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Dec 2011

Volume 5, Issue 4, Articles (04xxxx)

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Biomicrofluidics 5, 044119 (2011); http://dx.doi.org/10.1063/1.3672190 (12 pages)

Qianqian Cao, Chuncheng Zuo, Lujuan Li, and Guang Yan
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The microfluidic system for studies of carcinoma and normal cells interactions after photodynamic therapy (PDT) procedures

Elzbieta Jedrych, Michal Chudy, Artur Dybko, and Zbigniew Brzozka

Biomicrofluidics 5, 041101 (2011); http://dx.doi.org/10.1063/1.3658842 (6 pages)

Online Publication Date: 11 November 2011

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This study reports on the use of a microsystem for evaluation of photodynamic therapy (PDT) procedures on the “mixed” (carcinoma-normal) cultures. Balb/3T3 (normal mouse embryo) and A549 (human lung carcinoma) cells were tested in separated and “mixed” cultures. Interactions and migration of cells cultured together were observed. The PDT procedures were examined in the hybrid (PDMS/glass) microsystem which contains cell culture microchambers integrated with network of microchannels. We investigated that the number of dead cells after PDT procedures is dependent on the kind of cell culture. Moreover, the influence of the carcinoma cells on the viability of normal cells in the “mixed” culture was observed.
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87.50.wp Therapeutic applications
87.80.Ek Mechanical and micromechanical techniques
87.85.Ox Biomedical instrumentation and transducers, including micro-electro-mechanical systems (MEMS)
85.85.+j Micro- and nano-electromechanical systems (MEMS/NEMS) and devices
87.17.Uv Biotechnology of cell processes
87.19.xj Cancer
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Three-dimensional cellular focusing utilizing a combination of insulator-based and metallic dielectrophoresis

Ching-Te Huang, Cheng-Hsin Weng, and Chun-Ping Jen

Biomicrofluidics 5, 044101 (2011); http://dx.doi.org/10.1063/1.3646757 (11 pages) | Cited 1 time

Online Publication Date: 3 October 2011

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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.
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87.80.Ek Mechanical and micromechanical techniques
02.60.-x Numerical approximation and analysis
07.10.Cm Micromechanical devices and systems
82.45.-h Electrochemistry and electrophoresis
87.16.-b Subcellular structure and processes

Effect of wall permittivity on electroviscous flow through a contraction

J. D. Berry, M. R. Davidson, R. P. Bharti, and D. J. E. Harvie

Biomicrofluidics 5, 044102 (2011); http://dx.doi.org/10.1063/1.3645194 (17 pages)

Online Publication Date: 12 October 2011

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The electroviscous flow at low Reynolds number through a two-dimensional slit contraction with electric double-layer overlap is investigated numerically for cases where the permittivity of the wall material is significant in comparison with the permittivity of the liquid. The liquid-solid interface is assumed to have uniform surface-charge density. It is demonstrated that a finite wall permittivity has a marked effect on the distribution of ions in and around the contraction, with a significant build-up of counter-ions observed at the back-step. The development length of the flow increases substantially as the wall permittivity becomes significant, meaning that the electric double-layers require a longer distance to develop within the contraction. Consequently, there is a corresponding decrease in the hydrodynamic and electro-potential resistance caused by the contraction. The effect of wall-region width on the flow characteristics is also quantified, demonstrating that the development length increases with increasing wall-region width for widths up to 5 channel widths.
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47.65.Gx Electrorheological fluids
66.20.-d Viscosity of liquids; diffusive momentum transport
47.15.Rq Laminar flows in cavities, channels, ducts, and conduits
47.60.Dx Flows in ducts and channels

Microfluidic droplet encapsulation of highly motile single zoospores for phenotypic screening of an antioomycete chemical

Haifeng Yang, Xuan Qiao, Madan K. Bhattacharyya, and Liang Dong

Biomicrofluidics 5, 044103 (2011); http://dx.doi.org/10.1063/1.3651620 (11 pages)

Online Publication Date: 13 October 2011

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Highly motile Phytophthora sojae (P. sojae) zoospores of an oomycete plant pathogen and antioomycete candidate chemicals were encapsulated into microdroplets. Random fast self-motion of P. sojae zoospores was overcome by choosing an appropriate flow rate for a zoospore suspension. To influence stochastic loading of zoospores into a microfluidic channel, a zoospore suspension was directly preloaded into a microtubing with a largely reduced inner diameter. A relatively high single zoospore encapsulation rate of 60.5% was achieved on a most trivial T-junction droplet generator platform, without involving any specially designed channel geometry. We speculated that spatial reduction in the diameter direction of microtubing added a degree of zoospore ordering in the longitudinal direction of microtubing and thus influenced positively to change the inherent limitation of stochastic encapsulation of zoospores. Comparative phenotypic study of a plant oomycete pathogen at a single zoospore level had not been achieved earlier. Phenotypic changes of zoospores responding to various chemical concentration conditions were measured in multiple droplets in parallel, providing a reliable data set and thus an improved statistic at a low chemical consumption. Since each droplet compartment contained a single zoospore, we were able to track the germinating history of individual zoospores without being interfered by other germinating zoospores, achieving a high spatial resolution. By adapting some existing droplet immobilization and concentration gradient generation techniques, the droplet approach could potentially lead to a medium-to-high throughput, reliable screening assay for chemicals against many other highly motile zoospores of pathogens.
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87.80.Ek Mechanical and micromechanical techniques
47.63.Gd Swimming microorganisms
47.85.Np Fluidics
85.85.+j Micro- and nano-electromechanical systems (MEMS/NEMS) and devices
87.15.R- Reactions and kinetics

