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1.
Focusing cells into a single stream is usually a necessary step prior to counting and separating them in microfluidic devices such as flow cytometers and cell sorters. This work presents a sheathless electrokinetic focusing of yeast cells in a planar serpentine microchannel using dc-biased ac electric fields. The concurrent pumping and focusing of yeast cells arise from the dc electrokinetic transport and the turn-induced ac∕dc dielectrophoretic motion, respectively. The effects of electric field (including ac to dc field ratio and ac field frequency) and concentration (including buffer concentration and cell concentration) on the cell focusing performance were studied experimentally and numerically. A continuous electrokinetic filtration of E. coli cells from yeast cells was also demonstrated via their differential electrokinetic focusing in a serpentine microchannel.  相似文献   

2.
A featured microchip owning three big reservoirs and long turned geometry channel was designed to improve the detection limit of DNA fragments by using floating electrokinetic supercharging (FEKS) method. The novel design matches the FEKS preconcentration needs of a large sample volume introduction with electrokinetic injection (EKI), as well as long duration of isotachophoresis (ITP) process to enrich low concentration sample. In the curved channel [∼45.6 mm long between port 1 (P1) and the intersection point of two channels], EKI and ITP were performed while the side port 3 (P3) was electrically floated. The turn-induced band broadening with or without ITP process was investigated by a computer simulation (using CFD-ACE+ software) when the analytes traveling through the U-shaped geometry. It was found that the channel curvature determined the extent of band broadening, however, which could be effectively eliminated by the way of ITP. After the ITP-stacked zones passed the intersection point from P1, they were rapidly destacked for separation and detection from ITP to zone electrophoresis by using leading ions from P3. The FEKS carried on the novel chip successfully contributed to higher sensitivities of DNA fragments in comparison with our previous results realized on either a single channel or a cross microchip. The analysis of low concentration 50 bp DNA step ladders (0.23 μg∕ml after 1500-fold diluted) was achieved with normal UV detection at 260 nm. The obtained limit of detections (LODs) were on average 100 times better than using conventional pinched injection, down to several ng∕ml for individual DNA fragment.  相似文献   

3.
In this paper, we demonstrate for the first time that insulative dielectrophoresis can induce size-dependent trajectories of DNA macromolecules. We experimentally use λ (48.5 kbp) and T4GT7 (165.6 kbp) DNA molecules flowing continuously around a sharp corner inside fluidic channels with a depth of 0.4 μm. Numerical simulation of the electrokinetic force distribution inside the channels is in qualitative agreement with our experimentally observed trajectories. We discuss a possible physical mechanism for the DNA polarization and dielectrophoresis inside confining channels, based on the observed dielectrophoresis responses due to different DNA sizes and various electric fields applied between the inlet and the outlet. The proposed physical mechanism indicates that further extensive investigations, both theoretically and experimentally, would be very useful to better elucidate the forces involved at DNA dielectrophoresis. When applied for size-based sorting of DNA molecules, our sorting method offers two major advantages compared to earlier attempts with insulative dielectrophoresis: Its continuous operation allows for high-throughput analysis, and it only requires electric field strengths as low as ∼10 V∕cm.  相似文献   

4.
This paper reports the improvement of rectification effects in diffuser∕nozzle structures with viscoelastic fluids. Since rectification in a diffuser∕nozzle structure with Newtonian fluids is caused by inertial effects, micropumps based on this concept require a relatively high Reynolds numbers and high pumping frequencies. In applications with relatively low Reynolds numbers, anisotropic behavior can be achieved with viscoelastic effects. In our investigations, a solution of dilute polyethylene oxide was used as the viscoelastic fluid. A microfluidic device was fabricated in silicon using deep reactive ion etching. The microfluidic device consists of access ports for pressure measurement, and a series of ten diffuser∕nozzle structures. Measurements were carried out for diffuser∕nozzle structures with opening angles ranging from 15° to 60°. Flow visualization, pressure drop and diodicity of de-ionized water and the viscoelastic fluid were compared and discussed. The improvement of diodicity promises a simple pumping concept at low Reynolds numbers for lab-on-a-chip applications.  相似文献   

