Parallel separation of multiple samples with negative pressure sample injection on a 3‐D microfluidic array chip

A digital quantification method for the detection of biomarkers on a microfluidic array chip

In this paper, we present an integrated and automated prototype system which has been developed for real-time polymerase chain reaction (PCR) analysis based on microfluidic PCR array chips. The system integrates the PCR thermal cycling and optical detection capabilities to enable real-time fluorescence imaging and image processing for data analysis. The main advantage of the system is that it provides a solution that can rapidly perform and evaluate PCR experiment simultaneously on microfluidic PCR array chips. The system has demonstrated fast and efficient on-chip real-time PCR analysis using human genomic DNA samples. The implementation of the system integration is a multi-thread Windows software with component structure which is written in Visual C++.

A novel open-access microfluidic patch-clamp array chip with lateral cell trapping sites raised above the bottom plane of the chip was developed by combining both a microscale soft-lithography and a macroscale polymer fabrication method. This paper demonstrates the capability of using such an open-access fluidic system for patch-clamp measurements. The surface of the open-access patch-clamp sites prepared by the macroscale hole patterning method of soft-state elastic polydimethylsiloxane (PDMS) is examined; the seal resistances are characterized and correlated with the aperture dimensions. Whole cell patch-clamp measurements are carried out with CHO cells expressing Kv2.1 ion channels. Kv2.1 ion channel blocker (TEA) dosage response is characterized and the binding activity is examined. The results demonstrate that the system is capable of performing whole cell measurements and drug profiling in a more efficient manner than the traditional patch-clamp set-up.

We assessed blood interaction with different micrometer-scale topographies under flow conditions using a micro-fluidic array system. The channels of the micro-fluidic array chip were coated with azobenzene polymer films, which were then topographically structured using a one-step non-contact optical technique. A set of surfaces with different topographies was produced varying laser irradiation duration. These surfaces were then exposed to blood flow. The blood flow rate was measured with a micro-channel array flow analyzer. The measured blood flow rates decreased with time for all the samples, indicating formation of platelet clots which obstruct the channels during flow. This effect appeared enhanced on polymer surfaces having a sinusoidal profile with 200-nm-high ridges and 1.2-μm-grating spacing. The morphology of platelets that adhered on the polymer films was studied by scanning electron microscopy. Platelets adhered on azobenzene surfaces with flat topographies, typically exhibiting filopodia. Platelets adhered on optically structured surfaces also exhibited lamellipodia and appeared flattened on surfaces with the highest ridges. We conclude that surface topography influences blood behaviour on azobenzene polymer films.

In this work, an integrated biosensor consisting of spiral microfluidic array and microwave sensors is proposed for simultaneous separation and detection of cells. The biosensor integrated by plasma processing technology is fabricated by soft lithography and glass‐based IC process, which has the advantages of simple preparation, low cost, and reliable structure. In the field of clinical medicine, Escherichia coli ( E. coli ) is the causative agent of urinary tract infections, which leads to an increase in the number of white blood cells (WBCs) present in the urine. Different concentrations of E. coli and WBCs mixed solution are configured to perform biological cell experiments and the capability of the biosensor in separating and detecting WBCs is verified. Interdigital capacitors (IDCs) and split ring resonators (SRRs) are employed to detect the WBCs obtained by microfluidic array separation. The microfluidic array exhibits a WBC collection rate of 92.7%. The capacitance of the IDC and the resonant amplitude of the SRR exhibit a decrease of 51.36 pF and 0.34 dB, which demonstrates a satisfactory linearity of 0.96 and 0.98, respectively. Consequently, the integrated biosensor used to simultaneously separate and detect WBCs has the potential for the early diagnosis of urinary tract infections in clinical medicine.

<p indent="0mm">Microfluidic chips have attracted increasing research attention in biology, chemistry, and medicine; in particular, microfluidic pipes containing metal electrodes have broad prospects in capillary electrophoresis, electrochemical microdetection, biomedical engineering, and flexible electronics. In this study, a simple on-demand method for fabricating arrayed flexible microfluidic pipes with embedded metal electrodes was proposed. The method is based on the direct-writing technology of electrohydrodynamic jet printing (E-jet), which is combined with the conventional mold turning and wet etching processes to obtain flexible microfluidic pipe arrays with embedded metal electrodes. First, a linear convex microstructure with a smooth surface (line width of 10–100 μm and aspect ratio &gt;1:2) was prepared by stacking direct-writing polyvinyl alcohol fibers using the E-jet printing system with a linear rotation-receiving substrate to serve as a template. Using the template, the arrayed microfluidic channels were obtained. Second, a flexible metal patterned conductive electrode (line width &lt;5 μm) was printed on the flexible microfluidic pipe array substrate using the E-jet printing system with a linear translation-receiving substrate and the wet etching process, with a direct-writing photoresist as the protective layer. Finally, the microfluidic pipes were electrically tested with different salt solution concentrations to verify the conductivity of the pipes and the metal electrode. The results indicate that the integrated manufacturing process based on the E-jet printing could be used to simply and efficiently fabricate flexible microfluidic pipe arrays embedded with gold electrodes on-demand and at a low cost. These arrays can be broadly applied in biomedical engineering and flexible electronics.

