP)-1 in the lower channel of the device. This microfluidic platform can be used to study complex cell-matrix and cell-cell interactions in environments that mimic those in native and tissue engineered blood vessels, and offers the potential for parallelization and increased throughput over conventional macroscale systems. 相似文献
14.
Schmolke H Demming S Edlich A Magdanz V Büttgenbach S Franco-Lara E Krull R Klages CP 《Biomicrofluidics》2010,4(4):44113
Polyelectrolyte multilayers (PEMs) based on the combinations poly(diallyldimethylammonium chloride)∕poly(acrylic acid) (PDADMAC∕PAA) and poly(allylamine hydrochloride)∕PAA (PAH∕PAA) were adsorbed on poly(dimethylsiloxane) (PDMS) and tested for nonspecific surface attachment of hydrophobic yeast cells using a parallel plate flow chamber. A custom-made graft copolymer containing poly(ethylene glycol) (PEG) side chains (PAA-g-PEG) was additionally adsorbed on the PEMs as a terminal layer. A suitable PEM modification effectively decreased the adhesion strength of Saccharomyces cerevisiae DSM 2155 to the channel walls. However, a further decrease in initial cell attachment and adhesion strength was observed after adsorption of PAA-g-PEG copolymer onto PEMs from aqueous solution. The results demonstrate that a facile layer-by-layer surface functionalization from aqueous solutions can be successfully applied to reduce cell adhesion strength of S. cerevisiae by at least two orders of magnitude compared to bare PDMS. Therefore, this method is potentially suitable to promote planktonic growth inside capped PDMS-based microfluidic devices if the PEM deposition is completed by a dynamic flow-through process. 相似文献
15.
L. Yu S. M. Grist S. S. Nasseri E. Cheng Y.-C. E. Hwang C. Ni K. C. Cheung 《Biomicrofluidics》2015,9(2)
Creating multicellular tumor spheroids is critical for characterizing anticancer treatments since they may provide a better model of the tumor than conventional monolayer culture. Moreover, tumor cell interaction with the extracellular matrix can determine cell organization and behavior. In this work, a microfluidic system was used to form cell-laden core-shell beads which incorporate elements of the extracellular matrix and support the formation of multicellular spheroids. The bead core (comprising a mixture of alginate, collagen, and reconstituted basement membrane, with gelation by temperature control) and shell (comprising alginate hydrogel, with gelation by ionic crosslinking) were simultaneously formed through flow focusing using a cooled flow path into the microfluidic chip. During droplet gelation, the alginate acts as a fast-gelling shell which aids in preventing droplet coalescence and in maintaining spherical droplet geometry during the slower gelation of the collagen and reconstituted basement membrane components as the beads warm up. After droplet gelation, the encapsulated MCF-7 cells proliferated to form uniform spheroids when the beads contained all three components: alginate, collagen, and reconstituted basement membrane. The dose-dependent response of the MCF-7 cell tumor spheroids to two anticancer drugs, docetaxel and tamoxifen, was compared to conventional monolayer culture. 相似文献
16.
Cara Young Kester Rozario Christophe Serra Laura Poole-Warren Penny Martens 《Biomicrofluidics》2013,7(4)
Biosynthetic microspheres have the potential to address some of the limitations in cell microencapsulation; however, the generation of biosynthetic hydrogel microspheres has not been investigated or applied to cell encapsulation. Droplet microfluidics has the potential to produce more uniform microspheres under conditions compatible with cell encapsulation. Therefore, the aim of this study was to understand the effect of process parameters on biosynthetic microsphere formation, size, and morphology with a co-flow microfluidic method. Poly(vinyl alcohol) (PVA), a synthetic hydrogel and heparin, a glycosaminoglycan were chosen as the hydrogels for this study. A capillary-based microfluidic droplet generation device was used, and by varying the flow rates of both the polymer and oil phases, the viscosity of the continuous oil phase, and the interfacial surface tension, monodisperse spheres were produced from ∼200 to 800 μm. The size and morphology were unaffected by the addition of heparin. The modulus of spheres was 397 and 335 kPa for PVA and PVA/heparin, respectively, and this was not different from the bulk gel modulus (312 and 365 for PVA and PVA/heparin, respectively). Mammalian cells encapsulated in the spheres had over 90% viability after 24 h in both PVA and PVA/heparin microspheres. After 28 days, viability was still over 90% for PVA-heparin spheres and was significantly higher than in PVA only spheres. The use of biosynthetic hydrogels with microfluidic and UV polymerisation methods offers an improved approach to long-term cell encapsulation. 相似文献
17.