5.
This paper presents a numerical study of a preconcentrator design that can effectively increase the binding rate at the sensor in a real time manner. The particle enrichment is realized by the ac electrothermal (ACET) effect, which induces fluid movement to carry nanoparticles toward the sensor. The ACET is the only electrical method to manipulate a biological sample of medium to high ionic strength (>0.1 S∕m, e.g., 0.06× phosphate buffered saline). The preconcentrator consists of a pair of electrodes striding over the sensor, simple to implement as it is electrically controlled. This preconcentrator design is compatible and can be readily integrated with many types of micro- to nanosensors. By applying an ac signal over the electrodes, local vortices will generate a large velocity perpendicular to the reaction surface, which enhances transport of analytes toward the sensor. Our simulation shows that the binding rate at the sensor surface is greatly enhanced. Our study also shows that the collection of analytes will be affected by various parameters such as channel height, inlet velocity, and sensor size, and our results will provide guidance in optimization of the preconcentrator design.  相似文献   

6.
Multi-target pathogen detection using heterogeneous medical samples require continuous filtering, sorting, and trapping of debris, bioparticles, and immunocolloids within a diagnostic chip. We present an integrated AC dielectrophoretic (DEP) microfluidic platform based on planar electrodes that form three-dimensional (3D) DEP gates. This platform can continuously perform these tasks with a throughput of 3 μL∕min. Mixtures of latex particles, Escherichia coli Nissle, Lactobacillus, and Candida albicans are sorted and concentrated by these 3D DEP gates. Surface enhanced Raman scattering is used as an on-chip detection method on the concentrated bacteria. A processing rate of 500 bacteria was estimated when 100 μl of a heterogeneous colony of 107 colony forming units ∕ml was processed in a single pass within 30 min.  相似文献   

7.
Electroosmotic (EO) pumps based on dc electroosmosis is plagued by bubble generation and other electrochemical reactions at the electrodes at voltages beyond 1 V for electrolytes. These disadvantages limit their throughput and offset their portability advantage over mechanical syringe or pneumatic pumps. ac electroosmotic pumps at high frequency (>100 kHz) circumvent the bubble problem by inducing polarization and slip velocity on embedded electrodes,1 but they require complex electrode designs to produce a net flow. We report a new high-throughput ac EO pump design based on induced-polarization on the entire channel surface instead of just on the electrodes. Like dc EO pumps, our pump electrodes are outside of the load section and form a cm-long pump unit consisting of three circular reservoirs (3 mm in diameter) connected by a 1×1 mm channel. The field-induced polarization can produce an effective Zeta potential exceeding 1 V and an ac slip velocity estimated as 1 mm∕sec or higher, both one order of magnitude higher than earlier dc and ac pumps, giving rise to a maximum throughput of 1 μl∕sec. Polarization over the entire channel surface, quadratic scaling with respect to the field and high voltage at high frequency without electrode bubble generation are the reasons why the current pump is superior to earlier dc and ac EO pumps.  相似文献   

8.
Electroosmotic flow was studied in thin film microchannels with silicon dioxide and silicon nitride sidewalls formed using plasma-enhanced chemical vapor deposition (PECVD). A sacrificial etching process was employed for channel fabrication allowing for cross-sections with heights of 3 μm, ranging from 2 μm to 50 μm in width. Flow rates were measured for single channels and multichannel electroosmotic pump structures for pH levels ranging from 2.6 to 8.3, and zeta potentials were calculated for both silicon dioxide and silicon nitride surfaces. Flow rates as high as 0.086 μL∕min were measured for nitride multichannel pumps at applied electric fields of 300 V∕mm. The surface characteristics of PECVD nitride were analyzed and compared to more well-known oxide surfaces to determine the density of amine sites compared to silanol sites.  相似文献   