Transportation of single cell and microbubbles by phase-shift introduced to standing leaky surface acoustic waves

Long Meng, Feiyan Cai, Zidong Zhang, Lili Niu, Qiaofeng Jin, Fei Yan, Junru Wu, Zhanhui Wang, and Hairong Zheng

Biomicrofluidics 5, 044104 (2011); http://dx.doi.org/10.1063/1.3652872 (10 pages) | Cited 2 times

Online Publication Date: 20 October 2011

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A microfluidic device was developed to precisely transport a single cell or multiple microbubbles by introducing phase-shifts to a standing leaky surface acoustic wave (SLSAW). The device consists of a polydimethyl-siloxane (PDMS) microchannel and two phase-tunable interdigital transducers (IDTs) for the generation of the relative phase for the pair of surface acoustic waves (SAW) propagating along the opposite directions forming a standing wave. When the SAW contacts the fluid medium inside the microchannel, some of SAW energy is coupled to the fluid and the SAW becomes the leaky surface wave. By modulating the relative phase between two IDTs, the positions of pressure nodes of the SLSAW in the microchannel change linearly resulting in the transportation of a single cell or microbubbles. The results also reveal that there is a good linear relationship between the relative phase and the displacement of a single cell or microbubbles. Furthermore, the single cell and the microbubbles can be transported over a predetermined distance continuously until they reach the targeted locations. This technique has its distinct advantages, such as precise position-manipulation, simple to implement, miniature size, and noninvasive character, which may provide an effective method for the position-manipulation of a single cell and microbubbles in many biological and biomedical applications.
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87.85.Ox Biomedical instrumentation and transducers, including micro-electro-mechanical systems (MEMS)
87.17.-d Cell processes
43.38.Rh Surface acoustic wave transducers

An insulator-based dielectrophoretic microdevice for the simultaneous filtration and focusing of biological cells

Chun-Ping Jen and Wei-Fu Chen

Biomicrofluidics 5, 044105 (2011); http://dx.doi.org/10.1063/1.3658644 (11 pages) | Cited 2 times

Online Publication Date: 31 October 2011

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Manipulating and discriminating biological cells of interest using microfluidic and micro total analysis system (μTAS) devices have potential applications in clinical diagnosis and medicine. Cellular focusing in microfluidic devices is a prerequisite for medical applications, such as cell sorting, cell counting, or flow cytometry. In the present study, an insulator-based dielectrophoretic microdevice is designed for the simultaneous filtration and focusing of biological cells. The cells are introduced into the microchannel and hydrodynamically pre-confined by funnel-shaped insulating structures close to the inlet. There are ten sets of X-patterned insulating structures in the microfluidic channel. The main function of the first five sets of insulating structures is to guide the cells by negative dielectrophoretic responses (viable HeLa cells) into the center region of the microchannel. The positive dielectrophoretic cells (dead HeLa cells) are attracted to regions with a high electric-field gradient generated at the edges of the insulating structures. The remaining five sets of insulating structures are mainly used to focus negative dielectrophoretic cells that have escaped from the upstream region. Experiments employing a mixture of dead and viable HeLa cells are conducted to demonstrate the effectiveness of the proposed design. The results indicate that the performance of both filtration and focusing improves with the increasing strength of the applied electric field and a decreasing inlet sample flow rate, which agrees with the trend predicted by the numerical simulations. The filtration efficiency, which is quantitatively investigated, is up to 88% at an applied voltage of 50 V peak-to-peak (1 kHz) and a sample flow rate of 0.5 μl/min. The proposed device can focus viable cells into a single file using a voltage of 35 V peak-to-peak (1 kHz) at a sample flow rate of 1.0 μl/min.
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87.85.Ox Biomedical instrumentation and transducers, including micro-electro-mechanical systems (MEMS)
47.63.mh Transport processes and drug delivery
85.85.+j Micro- and nano-electromechanical systems (MEMS/NEMS) and devices
87.17.-d Cell processes
47.60.Dx Flows in ducts and channels
87.80.Kc Electrochemical techniques

Simulation of conformational preconditioning strategies for electrophoretic stretching of DNA in a microcontraction

Chih-Chen Hsieh and Tsung-Hsien Lin

Biomicrofluidics 5, 044106 (2011); http://dx.doi.org/10.1063/1.3655565 (17 pages)

