共查询到20条相似文献，搜索用时 31 毫秒
1.
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
2.
A new strategy for magnetically manipulating and isolating adherent cells with extremely high post-collection purity and viability is reported. Micromolded magnetic elements (termed microrafts) were fabricated in an array format and used as culture surfaces and carriers for living, adherent cells. A poly(styrene-co-acrylic acid) polymer containing well dispersed magnetic nanoparticles was developed for creating the microstructures by molding. Nanoparticles of γFe(2)O(3) at concentrations up to 1% wt.∕wt. could be used to fabricate microrafts that were optically transparent, highly magnetic, biocompatible, and minimally fluorescent. To prevent cellular uptake of nanoparticles from the magnetic polymer, a poly(styrene-co-acrylic acid) layer lacking γFe(2)O(3) nanoparticles was placed over the initial magnetic microraft layer to prevent cellular uptake of the γFe(2)O(3) during culture. The microraft surface geometry and physical properties were altered by varying the polymer concentration or layering different polymers during fabrication. Cells plated on the magnetic microrafts were visualized using standard imaging techniques including brightfield, epifluorescence, and confocal microscopy. Magnetic microrafts possessing cells of interest were dislodged from the array and efficiently collected with an external magnet. To demonstrate the feasibility of cell isolation using the magnetic microrafts, a mixed population of wild-type cells and cells stably transfected with a fluorescent protein was plated onto an array. Microrafts possessing single, fluorescent cells were released from the array and magnetically collected. A post-sorting single-cell cloning rate of 92% and a purity of 100% were attained. 相似文献
3.
A non-positional (or suspension) cell microarray was developed using shape-coded SU-8 photoresist microboards for potential application in multiplex and high-throughput cell-based assays. A conventional photolithography process on glass slides produced various shapes of SU-8 micropatterns that had a lateral dimension of 200 μm and a thickness of 40 μm. The resultant micropatterns were detached from the slides by sonication and named "microboards" due to the fact that had a much larger lateral dimension than thickness. The surfaces of the SU-8 microboards were modified with collagen to promote cell adhesion, and it was confirmed that collagen-coated SU-8 microboards supported cell adhesion and proliferation. Seeding of cells into poly(ethylene glycol)(PEG) hydrogel-coated well plates containing collagen-modified microboards resulted in selective cell adhesion onto the microboards due to the non-adhesiveness of PEG hydrogel toward cells, thereby creating non-positional arrays of microboards carrying cells. Finally, two different cell types (fibroblasts and HeLa cells) were separately cultured on different shapes of microboards and subsequently mixed together to create a non-positional cell microarray consisting of multiple cell types where each cell could be easily identified by the shape of the microboard to which they had adhered. Because numerous unique shapes of microboards can be fabricated using this method by simply changing the photomask designs, high throughput and multiplex cell-based assays would be easily achieved with this system in the future. 相似文献
4.
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
5.
The development of widely applicable point-of-care sensing and diagnostic devices can benefit from simple and inexpensive fabrication techniques that expedite the design, testing, and implementation of lab-on-a-chip devices. In particular, electrodes integrated within microfluidic devices enable the use of electrochemical techniques for the label-free detection of relevant analytes. This work presents a novel, simple, and cost-effective bench-top approach for the integration of high surface area three-dimensional structured electrodes fabricated on polystyrene (PS) within poly(dimethylsiloxane) (PDMS)-based microfluidics. Optimization of PS-PDMS bonding results in integrated devices that perform well under pressure and fluidic flow stress. Furthermore, the fabrication and bonding processes are shown to have no effect on sensing electrode performance. Finally, the on-chip sensing capabilities of a three-electrode electrochemical cell are demonstrated with a model redox compound, where the high surface area structured electrodes exhibit ultra-high sensitivity. We propose that the developed approach can significantly expedite and reduce the cost of fabrication of sensing devices where arrays of functionalized electrodes can be used for point-of-care analysis and diagnostics. 相似文献
6.