9.
We have designed, built, and evaluated a microfluidic device that uses deterministic lateral displacement for size-based separation. The device achieves almost 100% purity and recovery in continuously sorting two, four, and six micrometer microspheres. We have applied this highly efficient device to the purification of fungal (Aspergillus) spores that are spherical (∼4 μm diameter) with a narrow size distribution. Such separation directly from culture using unfiltered A. niger suspensions is difficult due to a high level of debris. The device produces a two to three increase in the ratio of spores to debris as measured by light scatter in a flow cytometer. The procedure is feasible at densities up to 4.4×106 spores∕ml. This is one of the first studies to apply microfluidic techniques to spore separations and has demonstrated that a passive separation system could significantly reduce the amount of debris in a suspension of fungal spores with virtually no loss of spore material.  相似文献   

10.
Ultrafast microfluidics using surface acoustic waves   总被引：2，自引：0，他引：2  
We demonstrate that surface acoustic waves (SAWs), nanometer amplitude Rayleigh waves driven at megahertz order frequencies propagating on the surface of a piezoelectric substrate, offer a powerful method for driving a host of extremely fast microfluidic actuation and micro∕bioparticle manipulation schemes. We show that sessile drops can be translated rapidly on planar substrates or fluid can be pumped through microchannels at 1–10 cm∕s velocities, which are typically one to two orders quicker than that afforded by current microfluidic technologies. Through symmetry-breaking, azimuthal recirculation can be induced within the drop to drive strong inertial microcentrifugation for micromixing and particle concentration or separation. Similar micromixing strategies can be induced in the same microchannel in which fluid is pumped with the SAW by merely changing the SAW frequency to rapidly switch the uniform through-flow into a chaotic oscillatory flow by exploiting superpositioning of the irradiated sound waves from the sidewalls of the microchannel. If the flow is sufficiently quiescent, the nodes of the transverse standing wave that arises across the microchannel also allow for particle aggregation, and hence, sorting on nodal lines. In addition, the SAW also facilitates other microfluidic capabilities. For example, capillary waves excited at the free surface of a sessile drop by the SAW underneath it can be exploited for micro∕nanoparticle collection and sorting at nodal points or lines at low powers. At higher powers, the large accelerations off the substrate surface as the SAW propagates across drives rapid destabilization of the drop free surface giving rise to inertial liquid jets that persist over 1–2 cm in length or atomization of the entire drop to produce 1–10 μm monodispersed aerosol droplets, which can be exploited for ink-jet printing, mass spectrometry interfacing, or pulmonary drug delivery. The atomization of polymer∕protein solutions can also be used for the rapid synthesis of 150–200 nm polymer∕protein particles or biodegradable polymeric shells in which proteins, peptides, and other therapeutic molecules are encapsulated within for controlled release drug delivery. The atomization of thin films behind a translating drop containing polymer solutions also gives rise to long-range spatial ordering of regular polymer spots whose size and spacing are dependent on the SAW frequency, thus offering a simple and powerful method for polymer patterning without requiring surface treatment or physical∕chemical templating.  相似文献   

11.
Chaurey V  Polanco C  Chou CF  Swami NS 《Biomicrofluidics》2012,6(1):12806-1280614
We present an electrokinetic framework for designing insulator constriction-based dielectrophoresis devices with enhanced ability to trap nanoscale biomolecules in physiological media of high conductivity, through coupling short-range dielectrophoresis forces with long-range electrothermal flow. While a 500-fold constriction enables field focusing sufficient to trap nanoscale biomolecules by dielectrophoresis, the extent of this high-field region is enhanced through coupling the constriction to an electrically floating sensor electrode at the constriction floor. However, the enhanced localized fields due to the constriction and enhanced current within saline media of high conductivity (1 S/m) cause a rise in temperature due to Joule heating, resulting in a hotspot region midway within the channel depth at the constriction center, with temperatures of ∼8°–10°K above the ambient. While the resulting vortices from electrothermal flow are directed away from the hotspot region to oppose dielectrophoretic trapping, they also cause a downward and inward flow towards the electrode edges at the constriction floor. This assists biomolecular trapping at the sensor electrode through enabling long-range fluid sampling as well as through localized stirring by fluid circulation in its vicinity.  相似文献   