Online Publication Date: 10 November 2011

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We have used Brownian dynamics-finite element method to examine two conformational preconditioning approaches for improving DNA stretching in a microcontraction for the purpose of direct gene analysis. The newly proposed “pre-stretching” strategy is found to significantly improve the degree of DNA extension at the exit of the contraction. On the other hand, applying an oscillating extensional field to DNA yields no preconditioning effect. Detailed analysis of the evolution of DNA extension and conformation reveals that the success of our “pre-stretching” strategy relies on the “non-local” effect that cannot be predicted using simple kinematics analysis. In other words, accurate prediction can only be obtained using detailed simulations. Comparing to the existing preconditioning strategies, our “pre-stretching” method is easy to implement while still providing a very good performance. We hope that the insight gained from this study can be useful for future design of biomicrofluidic devices for DNA manipulation.
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87.15.A- Theory, modeling, and computer simulation
05.40.Jc Brownian motion
36.20.Hb Configuration (bonds, dimensions)
87.15.Tt Electrophoresis
87.10.Kn Finite element calculations
87.14.gk DNA

Manipulating particle trajectories with phase-control in surface acoustic wave microfluidics

Nathan D. Orloff, Jaclyn R. Dennis, Marco Cecchini, Ethan Schonbrun, Eduard Rocas, Yu Wang, David Novotny, Raymond W. Simmonds, John Moreland, Ichiro Takeuchi, and James C. Booth

Biomicrofluidics 5, 044107 (2011); http://dx.doi.org/10.1063/1.3661129 (9 pages) | Cited 2 times

Online Publication Date: 14 November 2011

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We present a 91 MHz surface acoustic wave resonator with integrated microfluidics that includes a flow focus, an expansion region, and a binning region in order to manipulate particle trajectories. We demonstrate the ability to change the position of the acoustic nodes by varying the electronic phase of one of the transducers relative to the other in a pseudo-static manner. The measurements were performed at room temperature with 3 μm diameter latex beads dispersed in a water-based solution. We demonstrate the dependence of nodal position on pseudo-static phase and show simultaneous control of 9 bead streams with spatial control of −0.058 μm/deg ± 0.001 μm/deg. As a consequence of changing the position of bead streams perpendicular to their flow direction, we also show that the integrated acoustic-microfluidic device can be used to change the trajectory of a bead stream towards a selected bin with an angular control of 0.008 deg/deg ± 0.000(2) deg/deg.
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87.80.Ek Mechanical and micromechanical techniques
43.38.Rh Surface acoustic wave transducers
85.85.+j Micro- and nano-electromechanical systems (MEMS/NEMS) and devices

Contrast agent-free sonoporation: The use of an ultrasonic standing wave microfluidic system for the delivery of pharmaceutical agents

Dario Carugo, Dyan N. Ankrett, Peter Glynne-Jones, Lorenzo Capretto, Rosemary J. Boltryk, Xunli Zhang, Paul A. Townsend, and Martyn Hill

Biomicrofluidics 5, 044108 (2011); http://dx.doi.org/10.1063/1.3660352 (15 pages)

Online Publication Date: 15 November 2011

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Sonoporation is a useful biophysical mechanism for facilitating the transmembrane delivery of therapeutic agents from the extracellular to the intracellular milieu. Conventionally, sonoporation is carried out in the presence of ultrasound contrast agents, which are known to greatly enhance transient poration of biological cell membranes. However, in vivo contrast agents have been observed to induce capillary rupture and haemorrhage due to endothelial cell damage and to greatly increase the potential for cell lysis in vitro. Here, we demonstrate sonoporation of cardiac myoblasts in the absence of contrast agent (CA-free sonoporation) using a low-cost ultrasound-microfluidic device. Within this device an ultrasonic standing wave was generated, allowing control over the position of the cells and the strength of the acoustic radiation forces. Real-time single-cell analysis and retrospective post-sonication analysis of insonated cardiac myoblasts showed that CA-free sonoporation induced transmembrane transfer of fluorescent probes (CMFDA and FITC-dextran) and that different mechanisms potentially contribute to membrane poration in the presence of an ultrasonic wave. Additionally, to the best of our knowledge, we have shown for the first time that sonoporation induces increased cell cytotoxicity as a consequence of CA-free ultrasound-facilitated uptake of pharmaceutical agents (doxorubicin, luteolin, and apigenin). The US-microfluidic device designed here provides an in vitro alternative to expensive and controversial in vivo models used for early stage drug discovery, and drug delivery programs and toxicity measurements.
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87.50.yg Biophysical mechanisms of interaction
87.16.dp Transport, including channels, pores, and lateral diffusion

Refinement of the theory for extracting cell dielectric properties from dielectrophoresis and electrorotation experiments

U. Lei, Pei-Hou Sun, and Ronald Pethig

Biomicrofluidics 5, 044109 (2011); http://dx.doi.org/10.1063/1.3659282 (16 pages)