Cell culture and harvest are the most upstream operation for a completely integrated cell assay chip. In our previous work, thermoresponsive poly(N-isopropylacrylamide) (PNIPAAm) was successfully grafted onto polydimethylsiloxane (PDMS) surface via benzophenone-initiated photopolymerization. In the present work, the PNIPAAm-grafted-PDMS (PNIPAAm-g-PDMS) surface was explored for thermomodulated cell culture and noninvasive harvest in microfluidic channels. Using COS 7 fibroblast from African green monkey kidney as the model cells, the thermomodulated adhering and detaching behaviors of the cells on the PNIPAAm-g-PDMS surfaces were optimized with respect to PNIPAAm-grafting yields and gelatin modification. The viability of the cells cultured on and harvested from the PNIPAAm-g-PDMS surface with the thermomodulated noninvasive protocol was estimated against the traditional cell culture∕harvest method involving trypsin digestion. The configuration of the microchannel on the PNIPAAm-g-PDMS chip was evaluated for static cell culture. Using a pipette-shaped PNIPAAm-g-PDMS microchannel, long-term cell culture could be achieved at 37 °C with periodic change of the culture medium every 12 h. After moving the microchip from the incubator set at 37 °C to the room temperature, the proliferated cells could be spontaneously detached from the PNIPAAm-g-PDMS surface of the upstream chamber and transferred by a gentle fluid flow to the downstream chamber, wherein the transferred cells could be subcultured. The thermomodulated cell culture, harvest, and passage operations on the PNIPAAm-g-PDMS microfluidic channels were demonstrated. 相似文献
7.
Particle focusing in microfluidic devices is a necessary step in medical applications, such as detection, sorting, counting, and flow cytometry. This study proposes a microdevice that combines insulator-based and metal-electrode dielectrophoresis for the three-dimensional focusing of biological cells. Four insulating structures, which form an X pattern, are employed to confine the electric field in a conducting solution, thereby creating localized field minima in the microchannel. These electrodes, 56-μm-wide at the top and bottom surfaces, are connected to one electric pole of the power source. The electrodes connected to the opposite pole, which are at the sides of the microchannel, have one of three patterns: planar, dual-planar, or three-dimensional. Therefore, low-electric-field regions at the center of the microchannel are generated to restrain the viable HeLa cells with negative dielectrophoretic response. The array of insulating structures aforementioned is used to enhance the performance of confinement. According to numerical simulations, three-dimensional electrodes exhibit the best focusing performance, followed by dual-planar and planar electrodes. Experimental results reveal that increasing the strength of the applied electric field or decreasing the inlet flow rate significantly enhances focusing performance. The smallest width of focusing is 17 μm for an applied voltage and an inlet flow rate of 35 V and 0.5 μl/min, respectively. The effect of the inlet flow rate on focusing is insignificant for an applied voltage of 35 V. The proposed design retains the advantages of insulator-based dielectrophoresis with a relatively low required voltage. Additionally, complicated flow controls are unnecessary for the three-dimensional focusing of cells. 相似文献
8.
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
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.
A new method is demonstrated to transport particles, cells, and other microorganisms using rectified ac electro-osmotic flows in open microchannels. The rectified flow is obtained by synchronous zeta potential modulation with the driving potential in the microchannel. Experiments were conducted to transport both neutral, charged particles, and microorganisms of various sizes. A maximum speed of 50 μm∕s was obtained for 8 μm polystyrene beads, without any electrolysis, using a symmetrical square waveform driving electric field of 5 V∕mm at 10 Hz and a 360 V gate potential with its polarity synchronized with the driving potential (phase lag=0°). 相似文献
11.
Precise patterning of metals is required for diverse microfluidic and microelectromechanical system (MEMS) applications ranging from the separation of proteins to the manipulation of single cells and drops of water-in-oil emulsions. Here we present a very simple, inexpensive method for fabricating micropatterned electrodes. We deposit a thin metal layer of controlled thickness using wet chemistry, thus eliminating the need for expensive equipment typically required for metal deposition. We demonstrate that the resulting deposited metal can be used to fabricate functional electrodes: The wet-deposited metal film can sustain patterning by photolithography down to micron-sized features required for MEMS and microfluidic applications, and its properties are suitable for operative electrodes used in a wide range of microfluidic applications for biological studies. 相似文献
12.
Precisely controlling the spatial distribution of biomolecules on biomaterial surface is important for directing cellular activities in the controlled cell microenvironment. This paper describes a polydimethylsiloxane (PDMS) gradient-generating microfluidic device to immobilize the gradient of cellular adhesive Arg-Gly-Asp (RGD) peptide on poly (ethylene glycol) (PEG) hydrogel. Hydrogels are formed by exposing the mixture of PEG diacrylate (PEGDA), acryloyl-PEG-RGD, and photo-initiator with ultraviolet light. The microfluidic chip was simulated by a fluid dynamic model for the biomolecule diffusion process and gradient generation. PEG hydrogel covalently immobilized with RGD peptide gradient was fabricated in this microfluidic device by photo-polymerization. Bone marrow derived rat mesenchymal stem cells (MSCs) were then cultured on the surface of RGD gradient PEG hydrogel. Cell adhesion of rat MSCs on PEG hydrogel with various RGD gradients were then qualitatively and quantitatively analyzed by immunostaining method. MSCs cultured on PEG hydrogel surface with RGD gradient showed a grated fashion for cell adhesion and spreading that was proportional to RGD concentration. It was also found that 0.107–0.143 mM was the critical RGD concentration range for MSCs maximum adhesion on PEG hydrogel. 相似文献
13.