12.
A microfluidic device with planar square electrodes is developed for capturing particles from high conductivity media using negative dielectrophoresis (n-DEP). Specifically, Bacillus subtilis and Clostridium sporogenes spores, and polystyrene particles are tested in NaCl solution (0.05 and 0.225 S∕m), apple juice (0.225 S∕m), and milk (0.525 S∕m). Depending on the conductivity of the medium, the Joule heating produces electrothermal flow (ETF), which continuously circulates and transports the particles to the DEP capture sites. Combination of the ETF and n-DEP results in different particle capture efficiencies as a function of the conductivity. Utilizing 20 μm height DEP chambers, “almost complete” and rapid particle capture from lower conductivity (0.05 S∕m) medium is observed. Using DEP chambers above 150 μm in height, the onset of a global fluid motion for high conductivity media is observed. This motion enhances particle capture on the electrodes at the center of the DEP chamber. The n-DEP electrodes are designed to have well defined electric field minima, enabling sample concentration at 1000 distinct locations within the chip. The electrode design also facilitates integration of immunoassay and other surface sensors onto the particle capture sites for rapid detection of target micro-organisms in the future.  相似文献   

13.
We present a microfluidic approach to characterizing temperature-dependent biomolecular interactions. Solvated L-arginine vasopressin (AVP) and its immobilized RNA aptamer (spiegelmer) were allowed to achieve equilibrium binding in a microchip at a series of selected temperatures. Unbound AVP were collected and analyzed with matrix-assisted laser desorption∕ionization mass spectrometry (MALDI-MS), yielding melting curves that reveal highly temperature-dependent zones in which affinity binding (36–45 °C) or dissociation (25–33 °C and 50–65 °C) occurs. Additionally, temperature-dependent binding isotherms were constructed; from these, thermodynamic quantities involved in binding were extracted. The results illustrated a strong change in heat capacity of interaction for this system, suggesting a considerable thermodynamic influence controlling vasopressin-spiegelmer interaction.  相似文献   