A novel sample injection method for chip CE was presented. This injection method uses hydrostatic pressure, generated by emptying the sample waste reservoir, for sample loading and electrokinetic force for dispensing. The injection was performed on a double-cross microchip. One cross, created by the sample and separation channels, is used for formation of a sample plug. Another cross, formed by the sample and controlling channels, is used for plug control. By varying the electric field in the controlling channel, the sample plug volume can be linearly adjusted. Hydrostatic pressure takes advantage of its ease of generation on a microfluidic chip, without any electrode or external pressure pump, thus allowing a sample injection with a minimum number of electrodes. The potential of this injection method was demonstrated by a four-separation-channel chip CE system. In this system, parallel sample separation can be achieved with only two electrodes, which is otherwise impossible with conventional injection methods. Hydrostatic pressure maintains the sample composition during the sample loading, allowing the injection to be free of injection bias.

This paper presents an experimental investigation on the use of high-resolution injection techniques to deliver sample plugs within electrophoresis microchips. Two novel injection microfluidic chips are proposed, which employ conventional cross-shaped and U-shaped injection system combined with an expander to deliver high-quality sample plugs for detection in separation channel. The valving characteristics on microfluidic devices are controlled through appropriate manipulations of the electric potential strengths during the sample injection and separation steps. These novel injection techniques developed in this study has an exciting potential for use in high-quality, high-throughput chemical analysis applications and in many other applications throughout the field of micro-total-analysis systems.

Analysis of oral fluid (e.g., saliva) has the potential to aid diagnosis of both oral and systemic diseases. Oral fluids offer advantages over other bodily fluids, including quick, non-invasive, and inexpensive collection in both clinical and non-clinical settings. Nevertheless, current assay techniques using saliva are time-consuming, require large sample volumes, and are not amenable to automation or portability. We report on a robust microfluidic platform that performs rapid immunoassay-based analysis (<3 minutes) of saliva with low sample volume requirements (10 /spl mu/L) and appreciable sensitivity (10/sup -12/ M) with a high-degree of autonomy. Immunoassays for the detection of the systemic disease biomarker, C-reactive protein, and the cytokines interleukin-6 and tumor necrosis factor-/spl alpha/ were conducted in spiked human saliva using a microfluidic chip. The immunoassay developed is based on binding of fluorescently-labeled reporter antibodies to analytes followed by electrophoretic separation and quantitation of bound and unbound reporter. The separation selectivity was achieved using native polyacrylamide gel electrophoresis in a microchip containing in situ photolithographically patterned cross-linked polyacrylamide. Native electrophoresis conditions maintained the integrity of fragile immune complexes and provided rapid separations of complex samples. On-going work focuses on integration of the on-chip immunoassay with miniaturized hardware towards developing a portable prototype.

The fabrication and performance of a microfluidic device with integrated liquid-core optical waveguides for laser induced fluorescence DNA fragment analysis is presented. The device was fabricated through poly(dimethylsiloxane) (PDMS) soft lithography and waveguides are formed in dedicated channels through the addition of a liquid PDMS pre-polymer of higher refractive index. Once a master has been fabricated, microfluidic chips can be produced in less than 3 h without the requirement for a cleanroom, yet this method provides an optical system that has higher performance than a conventional confocal optical assembly. Optical coupling was achieved through the insertion of optical fibers into fiber-to-waveguide couplers at the edge of the chip and the liquid-fiber interface results in low reflection and scattering losses. Waveguide propagation losses are measured to be 1.8 dB cm(-1) (532 nm) and 1.0 dB cm(-1) (633 nm). The chip displays an average total coupling loss of 7.6 dB due to losses at the optical fiber interfaces. In the electrophoretic separation and detection of a BK virus PCR product, the waveguide system achieves an average signal-to-noise ratio of 570 +/- 30 whereas a commercial confocal benchtop electrophoresis system achieves an average SNR of 330 +/- 30. To our knowledge, this is the first time that a waveguide-based system has been demonstrated to have a SNR comparable to a commercially available confocal-based system for microchip capillary electrophoresis.