A size-selective cell sorting microfluidic device that utilizes optical force is developed. The device consists of a three-dimensional polydimethylsiloxane microstructure comprised of two crossed microchannels in a three-dimensional configuration. A line shaped focused laser beam is used for automatic size-selective cell sorting in a continuous flow environment. As yeast cells in an aqueous medium are fed continuously into a lower channel, the line shaped focused laser beam is applied (perpendicular to the direction of flow) at the junction of the two crossed channels. The scattering force of the laser beam was employed to push cells matching specific criteria upward from one channel to another. The force depends on the size of the cells, the laser power, and the fluid flow speed. The variation in size of yeast cells causes them to follow different routes at the intersection. For flow speeds below 30 μm∕s, all yeast cells larger than 3 μm were removed from the main stream. As a result, a high purity sample of small cells can be collected at the outlet of bottom channel. 相似文献
14.
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
15.
Definable surface chemistry is essential for many applications of microfluidic polymer systems. However, small cross-section channels with a high surface to volume ratio enhance passive adsorption of molecules that depletes active molecules in solution and contaminates the channel surface. Here, we present a one-step photochemical process to coat the inner surfaces of closed microfluidic channels with a nanometer thick layer of poly(ethylene glycol) (PEG), well known to strongly reduce non-specific adsorption, using only commercially available reagents in an aqueous environment. The coating consists of PEG diacrylate (PEGDA) covalently grafted to polymer surfaces via UV light activation of the water soluble photoinitiator benzoyl benzylamine, a benzophenone derivative. The PEGDA coating was shown to efficiently limit the adsorption of antibodies and other proteins to <5% of the adsorbed amount on uncoated polymer surfaces. The coating could also efficiently suppress the adhesion of mammalian cells as demonstrated using the HT-29 cancer cell line. In a subsequent equivalent process step, protein in aqueous solution could be anchored onto the PEGDA coating in spatially defined patterns with a resolution of <15 μm using an inverted microscope as a projection lithography system. Surface patterns of the cell binding protein fibronectin were photochemically defined inside a closed microfluidic device that was initially homogeneously coated by PEGDA. The resulting fibronectin patterns were shown to greatly improve cell adhesion compared to unexposed areas. This method opens for easy surface modification of closed microfluidic systems through combining a low protein binding PEG-based coating with spatially defined protein patterns of interest. 相似文献
16.
Weiran Liu Weixing Chen Ran Liu Yuan Ou Haoran Liu Lan Xie Ying Lu Caixia Li Bin Li Jing Cheng 《Biomicrofluidics》2015,9(4)
In sexual assault cases, forensic samples are a mixture of sperm from the perpetrator and epithelial cells from the victim. To obtain an independent short tandem repeat (STR) profile of the perpetrator, sperm cells must be separated from the mixture of cells. However, the current method used in crime laboratories, namely, differential extraction, is a time-consuming and labor-intensive process. To achieve a rapid and automated sample pretreatment process, we fabricated a microdevice for hydrodynamic and size-based separation of sperm and epithelial cells. When cells in suspension were introduced into the device''s microfluidic channels, they were forced to flow along different streamlines and into different outlets due to their different diameters. With the proposed microdevice, sperm can be separated within a short period of time (0.5 h for a 50-μl mock sample). The STR profiles of the products in the sperm outlet reservoir demonstrated that a highly purified male DNA fraction could be obtained (94.0% male fraction). This microdevice is of low-cost and can be easily integrated with other subsequent analysis units, providing great potential in the process of analyzing sexual assault evidence as well as in other areas requiring cell sorting. 相似文献
17.