14.
A droplet-based micro-total-analysis system involving biosensor performance enhancement by integrated surface-acoustic-wave (SAW) microstreaming is shown. The bioreactor consists of an encapsulated droplet with a biosensor on its periphery, with in situ streaming induced by SAW. This paper highlights the characterization by particle image tracking of the speed distribution inside the droplet. The analyte-biosensor interaction is then evaluated by finite element simulation with different streaming conditions. Calculation of the biosensing enhancement shows an optimum in the biosensor response. These results confirm that the evaluation of the Damköhler and Peclet numbers is of primary importance when designing biosensors enhanced by streaming.It has been pointed out that biosensing performances can be limited by the diffusion of the analytes near the sensing surface.1 In the case of low Peclet number hydrodynamic flows, typical of microfluidic systems, molecule displacements are mainly governed by diffusive effects that affect time scales and sensitivity. To overcome this problem, the enhancement of biosensor performance by electrothermal stirring within microchannels was first reported by Meinhart et al.2 Other authors3, 4 numerically studied the analyte transport as a function of the position of a nanowire-based sensor inside a microchannel, stressing on the fact that the challenge for nanobiosensors is not the sensor itself but the fluidic system that delivers the sample. Addressing this problem, Squires et al.5 developed a simple model applicable to biosensors embedded in microchannels. However, the presented model is limited to the case of a steady flow. The use of surface-acoustic waves (SAWs) for stirring in biomicrofluidic and chemical systems is becoming a popular investigation field,6, 7, 8, 9 especially to overcome problems linked to steady flows by enhancing the liquid∕surface interaction.1, 10, 11 The main challenges that need to be addressed when using SAW-induced stirring are the complexity of the flow and its poor reproducibility. However, some technical solutions were proposed to yield a simplified microstreaming. Yeo et al. presented a centrifugation system based on SAW that produces the rotation of the liquid in a droplet in a reproducible way by playing on the configuration of the transducers and reflectors,12 and presented a comprehensive experimental study of the three-dimensional (3D) flow that causes particle concentration in SAW-stirred droplets,13 revealing the presence of an azimuthal secondary flow in addition to the main vortexlike circular flow present in acoustically stirred droplets. The efficiency of SAW stirring in microdroplets to favorably cope with mass transport issues was finally shown by Galopin et al.,14 but the effect of the stirring on the analyte∕biosensor interaction was not studied. It is expected to overcome mass transport limitations by bringing fresh analytes from the bulk solution to the sensing surface.The studied system, described in Fig. ​Fig.1,1, consists of a microliter droplet microchamber squeezed between a hydrophobic piezoelectric substrate and a hydrophobic glass cover. Rayleigh SAWs are generated using interdigitated transducers (interdigital spacing of 50 μm) laid on an X-cut LiNbO3 substrate.1, 15, 16 The hydrophobicity of the substrate and the cover are obtained by grafting octadecyltrichlorosilane (OTS) self-assembled monolayers (contact angle of 108° and hysteresis of 9°). To do so, the surface is first hydroxylized using oxygen plasma (150 W, 100 mT, and 30 sccm3 O2) during 1 min and then immersed for 3 h into a 1 mM OTS solution with n-hexane as a solvent.Open in a separate windowFigure 1(a) General view of the considered system. (b) Mean value of the measured speeds within the droplet as a function of the inlet power before amplification.When Rayleigh waves are radiated toward one-half of the microchamber, a vortex is created in the liquid around an axis orthogonal to the substrate due to the momentum transfer between the solid and the liquid. This wave is generated under the Rayleigh angle into the liquid.Speed cartographies of the flow induced in the droplet are realized using the particle image tracking technique for different SAW generation powers. To do so, instantaneous images of the flow are taken with a high-speed video camera at 200 frames∕s and an aperture time of 500 μs on a 0.25 μl droplet containing 1 μm diameter fluorescent particles. Figure ​Figure11 shows the mean speed measured in the droplet as a function of the inlet power. The great dependence of the induced mean speed with the SAW power enables a large range of flow speeds in the stirred droplet. Moreover, the flow was visualized with a low depth of field objective. It was found to be circular and two dimensional (2D) in a large thickness range of the droplet.The binding of analytes to immobilized ligands on a biosensor is a two step process, including the mass transport of the analyte to the surface, followed by a complexation step,Abulk↔kmAsurface+B↔ka,kdAB(1)with km as the constant rate for mass transport from and to the sensor, and ka and kd as the constant rates of association and dissociation of the complex.At the biosensor surface, the reaction kinetics consumes analytes but their transport is limited by diffusive effects. In this case, the Damköhler number brings valuable information by comparing these two effects. Calling the characteristic time of reaction and diffusion, respectively, τC and τM, the mixing time in diffusion regime can be approximated by τM≈h2∕D with D as the diffusion coefficient and h a characteristic length of the microchannel. Calling RT the ligand concentration on the surface in mole∕m2, the Damköhler number (Da) can be written asDa=τMτC=kaRThD.