Phase modulators in surface plasmon resonance phase-differential imaging (SPR-PI) sensing systems reported so far are sensitive to temperature fluctuations or mechanical vibrations and thus their applications are limited. In this paper, we propose a novel prism phase modulator (PPM) to replace a traditional modulator. The PPM consists of a parallel prism, a rotation stage and a mirror. The PPM shows great stability in our experiment, and helps achieve high detection sensitivity in our SPR-PI system. Moreover, the cost of our PPM is much lower than that of a traditional modulator and is thus suitable for commercialization. A polydimethylsiloxane (PDMS) microfluidic chip is fabricated to control the flow velocity and realize parallel detection in our experiment. Measured result of glycerine solution shows that the resolution of our SPR biosensor array is about 9.11×10-7 refractive index unit (RIU). Real time monitoring of interaction between bovine IgG and anti-bovine IgG is also realized. The proposed PPM-based microfluidic SPR-PI biosensor array is promising for future practical applications.

This chapter introduces the layout design tools for microfluidic devices and presents a new script-based method to generate complex layouts of microfluidic devices using Nanolithography Toolbox (NT) software. Compared with other computer-aided design or integrated circuits-based software, NT software is based on an extensive set of parameterized blocks and features, formulated into a simple text file, which is converted into a GDSII format to fabricate lithography masks. The utilization of NT software for structures such as capillary electrophoresis chips, microfluidic emulsion generator array chips using centrifugal forces, chambers, splitters and mixers, etc., is demonstrated. This NT software is a powerful yet easy-to-use tool to generate the layout of microfluidic devices.

Electroporation is pivotal in bioelectrochemistry for cellular manipulation, with prominent applications in drug delivery and cell membrane studies. A comprehensive understanding of pore generation requires an in-depth analysis of the critical pore size and the corresponding energy barrier at the onset of cell rupture. However, many studies have been limited to basic models such as artificial membranes or theoretical simulations. Challenging this paradigm, our study pioneers using a microfluidic electroporation chip array. This tool subjects live breast cancer cell species to a diverse spectrum of alternating current electric field conditions, driving electroporation-induced cell rupture. We conclusively determined the rupture voltages across varying applied voltage loading rates, enabling an unprecedented characterization of electric cell rupture dynamics encompassing critical pore radius and energy barrier. Further bolstering our investigation, we probed cells subjected to cholesterol depletion via methyl-β-cyclodextrin and revealed a strong correlation with electroporation. This work not only elucidates the dynamics of electric rupture in live cell membranes but also sets a robust foundation for future explorations into the mechanisms and energetics of live cell electroporation.

Recently there has been an increased focus on the use of microfluidics for the synthesis of different products. One of the products proposed for synthesis is quantum dots. Microfluidics often uses Polydimethylsiloxane for structure in microfluidic chips, but quantum dots use octadecene in several synthesis steps. The purpose of this work was to create a lab setup capable of anodically bonding 4” diameter wafers, and to characterize the bond formed using the Plaza test chip so that microfluidic devices using glass and silicon as substrates could be created. Two stainless steel electrodes placed on top of a hot plate were attached to a high power voltage supply to perform anodic bonding. A Plaza test mask was created and used to pattern P type silicon wafers. The channels etched were between 300 and 500nm deep and ranged between 1000µm and 50µm. These wafers were then anodically bonded to Corning 7740 glass wafers. Bonding stopped once the entire surface of the wafer was bonded, determined by visual inspection. All bonds were formed at 400°C and the bond strength and toughness between wafers bonded at 400V and 700V was compared. A beam model was used to predict the interfacial fracture toughness, and the stress at the bond was calculated with a parallel spring model. By measuring the crack length of the test structures under a light microscope the load conditions of the beam could be found. It was concluded that the electrostatic forces between the wafers give the best indication of what the bond quality will be. This was seen by the large difference in crack length between samples that were bonded using a thick glass wafer (1 mm) and a thin glass wafer (500µm). The observed crack lengths for the thick glass wafers were between 40 and 60µm. Thin glass wafers had a crack length between 20 and 40µm. The fracture toughness was calculated using the beam model approximation. Fracture toughness of the thin glass wafers was 7MPa m1/2, and of the thick glass wafers was 30 MPa m1/2. The fracture toughness of the thick glass wafers agreed with results found through the use of the double cantilever beam samples in literature. The maximum observed interfacial stress was 70 MPa. Finally, to measure the change in the size of the sodium depletion zone formed during bonding, samples were placed under a scanning electron microscope (SEM). Depletion zones were found to be between

A numerical study is presented for the electroosmotic flow (EOF) at the cross region in microfluidic chips. The distributions of the electric potential due to the electric double layer (EDL) and the external electric field are discussed and the calculation of the latter can give rough speculations on the flow tendencies in the channels during various operation modes. Simplification of the two-dimensional Navier-Stokes (N-S) equations is obtained by focusing on the solution of interior flows, and the numerical calculation results show good agreement with the experimental images. The sample leakage to the separation channel during the "float" sampling proved to be caused not only by the sample diffusion, but also by the weak extension of the sampling electric field. It is also verified that with suitable voltage configuration, the "pinch" sampling mode is better than the "float" mode in sample plug control.