Found in all eukaryotic cells, linker histones H1 are known to bind to and rearrange nucleosomal linker DNA. In vitro, the fundamental nature of H1∕DNA interactions has attracted wide interest among research communities-from biologists to physicists. Hence, H1∕DNA binding processes and structural and dynamical information about these self-assemblies are of broad importance. Targeting a quantitative understanding of H1 induced DNA compaction mechanisms, our strategy is based on using small-angle x-ray microdiffraction in combination with microfluidics. The usage of microfluidic hydrodynamic focusing devices facilitates a microscale control of these self-assembly processes, which cannot be achieved using conventional bulk setups. In addition, the method enables time-resolved access to structure formation in situ, in particular, to transient intermediate states. The observed time dependent structure evolution shows that the H1∕DNA interaction can be described as a two-step process: an initial unspecific binding of H1 to DNA is followed by a rearrangement of molecules within the formed assemblies. The second step is most likely induced by interactions between the DNA and the H1's charged side chains. This leads to an increase in lattice spacing within the DNA∕protein assembly and induces a decrease in the correlation length of the mesophases, probably due to a local bending of the DNA. 相似文献
18.
The successful encapsulation of human hepatocellular carcinoma (HepG2) cells would greatly assist a broad range of applications in tissue engineering. Due to the harsh conditions during standard chitosan fiber fabrication processes, encapsulation of HepG2 cells in chitosan fibers has been challenging. Here, we describe the successful wet-spinning of chitosan-alginate fibers using a coaxial flow microfluidic chip. We determined the optimal mixing conditions for generating chitosan-alginate fibers, including a 1:5 ratio of 2% (w∕w) water-soluble chitosan (WSC) solution to 2% (w∕w) alginate solution. Ratio including higher than 2% (w∕w) WSC solution increased aggregation throughout the mixture. By suspending cells in the WSC-alginate solution, we successfully fabricated HepG2 cell-laden fibers. The encapsulated HepG2 cells in the chitosan-alginate fibers were more viable than cells encapsulated in pure alginate fibers, suggesting that cross-linked chitosan provides a better environment for HepG2 cells than alginate alone. In addition, we found that the adhesion of HepG2 cells on the chitosan-alginate fiber is much better than that on the alginate fibers. 相似文献
19.
Microvascular network formation is a significant and challenging goal in the engineering of large three-dimensional artificial tissue structures. We show here the development of a fully patent, 3D endothelial cell (microvascular) microfluidic network that has a single inlet and outlet, created in only 28 h in a microdevice involving fluid flow equivalent to natural vasculature. Our microdevice features a tailored "multi-rung ladder" network, a stylized mimic of an arterial-to-venous pedicle, designed to also allow for systematic and reproducible cell seeding. Immunofluorescence staining revealed a highly contiguous endothelial monolayer (human umbilical vein endothelial cells) throughout the whole network after 24 h of continuous perfusion. This network persisted for up to 72 h of culture, providing a useful template from which the effects of surface chemistry, fluid flow, and environmental conditions on the development of artificial vascular networks ex vivo may be rapidly and robustly evaluated. 相似文献
20.
BackgroundThe search for innovative anti-tubercular agents has received increasing attention in tuberculosis chemotherapy because Mycobacterium tuberculosis infection has steadily increased over the years. This underlines the necessity for new methods of preparation for polymer-drug adducts to treat this important infectious disease. The use of poly(ethylene glycol)(PEG) is an alternative producing anti-tubercular derivatives. However, it is not yet known whether PEGylated isonicotinylhydrazide conjugates obtained by direct links with PEG are useful for therapeutic applications.ResultsHere, we synthesized a PEGylated isoniazid (PEG-g-INH or PEG–INH) by gamma radiation-induced polymerization, for the first time. The new prodrugs were characterized using Raman and UV/Vis spectrometry. The mechanism of PEGylated INH synthesis was proposed. The in vitro evaluation of a PEGylated isonicotinylhydrazide macromolecular prodrug was also carried out. The results indicated that PEG–INH inhibited the bacterial growth above 95% as compared with INH, which showed a lower value (80%) at a concentration of 0.25 μM. Similar trends are observed for 0.1, 1, and 5 μM.ConclusionsIn summary, the research suggests that it is possible to covalently attach the PEG onto INH by the proposed method and to obtain a slow-acting isoniazid derivative with little toxicity in vitro and higher anti-mycobacterial potency than the neat drug.How to cite: González-Torres M, Guzmán-Beltrán S, Mata-Gómez M, et al. Synthesis, characterization, and in vitro evaluation of gamma radiation-induced PEGylated isoniazid. Electron J Biotechnol 2019; 41. https://doi.org/10.1016/j.ejbt.2019.07.005. 相似文献