(2)Depending on the type of reaction, the calculation of Da helps determine if a specific biointeraction will benefit from a mass SAW-based microstreaming. If the Damköhler number is low, the reaction is slow compared to mass transport and the reaction will not significantly benefit from microstirring. For example, the hybridization of 19 base single stranded DNA in a microfluidic system with a characteristic length of 500 μm is characterized by a Damköhler number of 0.07 and is therefore not significantly influenced by mass transport. On the contrary, the binding of biotin to immobilized streptavidin is characterized by a Da number of approximately 104. In this case, the stirring solution will significantly improve the reaction rate.COMSOL numerical simulations were carried out to study the efficiency of the SAW stirring in the case of a droplet-based microbioreactor with a diameter of 1 mm. Assuming a 2D flow, the simulated model takes into account the convective and diffusive effects in the analyte-carrying fluid and the binding kinetics on the biosensor surface. This approach was thoroughly developed by Meinhart et al.2On the biosensor surface, the following equations are solved:∂B∂t=kacs(RT−B)−kdB,(3)∂B∂t=D|∂c∂y|y=0(4)with c as the local concentration of analytes in the droplet and B as the surface concentration of bound analytes on the biosensor surface. Simulation results show that a depleted zone is formed near the biosensor in the case of an interaction without stirring. This zone is characterized by a low concentration of analytes and results from the trapping of analytes on the biosensor surface, thus creating a concentration gradient on the vicinity of the biosensor. When stirring is applied, the geometry of the depleted zone is modified, as it is pushed in the direction of the flow. The geometry of the depleted zone then depends on many parameters, among which the diffusion coefficient D, the speed distribution of the flow (not only near the biosensor but also in the whole microfluidic system), and the reaction kinetics on the biosensor. In our case, which is assimilated to a simple circular flow, the depleted zone reaches a permanent state consisting of an analyte-poor layer situated in the exterior perimeter of the stirred droplet. The diffusion of analytes is then limited again by diffusion from the inner part of the droplet toward its exterior perimeter (see Fig. ​Fig.22).Open in a separate windowFigure 2(a) Mean concentration of bound analytes vs time for different mean flow speeds. (b) The obtained concentration profiles with and without circular stirring, t=10 000 s.The initial analyte and receptor concentrations are, respectively, 0.1 nM in the solution and 3.3×10−3 nM m on the biosensor surface, the diffusion coefficient is D=10−11 m2 s−1, and the reaction constants are ka=106 M−1 s−1 and kd=10−3 s−1. Simulations show that the mean concentration of bound analytes highly increases with the flow speed, improving the efficiency of the biosensing device. To evaluate the benefits of in situ microstreaming with SAW, the same simulations were conducted for Da numbers ranging from 104 to 108 M−1∕s, by ranging the diffusion coefficient from 4×10−12 to 4×10−9 m2∕s, and the association coefficient ka from 104 to 108 M−1∕s. The enhancement factor of analyte capture, defined as the ratio of the binding rate with streaming B and the binding rate without streaming B0, is plotted in Fig. ​Fig.33 for different values of Da. Calculations are done in the case of a mean flow speed of 0.5 mm∕s.Open in a separate windowFigure 3(a) Enhancement factor (defined as the ratio between binding rate with streaming B and binding rate without streaming B0) for different Damkhöler numbers and (b) normalized enhancement factor for different Peclet numbers.One can notice the saturation of the enhancement factor curve for large value of Da to the value of 3.5 for high Da. This can be explained by the fact that for large ka∕Da ratios, the analytes, which normally require penetration in the depleted zone by diffusion, do not have time to interact with the biosensor when they pass in the vicinity of its surface. The efficiency of the streaming is then reduced for large values of Da. In the case of our specific flow configuration, the enhancement factor reaches 3.2 for the interaction of streptavidin on immobilized biotin (Da=103).The reported simulation results can be compared to an experimental value obtained using the droplet-based surface plasmon resonance sensor streamed in situ using SAW reported by Yeo et al.12 By monitoring the streptavidin∕biotin binding interaction on an activated gold slide, they showed that SAW stirring brings an improvement factor of more than 2. This difference can be accounted to the high complexity of the induced 3D flow, which was modeled in a simple manner in our calculations.Other factors must be taken into account when optimizing the improvement factor, such as the flow velocity and the characteristic length of the mixing. To do so, the Peclet number allows the comparison of the convective and diffusive effects.17 For δC a typical variation in concentration on the distance h, the Peclet number is given byPe=UhD.(5)A significantly high Peclet number causes a decrease in biosensing efficiency as the analytes do not have enough time to interact with the biosensing surface by diffusion through the analyte-poor layer. On the contrary, the case of a low Peclet number corresponds to the diffusion-limited problem. Therefore, for each Damköhler number, there is a Peclet number optimizing this factor. To illustrate this fact, Fig. ​Fig.3b3b shows the calculation of the enhancement factor as a function of the Peclet number for a given Da.In this paper, we showed that surface loading of typical analytes on a droplet-based biosensor can be highly increased by SAW microstirring. The system permits the enhancement of the biosensing performances by the continuous renewal of the analyte-carrying fluid near the sensing surface. Thanks to mean flow speeds measured up to 1800 μm∕s, the SAW microstreaming can be beneficial to the biosensing of a large range of analyte∕ligand interactions. In addition to the biosensing performance improvement, such a method can be easily integrated in micro-micro-total-analysis systems, which makes it a convenient tool for liquid handling in future biochips.  相似文献   