Online Publication Date: 17 November 2011

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A modified theory is proposed for extracting cell dielectric properties from the peak frequency measurement of electrorotation (ER) and the crossover frequency measurement of dielectrophoresis (DEP). Current theory in the literature is based on the low frequency (DC) approximations for the equivalent cell permittivity and conductivity, which are valid when the measurements are performed in a medium with conductivity less than 1 mS/m. The present theory extracts the cell properties through optimizing an expression for the medium conductivity in terms of the peak ER, or DEP crossover, frequency according to its definition using full expressions of equivalent cell permittivity and conductivity. Various levels of approximation of the theory are proposed and discussed through a scaling analysis. The present theory can extract both membrane and interior properties from the low and the high peak ER, or DEP crossover, frequencies for any medium conductivity provided the peak ER, or DEP crossover, frequency exists. It can be reduced to the linear theory for the low peak ER and DEP crossover frequencies in the literature when the medium conductivity is less than 10 mS/m. However, we can determine the membrane capacitance and conductance via the slope and intercept, respectively, of the straight line fitting of the ER peak and DEP frequency against medium conductivity data according to the linear theory only when the intercept dominates the experimental uncertainty, which occurs when the medium conductivity is less than 1 mS/m in practice.
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87.16.D- Membranes, bilayers, and vesicles
82.45.-h Electrochemistry and electrophoresis

Optofluidic membrane interferometer: An imaging method for measuring microfluidic pressure and flow rate simultaneously on a chip

Wuzhou Song and Demetri Psaltis

Biomicrofluidics 5, 044110 (2011); http://dx.doi.org/10.1063/1.3664693 (11 pages)

Online Publication Date: 30 November 2011

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We present a novel image-based method to measure the on-chip microfluidic pressure and flow rate simultaneously by using the integrated optofluidic membrane interferometers (OMIs). The device was constructed with two layers of structured polydimethylsiloxane (PDMS) on a glass substrate by multilayer soft lithography. The OMI consists of a flexible air-gap optical cavity which upon illumination by monochromatic light generates interference patterns that depends on the pressure. These interference patterns were captured with a microscope and analyzed by computer based on a pattern recognition algorithm. Compared with the previous techniques for pressure sensing, this method offers several advantages including low cost, simple fabrication, large dynamic range, and high sensitivity. For pressure sensing, we demonstrate a dynamic range of 0-10 psi with an accuracy of ±2% of full scale. Since multiple OMIs can be integrated into a single chip for detecting pressures at multiple locations simultaneously, we also demonstrated a microfluidic flow sensing by measuring the differential pressure along a channel. Thanks to the simple fabrication that is compatible with normal microfluidics, such OMIs can be easily integrated into other microfluidic systems for in situ fluid monitoring.
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87.80.Ek Mechanical and micromechanical techniques
85.85.+j Micro- and nano-electromechanical systems (MEMS/NEMS) and devices
07.10.Cm Micromechanical devices and systems
07.07.Df Sensors (chemical, optical, electrical, movement, gas, etc.); remote sensing
07.60.Ly Interferometers
42.82.Cr Fabrication techniques; lithography, pattern transfer

Microfluidic concentration of bacteria by on-chip electrophoresis

Dietmar Puchberger-Enengl, Susann Podszun, Helene Heinz, Carsten Hermann, Paul Vulto, and Gerald A. Urban

Biomicrofluidics 5, 044111 (2011); http://dx.doi.org/10.1063/1.3664691 (10 pages)

Online Publication Date: 2 December 2011

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In this contribution, we present a system for efficient preconcentration of pathogens without affecting their viability. Development of miniaturized molecular diagnostic kits requires concentration of the sample, molecule extraction, amplification, and detection. In consequence of low analyte concentrations in real-world samples, preconcentration is a critical step within this workflow. Bacteria and viruses exhibit a negative surface charge and thus can be electrophoretically captured from a continuous flow. The concept of phaseguides was applied to define gel membranes, which enable effective and reversible collection of the target species. E. coli of the strains XL1-blue and K12 were used to evaluate the performance of the device. By suppression of the electroosmotic flow both strains were captured with efficiencies of up to 99%. At a continuous flow of 15 μl/min concentration factors of 50.17 ± 2.23 and 47.36 ± 1.72 were achieved in less than 27 min for XL1-blue and K12, respectively. These results indicate that free flow electrophoresis enables efficient concentration of bacteria and the presented device can contribute to rapid analyses of swab-derived samples.
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87.80.Ek Mechanical and micromechanical techniques
87.15.Tt Electrophoresis
85.85.+j Micro- and nano-electromechanical systems (MEMS/NEMS) and devices

Influences of electric field on living cells in a charged water-in-oil droplet under electrophoretic actuation

Do Jin Im, Jihoon Noh, Nam Woo Yi, Jaesung Park, and In Seok Kang

Biomicrofluidics 5, 044112 (2011); http://dx.doi.org/10.1063/1.3665222 (10 pages)

