Polydimethylsiloxane Chip Research Articles

Primary Hepatocytes Cultured on a Fiber-Embedded PDMS Chip to Study Drug Metabolism.

In vitro drug screening using reliable and predictable liver models remains a challenge. The identification of an ideal biological substrate is essential to maintain hepatocyte functions during in vitro culture. Here, we developed a fiber-embedded polydimethylsiloxane (PDMS) chip to culture hepatocytes. Hepatocyte spheroids formed in this device were subjected to different flow rates, of which a flow rate of 50 μL/min provided the optimal microenvironment for spheroid formation, maintained significantly higher rates of albumin and urea synthesis, yielded higher CYP3A1 (cytochrome P450 3A1) and CYP2C11 (cytochrome P450 2C11) enzyme activities for metabolism, and demonstrated higher expression levels of liver-specific genes. In vitro metabolism tests on tolbutamide and testosterone by hepatocytes indicated predicted clearance rates of 1.98 ± 0.43 and 40.80 ± 10.13 mL/min/kg, respectively, which showed a good in vitro–in vivo correspondence. These results indicate that this system provides a strategy for the construction of functional engineered liver tissue that can be used to study drug metabolism.

Direct 3D cell-printing of human skin with functional transwell system

Three-dimensional (3D) cell-printing has been emerging as a promising technology with which to build up human skin models by enabling rapid and versatile design. Despite the technological advances, challenges remain in the development of fully functional models that recapitulate complexities in the native tissue. Moreover, although several approaches have been explored for the development of biomimetic human skin models, the present skin models based on multistep fabrication methods using polydimethylsiloxane chips and commercial transwell inserts could be tackled by leveraging 3D cell-printing technology. In this paper, we present a new 3D cell-printing strategy for engineering a 3D human skin model with a functional transwell system in a single-step process. A hybrid 3D cell-printing system was developed, allowing for the use of extrusion and inkjet modules at the same time. We began by revealing the significance of each module in engineering human skin models; by using the extrusion-dispensing module, we engineered a collagen-based construct with polycaprolactone (PCL) mesh that prevented the contraction of collagen during tissue maturation; the inkjet-based dispensing module was used to uniformly distribute keratinocytes. Taking these features together, we engineered a human skin model with a functional transwell system; the transwell system and fibroblast-populated dermis were consecutively fabricated by using the extrusion modules. Following this process, keratinocytes were uniformly distributed onto the engineered dermis by the inkjet module. Our transwell system indicates a supportive 3D construct composed of PCL, enabling the maturation of a skin model without the aid of commercial transwell inserts. This skin model revealed favorable biological characteristics that included a stabilized fibroblast-stretched dermis and stratified epidermis layers after 14 days. It was also observed that a 50 times reduction in cost was achieved and 10 times less medium was used than in a conventional culture. Collectively, because this single-step process opens up chances for versatile designs, we envision that our cell-printing strategy could provide an attractive platform in engineering various human skin models.

Toward integrated detection and graphene-based removal of contaminants in a lab-on-a-chip platform

A novel, miniaturized microfluidic platform was developed for the simultaneous detection and removal of polybrominated diphenyl ethers (PBDEs). The platform consists of a polydimethylsiloxane (PDMS) microfluidic chip for an immunoreaction step, a PDMS chip with an integrated screen-printed electrode (SPCE) for detection, and a PDMS-reduced graphene oxide (rGO) chip for physical adsorption and subsequent removal of PBDE residues. The detection was based on competitive immunoassay-linked binding between PBDE and PBDE modified with horseradish peroxidase (HRP-PBDE) followed by the monitoring of enzymatic oxidation of o-aminophenol (o-AP) using square wave anodic stripping voltammetry (SW-ASV). PBDE was detected with good sensitivity and a limit of detection similar to that obtained with a commercial colorimetric test (0.018 ppb), but with the advantage of using lower reagent volumes and a reduced analysis time. The use of microfluidic chips also provides improved linearity and a better reproducibility in comparison to those obtained with batch-based measurements using screen-printed electrodes. In order to design a detection system suitable for toxic compounds such as PBDEs, a reduced graphene oxide–PDMS composite was developed and optimized to obtain increased adsorption (based on both the hydrophobicity and π–π stacking between rGO and PBDE molecules) compared to those of non-modified PDMS. To the best of our knowledge, this is the first demonstration of electrochemical detection of flame retardants and a novel application of the rGO-PDMS composite in a biosensing system. This system can be easily applied to detect any analyte using the appropriate immunoassay and it supports operation in complex matrices such as seawater.