15.
This paper presents a study of electrokinetic transport in single nanopores integrated into vertically stacked three-dimensional hybrid microfluidic∕nanofluidic structures. In these devices, single nanopores, created by focused ion beam (FIB) milling in thin polymer films, provide fluidic connection between two vertically separated, perpendicular microfluidic channels. Experiments address both systems in which the nanoporous membrane is composed of the same (homojunction) or different (heterojunction) polymer as the microfluidic channels. These devices are then used to study the electrokinetic transport properties of synthetic (i.e., polystyrene sulfonate and polyallylamine) and biological (i.e., DNA) polyelectrolytes across these nanopores using both electrical current measurements and confocal microscopy. Both optical and electrical measurements indicate that electro-osmotic transport is predominant over electrophoresis in single nanopores with d>180 nm, consistent with results obtained under similar conditions for nanocapillary array membranes.  相似文献   

16.
This work presents the development of an array of bioreactors where finely controlled stirring is provided at the microliter scale (100–300 μl). The microliter-bioreactor array is useful for performing protocol optimization in up to 96 parallel experiments of hematopoietic stem cell (HSC) cultures. Exploring a wide range of experimental conditions at the microliter scale minimizes cost and labor. Once the cell culture protocol is optimized, it can be applied to large-scale bioreactors for stem cell production at the clinical level. The controlled stirring inside the wells of a standard 96-well plate is provided by buoyancy-driven thermoconvection. The temperature and velocity fields within the culture volume are determined with numerical simulations. The numerical results are verified with experimental velocity measurements using microparticle image velocimetry (μPIV) and are used to define feasible experimental conditions for stem cell cultures. To test the bioreactor array’s functionality, human umbilical cord blood-derived CD34+ cells were cultured for 7 days at five different stirring conditions (0.24–0.58 μm∕s) in six repeated experiments. Cells were characterized in terms of proliferation, and flow cytometry measurements of viability and CD34 expression. The microliter-bioreactor array demonstrates its ability to support HSC cultures under stirred conditions without adversely affecting the cell behavior. Because of the highly controlled operative conditions, it can be used to explore culture conditions where the mass transport of endogenous and exogenous growth factors is selectively enhanced, and cell suspension provided. While the bioreactor array was developed for culturing HSCs, its application can be extended to other cell types.  相似文献   