Online Publication Date: 2 December 2011

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We experimentally investigate the effects of high electric field on living cells inside a charged droplet under electrophoretic actuation. When an aqueous droplet suspended in a dielectric liquid contacts with electrified electrode, the droplet acquires charge. This charged droplet undergoes electrophoretic motion under strong electric field (1–3 kV/cm), which can be used as a droplet manipulation method in biomicrofluidic applications. However, because strong electric field and use of dielectric oil can be a harmful environment for living cells, the biological feasibilities have been tested. Trypan blue test and cell growth test have been performed to check the viability and proliferation of cells in a droplet under various electric field strengths and actuation times. We have not observed any noticeable influence of electric field and silicone oil on the viability and proliferation of cells, which indicates that electrophoresis could be safely used as a manipulation method for a droplet containing living biological system.
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87.50.ch Electrophoresis/dielectrophoresis and other mechanical effects
87.80.Ek Mechanical and micromechanical techniques
47.85.Np Fluidics
87.17.Ee Growth and division

Stable, biocompatible lipid vesicle generation by solvent extraction-based droplet microfluidics

Shia-Yen Teh, Ruba Khnouf, Hugh Fan, and Abraham P. Lee

Biomicrofluidics 5, 044113 (2011); http://dx.doi.org/10.1063/1.3665221 (12 pages)

Online Publication Date: 9 December 2011

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In this paper, we present a microfluidic platform for the continuous generation of stable, monodisperse lipid vesicles 20–110 μm in diameter. Our approach utilizes a microfluidic flow-focusing droplet generation design to control the vesicle size by altering the system’s fluid flow rates to generate vesicles with narrow size distribution. Double emulsions are first produced in consecutive flow-focusing channel geometries and lipid membranes are then formed through a controlled solvent extraction process. Since no strong solvents are used in the process, our method allows for the safe encapsulation and manipulation of an assortment of biological entities, including cells, proteins, and nucleic acids. The vesicles generated by this method are stable and have a shelf life of at least 3 months. Here, we demonstrate the cell-free in vitro synthesis of proteins within lipid vesicles as an initial step towards the development of an artificial cell.
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87.16.dt Structure, static correlations, domains, and rafts
87.80.Ek Mechanical and micromechanical techniques
07.10.Cm Micromechanical devices and systems
82.70.Kj Emulsions and suspensions
87.14.E- Proteins

Characterization of low viscosity polymer solutions for microchip electrophoresis of non-denatured proteins on plastic chips

Takao Yasui, Mohamad Reza Mohamadi, Noritada Kaji, Yukihiro Okamoto, Manabu Tokeshi, and Yoshinobu Baba

Biomicrofluidics 5, 044114 (2011); http://dx.doi.org/10.1063/1.3668233 (9 pages)

Online Publication Date: 12 December 2011

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In this paper, we study characteristics of polymers (methylcellulose, hypromellose ((hydroxypropyl)methyl cellulose), poly(vinylpyrrolidone), and poly(vinyl alcohol)) with different chemical structures for microchip electrophoresis of non-denatured protein samples in a plastic microchip made of poly(methyl methacrylate) (PMMA). Coating efficiency of these polymers for controlling protein adsorption onto the channel surface of the plastic microchip, wettability of the PMMA surface, and electroosmotic flow in the PMMA microchannels in the presence of these polymers were compared. Also relative electrophoretic mobility of protein samples in solutions of these polymers was studied. We showed that when using low polymer concentrations (lower than the polymer entanglement point) where the sieving effect is substantially negligible, the interaction of the samples with the polymer affected the electrophoretic mobility of the samples. This effect can be used for achieving better resolution in microchip electrophoresis of protein samples.
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87.15.km Protein-protein interactions
87.15.B- Structure of biomolecules
87.15.R- Reactions and kinetics
82.45.-h Electrochemistry and electrophoresis
87.15.Tt Electrophoresis

A parallel microfluidic channel fixture fabricated using laser ablated plastic laminates for electrochemical and chemiluminescent biodetection of DNA

Thayne L. Edwards, Jason C. Harper, Ronen Polsky, DeAnna M. Lopez, David R. Wheeler, Amy C. Allen, and Susan M. Brozik

Biomicrofluidics 5, 044115 (2011); http://dx.doi.org/10.1063/1.3664694 (14 pages)