Flow lithography in ultraviolet-curable polydimethylsiloxane microfluidic chips.

Flow Lithography (FL) is the technique used for the synthesis of hydrogel microparticles with various complex shapes and distinct chemical compositions by combining microfluidics with photolithography. Although polydimethylsiloxane (PDMS) has been used most widely as almost the sole material for FL, PDMS microfluidic chips have limitations: (1) undesired shrinkage due to the thermal expansion of masters used for replica molding and (2) interfacial delamination between two thermally cured PDMS layers. Here, we propose the utilization of ultraviolet (UV)-curable PDMS (X-34-4184) for FL as an excellent alternative material of the conventional PDMS. Our proposed utilization of the UV-curable PDMS offers three key advantages, observed in our study: (1) UV-curable PDMS exhibited almost the same oxygen permeability as the conventional PDMS. (2) The almost complete absence of shrinkage facilitated the fabrication of more precise reverse duplication of microstructures. (3) UV-cured PDMS microfluidic chips were capable of much stronger interfacial bonding so that the burst pressure increased to ∼0.9 MPa. Owing to these benefits, we demonstrated a substantial improvement of productivity in synthesizing polyethylene glycol diacrylate microparticles via stop flow lithography, by applying a flow time (40 ms) an order of magnitude shorter. Our results suggest that UV-cured PDMS chips can be used as a general platform for various types of flow lithography and also be employed readily in other applications where very precise replication of structures on micro- or sub-micrometer scales and/or strong interfacial bonding are desirable.

Fluorescence radiation characteristics based on evanescent wave pumping in a microfluidic chip

A bare quartz optical fiber is implanted in a microfluidic channel of polydimethylsiloxane (PDMS) substrate. Pumping the microfluid by a continuous wave laser with a wavelength of 532 nm along the fiber axis, the fluorescent spectra from the channel filled with lower refractive index (RI) dye solution are obtained. Due to the fact that the evanescent field of the pump beam is homogeneous around fiber, the fluorescent emission from the rim of fiber is uniform. It is found experimentally that the fluorescent emission intensity decreases with the axial distance of fiber, and the intensity is very sensitive to the RI of the dye solution and the dye concentration. For the dye solution with a large RI, the emitted fluorescent intensity attenuates along the fiber axis more obviously than that of the dye solution with a small RI. For the high dye concentration solution, the emitted fluorescent intensity attenuates along the fiber axis also more significantly than that of the low dye concentration solution. Therefore, it is possible to obtain a uniform fluorescence radiation along the fiber axis by selecting a suitably smaller RI and a lower dye concentration solution. The observed experimental phenomena are well explained based on the mechanism of evanescent wave pumping fluorescent radiation. Based on the features of fluorescent emission in the microfluidic chip, a PDMS chip with three micro-channels is designed and fabricated. After injecting ethanol solutions of rhodamine 640, rhodamine B and rhodamine 6 G separately into the three channels and pumpingthese solutions by evanescent wave along the optical fiber axis, three fluorescence emissions with different wavelength ranges are successfully observed in a single PDMS chip.

A Vascular Permeability Assay Using an In Vitro Human Microvessel Model Mimicking the Inflammatory Condition.

The vascular barrier is an important function of the endothelium and its dysfunction is involved in several diseases. The barrier function of the endothelial cell monolayer is governed by cell-cell, cell-extracellular matrix (cell-ECM) contacts, and inflammatory factors such as thrombin, histamine or vascular endothelial growth factor. Several in vivo and in vitro assays that measure the vascular permeability induced by these factors have been developed. However, they suffer limitations such as being challenging for assessing details of biological processes at a cellular level or lacking the architecture of a vessel, that raise the need for new methods. In vitro 3D model-based assays have thus been developed but assays for investigating compounds that protects the barrier function are lacking. Here we describe the development of an in vitro three-dimensional (3D) vascular endothelium model in which we can manipulate the endothelial barrier function and permeability to molecules, which have a molecular weight similar to human serum albumin, allowing to assess the protective effect of compounds. A microvessel was prepared by culturing human umbilical vein endothelial cells (HUVECs) within a collagen gel on a polydimethylsiloxane (PDMS) chip. Using fluorescein isothiocyanate (FITC)-conjugated dextran (70 kDa, FITC-dextran) and confocal fluorescence microscopy, we showed that the microvessel presented an effective barrier function. We were then able to induce the loss of this barrier function by treatment with the inflammatory factor thrombin. The loss of barrier function was quantified by the extravasation of FITC-dextran into collagen matrix. Furthermore, we were able to analyze the protective effect on the endothelial barrier function of the cyclic adenosine monophosphate (cAMP) analog, 8-pCPT-2'-O-Me-cAMP (also called 007). In an attempt to understand the effects of thrombin and 007 in our model, we analyzed the adherens junctions and cytoskeleton through immunostaining of the vascular endothelial cadherin and actin, respectively. Our assay method could be used to screen for compounds modulating the barrier function of endothelial cells, as well as investigating mechanistic aspects of barrier dysfunction.