虾,蟹壳聚糖溶液粘度变化的研究 总被引：2，自引：0，他引：2
分别对虾、蟹壳聚糖溶液在不同浓度、p H值、温度下对其粘度变化的影响进行了研究 ,并对引起这些变化的原因进行了初步探讨 .结果表明 ,在溶液中蟹壳聚糖的降解速度比虾壳聚糖的降解速度快 ,在相同条件下虾壳聚糖溶液的粘度大于蟹壳聚糖溶液的粘度 .此结果对以壳聚糖为原料制备胶囊有着一定的理论意义和实际意义 相似文献
18.
Seamless integration of biological components with electrochemical sensors is critical in the development of microdevices for cell analysis. The present paper describes the integration miniature Au electrodes next to immune cells (macrophages) in order to detect cell-secreted hydrogen peroxide (H(2)O(2)). Photopatterning of poly(ethylene glycol) (PEG) hydrogels was used to both immobilize horseradish peroxidase molecules onto electrodes and to define regions for cell attachment in the vicinity of sensing electrodes. Electrodes micropatterned in such a manner were enclosed inside poly(dimethylsiloxane) fluid conduits and incubated with macrophages. The cells attached onto the exposed glass regions in the vicinity of the electrodes and nowhere else on the non-fouling PEG hydrogel surface. A microfluidic device was converted into an electrochemical cell by placing flow-through Ag∕AgCl reference and Pt wire counter electrodes at the outlet and inlet, respectively. This microdevice with integrated H(2)O(2)-sensing electrodes had sensitivity of 27 μA∕cm(2) mM with a limit of detection of 2 μM. Importantly, this microdevice allowed controllable seeding of macrophages next to electrodes, activation of these cells and on-chip monitoring of H(2)O(2) release in real time. In the future, this biosensor platform may be utilized for monitoring of macrophage responses to pathogens or for the study of inflammatory signaling in micropatterned cell cultures. 相似文献
19.
Kuo-yuan Chung Narayan Chandra Mishra Chen-chi Wang Feng-hui Lin Keng-hui Lin 《Biomicrofluidics》2009,3(2)
In this paper, we demonstrate for the first time the technique to using microfluidics to fabricate tissue engineering scaffolds with uniform pore sizes. We investigate both the bubble generation of the microfluidic device and the application of foam as a tissue engineering scaffold. Our microfluidic device consists of two concentric tapered channels, which are made by micropipettes. Nitrogen gas and aqueous alginate solution with Pluronic® F127 surfactant are pumped through the inner and the outer channels, respectively. We observe rich dynamic patterns of bubbles encapsulated in the liquid droplets. The size of the bubble depends linearly on the gas pressure and inversely on the liquid flow rate. In addition, monodisperse bubbles self-assemble into crystalline structures. The liquid crystalline foams are further processed into open-cell solid foams. The novel foam gel was used as a scaffold to culture chondrocytes. 相似文献
20.
We demonstrate the generation of water-in-water (w/w) jets and emulsions by combining droplet microfluidics and aqueous two-phase systems (ATPS). The application of ATPS in microfluidics has been hampered by the low interfacial tension between typical aqueous phases. The low tension makes it difficult to form w/w droplets with conventional droplet microfluidic approaches. We show that by mechanically perturbing a stable w/w jet, w/w emulsions can be prepared in a controlled and reproducible fashion. We also characterize the encapsulation ability of w/w emulsions and demonstrate that their encapsulation efficiency can be significantly enhanced by inducing formation of precipitates and gels at the w/w interfaces. Our work suggests a biologically and environmentally friendly platform for droplet microfluidics and establishes the potential of w/w droplet microfluidics for encapsulation-related applications. 相似文献