Online Publication Date: 15 December 2011

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Herein is described the fabrication and use of a plastic multilayer 3-channel microfluidic fixture. Multilayer devices were produced by laser machining of plastic polymethylmethacrylate and polyethyleneterapthalate laminates by ablation. The fixture consisted of an array of nine individually addressable gold or gold/ITO working electrodes, and a resistive platinum heating element. Laser machining of both the fluidic pathways in the plastic laminates, and the stencil masks used for thermal evaporation to form electrode regions on the plastic laminates, enabled rapid and inexpensive implementation of design changes. Electrochemiluminescence reactions in the fixture were achieved and monitored through ITO electrodes. Electroaddressable aryl diazonium chemistry was employed to selectively pattern gold electrodes for electrochemical multianalyte DNA detection from double stranded DNA (dsDNA) samples. Electrochemical detection of dsDNA was achieved by melting of dsDNA molecules in solution with the integrated heater, allowing detection of DNA sequences specific to breast and colorectal cancers with a non-specific binding control. Following detection, the array surface could be renewed via high temperature (95 °C) stripping using the integrated heating element. This versatile and simple method for prototyping devices shows potential for further development of highly integrated, multi-functional bioanalytical devices.
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87.85.Ox Biomedical instrumentation and transducers, including micro-electro-mechanical systems (MEMS)
87.80.Kc Electrochemical techniques
87.85.Va Micromachining
85.85.+j Micro- and nano-electromechanical systems (MEMS/NEMS) and devices
82.47.Rs Electrochemical sensors
87.14.gk DNA

Effect of pulse direct current signals on electrotactic movement of nematodes Caenorhabditis elegans and Caenorhabditis briggsae

Pouya Rezai, Sangeena Salam, Ponnambalam Ravi Selvaganapathy, and Bhagwati P. Gupta

Biomicrofluidics 5, 044116 (2011); http://dx.doi.org/10.1063/1.3665224 (9 pages)

Online Publication Date: 15 December 2011

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The nematodes (worms) Caenorhabditis elegans and Caenorhabditis briggsae are well-known model organisms to study the basis of animal development and behaviour. Their sinusoidal pattern of movement is highly stereotypic and serves as a tool to monitor defects in neurons and muscles that control movement. Until recently, a simple yet robust method to initiate movement response on-demand did not exist. We have found that the electrical stimulation in a microfluidic channel, using constant DC electric field, induces movement (termed electrotaxis) that is instantaneous, precise, sensitive, and fully penetrant. We have further characterized this behaviour and, in this paper, demonstrate that electrotaxis can also be induced using a pulse DC electric signal. Worms responded to pulse DC signals with as low as 30% duty cycle by moving towards the negative electrode at the same speed as constant DC fields (average speed of C. elegans = 296 ± 43 μm/s and C. briggsae = 356 ± 20 μm/s, for both constant and pulse DC electric fields with various frequencies). C. briggsae was found to be more sensitive to electric signals compared to C. elegans. We also investigated the turning response of worms to a change in the direction of constant and pulse DC signals. The response for constant DC signal was found to be instantaneous and similar for most worms. However, in the case of pulse DC signal, alterations in duty cycle affected the turning response time as well as the number of responding worms. Our findings show that pulse DC method allows quantitative measurement of response behaviour of worms and suggest that it could be used as a tool to study the neuronal basis of such a behaviour that is not observed under constant DC conditions.
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87.19.rs Movement
87.19.ld Electrodynamics in the nervous system
87.19.Ff Muscles

Microwell perfusion array for high-throughput, long-term imaging of clonal growth

Huaying Chen, Jingjing Li, Han Zhang, Musen Li, Gary Rosengarten, and Robert E. Nordon

Biomicrofluidics 5, 044117 (2011); http://dx.doi.org/10.1063/1.3669371 (13 pages)

Online Publication Date: 15 December 2011

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Continuous cell tracking by time-lapse microscopy has led to detailed study of cell differentiation pathways using single cell fate maps. There are a multitude of cell fate outcomes, so hundreds of clonal division histories are required to measure these stochastic branching processes. This study examines the principle of condensing cell imaging information into a relatively small region to maximize live cell imaging throughput. High throughput clonal analysis of non-adherent cells by continuous live cell tracking was possible using a microwell perfusion array with an internal volume of 16 μl and 600 microwells at the base. This study includes examination of biocompatibility of buffer systems, connecting tubing, cell culture substrates, and media degradation. An intermittent perfusion protocol was selected for long-term time-lapse imaging of KG1a cells in the microwell array; 1500 clones were simultaneously cultured and scanned every 3 min at 100 × magnifications for 6 days. The advantages of perfusion microwell culture are continuous long-term cell tracking, higher cell imaging throughput, and greater control over cell microenvironment. Microwell devices facilitate high throughput analysis of cell lineage development and measurement of the probability distribution for cell life events such as mitosis.
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87.17.Uv Biotechnology of cell processes
02.50.Cw Probability theory
02.50.Ey Stochastic processes
87.80.-y Biophysical techniques (research methods)

Growth propagation of yeast in linear arrays of microfluidic chambers over many generations

Li Wang, Jiaji Liu, Xin Li, Jian Shi, Jie Hu, Ran Cui, Zhi-Ling Zhang, Dai-Wen Pang, and Yong Chen

Biomicrofluidics 5, 044118 (2011); http://dx.doi.org/10.1063/1.3668243 (9 pages)