Separation of superparamagnetic particles through ratcheted Brownian motion and periodically switching magnetic fields.

Brownian ratchet based particle separation systems for application in lab on chip devices have drawn interest and are subject to ongoing theoretical and experimental investigations. We demonstrate a compact microfluidic particle separation chip, which implements an extended on-off Brownian ratchet scheme that actively separates and sorts particles using periodically switching magnetic fields, asymmetric sawtooth channel sidewalls, and Brownian motion. The microfluidic chip was made with Polydimethylsiloxane (PDMS) soft lithography of SU-8 molds, which in turn was fabricated using Proton Beam Writing. After bonding of the PDMS chip to a glass substrate through surface activation by oxygen plasma treatment, embedded electromagnets were cofabricated by the injection of InSn metal into electrode channels. This fabrication process enables rapid production of high resolution and high aspect ratio features, which results in parallel electrodes accurately aligned with respect to the separation channel. The PDMS devices were tested with mixtures of 1.51 μm, 2.47 μm, and 2.60 μm superparamagnetic particles suspended in water. Experimental results show that the current device design has potential for separating particles with a size difference around 130 nm. Based on the promising results, we will be working towards extending this design for the separation of cells or biomolecules.

Using injection molding and reversible bonding for easy fabrication of magnetic cell trapping and sorting devices

Magnetism and microfluidics are two key elements for the development of inexpensive and reliable tools dedicated to high-throughput biological analysis and providing a large panel of applications in domains ranging from fundamental biology to medical diagnostics. In this work, we introduce a simple protocol, relying on injection molding and reversible bonding for fabrication of magnetic cell trapping and sorting devices using only standard soft-lithography equipment. Magnetic strips or grids made of Polydimethylsiloxane (PDMS) doped with hard (NdFeB) or soft (carbonyl iron) magnetic powders were integrated at the bottom of whole PDMS chips. Preliminary results show the effective deviation/trapping of magnetic beads or magnetically-labeled bacteria as the sample flows through the microchannel, proving the potential of this rapid prototyping approach for easy fabrication of magnetic cell sorters.

Practical fabrication of microfluidic platforms for live-cell microscopy.

We describe a simple fabrication technique - targeted towards non-specialists - that allows for the production of leak-proof polydimethylsiloxane (PDMS) microfluidic devices that are compatible with live-cell microscopy. Thin PDMS base membranes were spin-coated onto a glass-bottom cell culture dish and then partially cured via microwave irradiation. PDMS chips were generated using a replica molding technique, and then sealed to the PDMS base membrane by microwave irradiation. Once a mold was generated, devices could be rapidly fabricated within hours. Fibronectin pre-treatment of the PDMS improved cell attachment. Coupling the device to programmable pumps allowed application of precise fluid flow rates through the channels. The transparency and minimal thickness of the device enabled compatibility with inverted light microscopy techniques (e.g. phase-contrast, fluorescence imaging, etc.). The key benefits of this technique are the use of standard laboratory equipment during fabrication and ease of implementation, helping to extend applications in live-cell microfluidics for scientists outside the engineering and core microdevice communities.

Self-regenerating and hybrid irreversible/reversible PDMS microfluidic devices.

This paper outlines a straightforward, fast, and low-cost method to fabricate polydimethylsiloxane (PDMS) chips. Termed sandwich bonding (SWB), this method requires only a laboratory oven. Initially, SWB relies on the reversible bonding of a coverslip over PDMS channels. The coverslip is smaller than the substrate, leaving a border around the substrate exposed. Subsequently, a liquid composed of PDMS monomers and a curing agent is poured onto the structure. Finally, the cover is cured. We focused on PDMS/glass chips because of their key advantages in microfluidics. Despite its simplicity, this method created high-performance microfluidic channels. Such structures featured self-regeneration after leakages and hybrid irreversible/reversible behavior. The reversible nature was achieved by removing the cover of PDMS with acetone. Thus, the PDMS substrate and glass coverslip could be detached for reuse. These abilities are essential in the stages of research and development. Additionally, SWB avoids the use of surface oxidation, half-cured PDMS as an adhesive, and surface chemical modification. As a consequence, SWB allows surface modifications before the bonding, a long time for alignment, the enclosure of sub-micron channels, and the prototyping of hybrid devices. Here, the technique was successfully applied to bond PDMS to Au and Al.

