Andrew M. Wo

Reduction of unsteady blade loading by beneficial use of vortical and potential disturbances in an axial compressor with rotor clocking

This paper demonstrates reduction of stator unsteady loading due to forced response in a large-scale, low-speed, rotor/stator/rotor axial compressor rig by clocking the downstream rotor. Data from the rotor/stator configuration showed that the stator response due to the upstream vortical disturbance reaches a maximum when the wake impinges against the suction surface immediately downstream of the leading edge. Results from the stator/rotor configuration revealed that the stator response due to the downstream potential disturbance reaches a minimum with a slight time delay after the rotor sweeps pass the stator trailing edge. For the rotor/stator/rotor configuration, with Gap1 = 10 percent chord and Gap2 = 30 percent chord, results showed a 60 percent reduction in the stator force amplitude by clocking the downstream rotor so that the time occurrence of the maximum force due to the upstream vortical disturbance coincides with that of the minimum force due to the downstream potential disturbance. This is the first time, the authors believe, that beneficial use of flow unsteadiness is definitively demonstrated to reduce the blade unsteady loading.

A planar interdigitated ring electrode array via dielectrophoresis for uniform patterning of cells

Uniform patterning of cells is highly desirable for most cellular studies involving cell-cell interactions but is often difficult in an in vitro environment. This paper presents the development of a collagen-coated planar interdigitated ring electrode (PIRE) array utilizing positive dielectrophoresis to pattern cells uniformly. Key features of the PIRE design include: (1) maximizing length along the edges where the localized maximum in the electric field exists; (2) making the inner gap slightly smaller than the outer gap in causing the electric field strength near the center of a PIRE being generally stronger than that near the outer edge of the same PIRE. Results of human hepatocellular carcinoma cells, HepG2, adhered on a 6x6 PIRE array show that cells patterned within minutes with good uniformity (48+/-6 cells per PIRE). Cell viability test revealed healthy patterned cells after 24h that were still confined to the collagen-coated PIREs. Furthermore, quantification of fluorescence intensity of living cells shows an acceptable reproducibility of cell viability among PIREs (mean normalized intensity per PIRE was 1+/-0.138). The results suggest that the PIRE array would benefit applications that desire uniform cellular patterning, and improve both response and reproducibility of cell-based biosensors.

Modeling of cancer metastasis and drug resistance via biomimetic nano-cilia and microfluidics

Three-dimensional (3D) tissue culture platforms that are capable of mimicking in vivo microenvironments to replicate physiological conditions are vital tools in a wide range of cellular and clinical studies. Here, learning from the nature of cilia in lungs - clearing mucus and pathogens from the airway - we develop a 3D culture approach via flexible and kinetic copolymer-based chains (nano-cilia) for diminishing cell-to-substrate adhesion. Multicellular spheroids or colonies were tested for 3-7 days in a microenvironment consisting of generated cells with properties of putative cancer stem cells (CSCs). The dynamic and reversible regulation of epithelial-mesenchymal transition (EMT) was examined in spheroids passaged and cultured in copolymer-coated dishes. The expression of CSC markers, including CD44, CD133, and ABCG2, and hypoxia signature, HIF-1α, was significantly upregulated compared to that without the nano-cilia. In addition, these spheroids exhibited chemotherapeutic resistance in vitro and acquired enhanced metastatic propensity, as verified from microfluidic chemotaxis assay designed to replicate in vivo-like metastasis. The biomimetic nano-cilia approach and microfluidic device may offer new opportunities to establish a rapid and cost-effective platform for the study of anti-cancer therapeutics and CSCs.

A travelling wave dielectrophoretic pump for blood delivery

The travelling wave dielectrophoretic pump studied here is essentially a rectangular straight micro-channel with an electrode array on part of its wall, and operated under an ac voltage with phase shift at neighbouring electrodes. The travelling wave dielectrophoretic force drives the cells, which drag the plasma, and after some sophisticated interaction between conventional dielectrophoresis, travelling wave dielectrophoresis and fluid mechanics, the whole blood is delivered. The pump was fabricated using MEMS techniques and studied in details for different parameters. It is found that the pumping velocity is maximized at an intermediate frequency around 20-30 MHz (varies with phase shift), and at an intermediate channel height at about 40 microm. The quasi-static average cell velocity can reach 15 microm s(-1) for a pump with 1 mm length and 16 electrodes (total array length 465 microm) operated at 5 V and 20 MHz with 90 degrees phase shift.

Patch clamping on plane glass—fabrication of hourglass aperture and high-yield ion channel recording

Planar patch-clamp has revolutionized ion-channel measurement by eliminating laborious manipulation from the traditional micropipette approach and enabling high throughput. However, low yield in gigaseal formation and/or relatively high cost due to microfabricated processes are two main drawbacks. This paper presents patch clamping on glass substrate-an economical solution without sacrificing gigaseal yield rate. Two-stage CO(2) laser drilling methodology was used to generate an hourglass, funnel-like aperture of a specified diameter with smooth and debris-free surfaces on 150 microm borosilicate cover glass. For 1-3 microm apertures as patch-clamp chips, seal resistance was tested on human embryonic kidney, Chinese hamster ovary, and Jurkat T lymphoma cells with a gigaseal success rate of 62.5%, 43.6% and 66.7% respectively. Results also demonstrated both whole-cell and single channel recording on endogenously expressed ion channels to confirm the capability of different patch configurations.