Online Publication Date: 16 December 2011

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The growth of microorganisms is often confined in restricting geometries. In this work, we designed a device to study the growth propagation of budding yeast along linear arrays of microfluidic chambers. Vacuum assisted cell loading was used to seed cells of limited numbers in the up-most chambers of each linear array. Once loaded, cells grow until confluent and then overgrow, pushing some of the newborns into the neighboring downstream chamber through connection channels. Such a scenario repeats sequentially along the whole linear chamber arrays. We observed that the propagation speed of yeast population along the linear arrays was strongly channel geometry dependent. When the connection channel is narrow and long, the amount of cells delivered into the downstream chamber is small so that cells grow over several generations in the same chamber before passing into the next chamber. Consequently, a population growth of more than 50 generations could be observed along a single linear array. We also provided a mathematical model to quantitatively interpret the observed growth dynamics.
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87.85.gf Fluid mechanics and rheology
07.10.Cm Micromechanical devices and systems
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Effects of chain stiffness and salt concentration on responses of polyelectrolyte brushes under external electric field

Qianqian Cao, Chuncheng Zuo, Lujuan Li, and Guang Yan

Biomicrofluidics 5, 044119 (2011); http://dx.doi.org/10.1063/1.3672190 (12 pages) | Cited 1 time

Online Publication Date: 21 December 2011

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We report a molecular dynamics study on non-equilibrium dynamics of polyelectrolyte brushes under external electric fields. In this work, the effects of chain stiffness and salt concentration on static and dynamic responses of the brushes are addressed in detail. Our simulations indicate that varying these parameters induce rich electro-responsive behavior of the brushes. The increase of salt concentration results in the enhancement of an opposite electric field formed by non-equilibrium distribution of cations and anions, which resists stretching or shrinkage of grafted chains. At strong positive electric fields, the flexible brushes are more sensitive to the change of salt concentration. When reversing the electric field, the stiff brushes undergo a conformational transition from collapse to complete stretching. At high salt concentrations, dynamic responsive magnitude of the brush thickness to added electric field is strongly reduced. It was found that the fall time for the stiff brush becomes much shorter than that for the flexible brush. Additionally, increasing ion concentration leads to an excess extension or shrinkage of flexible brushes. For strongly stiff brushes, such phenomenon occurs in the presence or absence of salt.
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87.15.rp Polymerization
87.15.ap Molecular dynamics simulation
02.60.-x Numerical approximation and analysis
87.50.C- Static and low-frequency electric and magnetic fields effects
82.35.Rs Polyelectrolytes

Asymmetry of red blood cell motions in a microchannel with a diverging and converging bifurcation

Vladimir Leble, Rui Lima, Ricardo Dias, Carla Fernandes, Takuji Ishikawa, Yohsuke Imai, and Takami Yamaguchi

Biomicrofluidics 5, 044120 (2011); http://dx.doi.org/10.1063/1.3672689 (15 pages) | Cited 1 time

Online Publication Date: 23 December 2011

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In microcirculation, red blood cells (RBCs) flowing through bifurcations may deform considerably due to combination of different phenomena that happen at the micro-scale level, such as: attraction effect, high shear, and extensional stress, all of which may influence the rheological properties and flow behavior of blood. Thus, it is important to investigate in detail the behavior of blood flow occurring at both bifurcations and confluences. In the present paper, by using a micro-PTV system, we investigated the variations of velocity profiles of two working fluids flowing through diverging and converging bifurcations, human red blood cells suspended in dextran 40 with about 14% of hematocrit level (14 Hct) and pure water seeded with fluorescent trace particles. All the measurements were performed in the center plane of rectangular microchannels using a constant flow rate of about 3.0 × 10−12 m3/s. Moreover, the experimental data was compared with numerical results obtained for Newtonian incompressible fluid. The behavior of RBCs was asymmetric at the divergent and convergent side of the geometry, whereas the velocities of tracer particles suspended in pure water were symmetric and well described by numerical simulation. The formation of a red cell-depleted zone immediately downstream of the apex of the converging bifurcation was observed and its effect on velocity profiles of RBCs flow has been investigated. Conversely, a cell-depleted region was not formed around the apex of the diverging bifurcation and as a result the adhesion of RBCs to the wall surface was enhanced in this region.
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87.85.gf Fluid mechanics and rheology
02.60.-x Numerical approximation and analysis
87.17.Rt Cell adhesion and cell mechanics
87.19.U- Hemodynamics
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Membrane-integrated microfluidic device for high-resolution live cell imaging

Alla A. Epshteyn, Steven Maher, Amy J. Taylor, Angela B. Holton, Jeffrey T. Borenstein, and Joseph D. Cuiffi

Biomicrofluidics 5, 046501 (2011); http://dx.doi.org/10.1063/1.3647824 (6 pages)