17.
Two microfluidic systems have been developed for specific analysis of L-glutamate in food based on substrate recycling fluorescence detection. L-glutamate dehydrogenase and a novel enzyme, D-phenylglycine aminotransferase, were covalently immobilized on (i) the surface of silicon microchips containing 32 porous flow channels of 235 μm depth and 25 μm width and (ii) polystyrene Poros™ beads with a particle size of 20 μm. The immobilized enzymes recycle L-glutamate by oxidation to 2-oxoglutarate followed by the transfer of an amino group from D-4-hydroxyphenylglycine to 2-oxoglutarate. The reaction was accompanied by reduction of nicotinamide adenine dinucleotide (NAD+) to NADH, which was monitored by fluorescence detection (εex=340 nm, εem=460 nm). First, the microchip-based system, L-glutamate was detected within a range of 3.1–50.0 mM. Second, to be automatically determined, sequential injection analysis (SIA) with the bead-based system was investigated. The bead-based system was evaluated by both flow injection analysis and SIA modes, where good reproducibility for L-glutamate calibrations was obtained (relative standard deviation of 3.3% and 6.6%, respectively). In the case of SIA, the beads were introduced and removed from the microchip automatically. The immobilized beads could be stored in a 20% glycerol and 0.5 mM ethylenediaminetetraacetic acid solution maintained at a pH of 7.0 using a phosphate buffer for at least 15 days with 72% of the activity remaining. The bead-based system demonstrated high selectivity, where L-glutamate recoveries were between 91% and 108% in the presence of six other L-amino acids tested.  相似文献   

18.
This paper presents the design and analysis of a liquid refractive index sensor that utilizes a unique physical mechanism of resonant optical tunneling effect (ROTE). The sensor consists of two hemicylindrical prisms, two air gaps, and a microfluidic channel. All parts can be microfabricated using an optical resin NOA81. Theoretical study shows that this ROTE sensor has extremely sharp transmission peak and achieves a sensitivity of 760 nm∕refractive index unit (RIU) and a detectivity of 85 000 RIU−1. Although the sensitivity is smaller than that of a typical surface plasmon resonance (SPR) sensor (3200 nm∕RIU) and is comparable to a 95% reflectivity Fabry–Pérot (FP) etalon (440 nm∕RIU), the detectivity is 17 000 times larger than that of the SPR sensor and 85 times larger than that of the FP etalon. Such ROTE sensor could potentially achieve an ultrahigh sensitivity of 10−9 RIU, two orders higher than the best results of current methods.  相似文献   

19.
Surface acoustic wave (SAW) devices with 64 μm wavelength were fabricated on a zinc oxide (ZnO) film deposited on top of an ultra-smooth nanocrystalline diamond (UNCD) layer. The smooth surface of the UNCD film allowed the growth of the ZnO film with excellent c-axis orientation and low surface roughness, suitable for SAW fabrication, and could restrain the wave from significantly dissipating into the substrate. The frequency response of the fabricated devices was characterized and a Rayleigh mode was observed at ∼65.4 MHz. This mode was utilised to demonstrate that the ZnO/UNCD SAW device can be successfully used for microfluidic applications. Streaming, pumping, and jetting using microdroplets of 0.5 and 20 μl were achieved and characterized under different powers applied to the SAW device, focusing more on the jetting behaviors induced by the ZnO SAW.  相似文献   

20.
Rapid concentration and detection of bacteria in integrated chips and microfluidic devices is needed for the advancement of lab-on-a-chip devices because current detection methods require high concentrations of bacteria which render them impractical. We present a new chip-scale rapid bacteria concentration technique combined with surface-enhanced Raman scattering (SERS) to enhance the detection of low bacteria count samples. This concentration technique relies on convection by a long-range converging vortex to concentrate the bacteria into a packed mound of 200 μm in diameter within 15 min. Concentration of bioparticle samples as low as 104 colony forming units (CFU)∕ml are presented using batch volumes as large as 150 μl. Mixtures of silver nanoparticles with Saccharomyces cerevisiae, Escherichia coli F-amp, and Bacillus subtilis produce distinct and noticeably different Raman spectra, illustrating that this technique can be used as a detection and identification tool.  相似文献   

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