Detection of Circulating Endothelial Cells via a Microfluidic Disk

BACKGROUND Circulating endothelial cells (CECs) in the blood are rare but have been shown to be associated with various diseases. With the ratio of CECs to peripheral blood mononuclear cells (PBMCs) less than 1 part per thousand, their separation from PBMCs and detection are challenging. We present a means of detecting CECs from PBMCs via an economical microfluidic disk with a model cell system [human umbilical vein endothelial cells (HUVECs) in PBMCs], along with demonstration of its efficacy clinically. METHODS To enrich these rare cells, we used immunomagnetic beads and a tailor-made magnet on the disk. CEC-simulating HUVECs, as target cells, were stained with primary anti-CD146-phycoerythrin antibody and bound with secondary antibody on antiphycoerythrin magnetic beads. PBMCs served as nontarget cells and were labeled with anti-CD45-FITC antibody. RESULTS When hundreds of HUVECs were mixed in 10(6) PBMCs, 95% of spiked HUVECs were detected. This yield also held for 60 HUVEC in <10(4) PBMCs. We compared data from flow cytometry with that from the disk: CEC counts in 50 μL blood from patients with systemic lupus erythematosus were 61.1 (21.5), significantly higher (P < 0.01) than those of healthy donors, 31.2 (13.3). CONCLUSIONS The count of CECs is a suitable marker for symptoms of systemic lupus erythematosus. The microfluidic disk system should be a viable platform for detection of CECs.

Direct Characterization of Motion-Dependent Parameters of Sperm in a Microfluidic Device: Proof of Principle

BACKGROUND Semen analysis is essential for evaluating male infertility. Besides sperm concentration, other properties, such as motility and morphology, are critical indicators in assessing sperm quality. Nevertheless, rapid and complete assessment of these measures still presents considerable difficulty and involves a range of complex issues. Here we present a microfluidic device capable of quantifying a range of properties of human sperm via the resistive pulse technique (RPT). METHODS An aperture, designed as a long channel, was used to allow the quantification of various properties as sperm swam through. RESULTS The time trace of the voltage drop across the aperture during sperm passage contained a wealth of information: the sperm volume was presented by the amplitude of the induced pulse, the swim velocity was evaluated via the duration, and the beat frequency was calculated from the voltage undulation superposed on the pulse signal. The RPT measurement of swim velocity and beat frequency showed a correlation with the same observation in a microscope (R(2) = 0.94 and 0.70, respectively). CONCLUSIONS The proposed proof of principle enables substantial quantification of the motion-dependent properties of sperm. Because this approach requires only a current/voltage source and data analysis, it is economically advantageous compared with optical methods for characterizing sperm motion. Furthermore, this approach may be used to characterize sperm morphology.

Hourglass-shaped aperture for cellular electrophysiological study

This work presents a simple process of reflow of melted glass by CO2 laser to form an hourglass-shaped aperture for cellular electrophysiological study. The fabricated aperture proves to be smooth, circular, debris-free, and freshly activated. Two-phase flow simulation illuminates details of the reflow process. The resulting seal resistance quality is well suited to measure ion-channel activities; a seal resistance of 1.9GΩ for PC-12 cell and a typical resistance of 200MΩ for Chinese hamster ovary cells were achieved. This approach can be integrated with microfluidics in a single cell patch or array configuration.

Ion channel electrophysiology via integrated planar patch-clamp chip with on-demand drug exchange

Planar patch clamp has revolutionized characterization of ion channel behavior in drug discovery primarily via advancement in high throughput. Lab use of planar technology, however, addresses different requirements and suffers from inflexibility to enable wide range of interrogation via a single cell. This work presents integration of planar patch clamp with microfluidics, achieving multiple solution exchanges for tailor‐specific measurement and allowing rapid replacement of the cell‐contacting aperture. Studies via endogenously expressed ion channels in HEK 293T cells were commenced to characterize the device. Results reveal the microfluidic concentration generator produces distinct solution/drug combination/concentrations on‐demand. Volume‐regulated chloride channel and voltage‐gated potassium channels in HEK 293T cells immersed in generated solutions under various osmolarities or drug concentrations show unique channel signature under specific condition. Excitation and blockage of ion channels in a single cell was demonstrated via serial solution exchange. Robustness of the reversible bonding and ease of glass substrate replacement were proven via repeated usage of the integrated device. The present approach reveals the capability and flexibility of integrated microfluidic planar patch‐clamp system for ion channel assays. Biotechnol. Bioeng. 2011; 108:1395–1403.

Enumeration and viability of rare cells in a microfluidic disk via positive selection approach

Recent studies have shown that specific rare cells in the blood can serve as an indicator of cancer prognosis, among other purposes. This article demonstrates the concept of separating and detecting rare cells from peripheral blood mononuclear cells via an economical microfluidic disk with a model system. MCF7, labeled with magnetic beads, was used to simulate circulating tumor cells as a target. Jurkat clone E6-1 was used to simulate leukocytes or other cells abundant in human blood. A tailored multistage magnet maximized the magnetic field to ensure optimal trapping efficiency. Results indicate that the yield of detected MCF7 was consistent at approximately 80% when fewer than hundreds of MCF7 cells were mixed in greater than 1 million Jurkat cells. The 80% yield also held for 10 MCF7 in 100 million Jurkat (rarity of 10(7)). Compared with the results from autoMACS, the performance was at least 20% higher and was more independent of the number of Jurkat. The viability of the enriched cells was approximately 90 ± 20%, showing that this method caused little damage to trapped cells. The microfluidic disk should be applicable for separation and detection of various rare cells, such as circulating tumor cells and circulating endothelial cells in human blood.