Online Publication Date: 17 October 2011

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The design and fabrication of a membrane-integrated microfluidic cell culture device (five layers,≤500 μm total thickness) developed for high resolution microscopy is reported here. The multi-layer device was constructed to enable membrane separated cell culture for tissue mimetic in vitro model applications and pharmacodynamic evaluation studies. The microdevice was developed via a unique combination of low profile fluidic interconnect design, substrate transfer methodology, and wet silane bonding. To demonstrate the unique high resolution imaging capability of this device, we used oil immersion microscopy to image stained nuclei and mitochondria in primary hepatocytes adhered to the incorporated membrane
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87.80.Ek Mechanical and micromechanical techniques
87.16.D- Membranes, bilayers, and vesicles
87.16.-b Subcellular structure and processes

Microscale pH regulation by splitting water

Li-Jing Cheng and Hsueh-Chia Chang

Biomicrofluidics 5, 046502 (2011); http://dx.doi.org/10.1063/1.3657928 (8 pages) | Cited 1 time

Online Publication Date: 2 November 2011

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We present a simple, flexible approach for pH regulation in micro-chambers by injecting controllable amounts of protons and hydroxide ions via field-enhanced dissociation of water molecules. Under a DC voltage bias, the polymeric bipolar membranes integrated in microfluidics devices generate and separate H+ and OH− ions without gas production or contaminant generation resulting from electron-transfer reactions. Robust local on-chip pH and pH gradients are sustained with no need of additional acidic/basic solutions that dilute analyte concentrations. The method could provide a better strategy for pH control in microfluidics.
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87.80.Ek Mechanical and micromechanical techniques
47.85.Np Fluidics
82.30.Fi Ion-molecule, ion-ion, and charge-transfer reactions
82.30.Lp Decomposition reactions (pyrolysis, dissociation, and fragmentation)
82.45.Tv Bioelectrochemistry
85.85.+j Micro- and nano-electromechanical systems (MEMS/NEMS) and devices

Sealing SU-8 microfluidic channels using PDMS

Zhiyi Zhang, Ping Zhao, Gaozhi Xiao, Benjamin R. Watts, and Changqing Xu

Biomicrofluidics 5, 046503 (2011); http://dx.doi.org/10.1063/1.3659016 (8 pages)

Online Publication Date: 9 November 2011

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A simple method of irreversibly sealing SU-8 microfluidic channels using PDMS is reported in this paper. The method is based on inducing a chemical reaction between PDMS and SU-8 by first generating amino groups on PDMS surface using N2 plasma treatment, then allowing the amino groups to react with the residual epoxy groups on SU-8 surface at an elevated temperature. The N2 plasma treatment of PDMS can be conducted using an ordinary plasma chamber and high purity N2, while the residual epoxy groups on SU-8 surface can be preserved by post-exposure baking SU-8 at a temperature no higher than 95 °C. The resultant chemical bonding between PDMS and SU-8 using the method create an interface that can withstand a stress that is greater than the bulk strength of PDMS. The bond is permanent and is long-term resistant to water. The method was applied in fabricating SU-8 microfluidi-photonic integrated devices, and the obtained devices were tested to show desirable performance.
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87.80.Ek Mechanical and micromechanical techniques
42.82.-m Integrated optics
85.85.+j Micro- and nano-electromechanical systems (MEMS/NEMS) and devices
87.17.-d Cell processes
87.50.wf Biophysical mechanisms of interaction

The effects of laser welding on heterogeneous immunoassay performance in a microfluidic cartridge

Anne Mäntymaa, Jussi Halme, Lasse Välimaa, and Pasi Kallio

Biomicrofluidics 5, 046504 (2011); http://dx.doi.org/10.1063/1.3668261 (11 pages)

Online Publication Date: 12 December 2011

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Sealing of a microfluidic cartridge is a challenge, because the cartridge commonly contains heat-sensitive biomolecules that must also be protected from contamination. In addition, the objective is usually to obtain a sealing method suitable for mass production. Laser welding is a rapid technique that can be accomplished with low unit costs. Even though the technique has been widely adopted in industry, the literature on its use in microfluidic applications is not large. This paper is the first to report the effects of laser welding on the performance of the heterogeneous immunoassay in a polystyrene microfluidic cartridge in which biomolecules are immobilized into the reaction surface of the cartridge before sealing. The paper compares the immunoassay performance of microfluidic cartridges that are sealed either with an adhesive tape or by use of laser transmission welding. The model analyte used is thyroid stimulating hormone (TSH). The results show that the concentration curves in the laser-welded cartridges are very close to the curves in the taped cartridges. This indicates, first, that laser welding does not cause any significant reduction in immunoassay performance, and second, that the polystyrene cover does not have significant effect on the signal levels. Interestingly, the coefficients of variance between parallel samples were lower in the laser-welded cartridges than in the taped cartridges.
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87.80.Ek Mechanical and micromechanical techniques
85.85.+j Micro- and nano-electromechanical systems (MEMS/NEMS) and devices
87.80.Qk Biochemical separation processes
87.15.R- Reactions and kinetics
42.62.Cf Industrial applications
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