Semi-contact-writing of polymer molds for prototyping PDMS chips with low surface roughness, sharp edges and locally varying channel heights

Microfluidic systems fabricated in polydimethylsiloxane (PDMS) enable a broad variety of applications and are widespread in the field of Lab-on-a-Chip. Here we demonstrate semi-contact-writing, a novel method for fabrication of polymer based molds for casting microfluidic PDMS chips in a highly flexible, time and cost-efficient manner. The method is related to direct-writing of an aqueous polymer solution on a planar glass substrate and substitutes conventional, time- and cost-consuming UV-lithography. This technique facilitates on-demand prototyping in a low-cost manner and is therefore ideally suited for rapid chip layout iterations. No cleanroom facilities and less expertise are required. Fabrication time from scratch to ready-to-use PDMS-chip is less than 5 h. This polymer writing method enables structure widths down to 140 μm and controllable structure heights ranging from 5.5 μm for writing single layers up to 98 μm by stacking. As a unique property, freely selectable height variations across a substrate can be achieved by application of local stacking. Furthermore, the molds exhibit low surface roughness (Ra = 24 nm, RRMS = 28 nm) and high fidelity edge sharpness. We validated the method by fabrication of molds to cast PDMS chips for droplet based flow-through PCR with single-cell sensitivity.

Autonomous Chemical Oscillator Circuit Based on Bidirectional Chemical‐Microfluidic Coupling

Autonomous Chemical Oscillator Circuit Based on Bidirectional Chemical‐Microfluidic Coupling

An integrated acoustic and dielectrophoretic particle manipulation in a microfluidic device for particle wash and separation fabricated by mechanical machining

In this study, acoustophoresis and dielectrophoresis are utilized in an integrated manner to combine the two different operations on a single polydimethylsiloxane (PDMS) chip in sequential manner, namely, particle wash (buffer exchange) and particle separation. In the washing step, particles are washed with buffer solution with low conductivity for dielectrophoretic based separation to avoid the adverse effects of Joule heating. Acoustic waves generated by piezoelectric material are utilized for washing, which creates standing waves along the whole width of the channel. Coupled electro-mechanical acoustic 3D multi-physics analysis showed that the position and orientation of the piezoelectric actuators are critical for successful operation. A unique mold is designed for the precise alignment of the piezoelectric materials and 3D side-wall electrodes for a highly reproducible fabrication. To achieve the throughput matching of acoustophoresis and dielectrophoresis in the integration, 3D side-wall electrodes are used. The integrated device is fabricated by PDMS molding. The mold of the integrated device is fabricated using high-precision mechanical machining. With a unique mold design, the placements of the two piezoelectric materials and the 3D sidewall electrodes are accomplished during the molding process. It is shown that the proposed device can handle the wash and dielectrophoretic separation successfully.

Highly Specific Electrochemiluminescence Detection of Cancer Cells with a Closed Bipolar Electrode

AbstractAn advanced biosensor based on a closed bipolar electrode (BPE)–electrochemiluminescence (ECL) strategy is proposed for cancer cell adhesion, specific recognition, and detection with dual biomarkers. The system consists of a two‐channel polydimethylsiloxane (PDMS) chip (sensing channel and reporting channel) connected through a U‐shaped indium tin oxide BPE at a glass surface. A sandwich‐type cancer cell detection model is developed at the cathode of the BPE with two recognition molecules, folic acid (FA) and an aptamer. On the basis of the high affinity between FA and the folate receptor that is overexpressed at a cancer cell membrane as well as the excellent conductivity of graphene–Au conjugates, this strategy could achieve the sensitive detection of HL‐60 cells, with a limit as low as 18 cells in 30 μL of cell suspension. Control experiments confirm that the sandwich assay specifically captures cancer cells and recognizes target cells.

A Microflow Cytometer Based on a Disposable Microfluidic Chip With Side Scatter and Fluorescence Detection Capability.

In this paper, we present a microflow cytometer based on a polydimethylsiloxane (PDMS) microfluidic chip. The PDMS chip has a smooth sidewall because of the proposed double molding process. Thus, the side scatter (SSC) detection can be used for the cytometer. The coincidence error using the sheathless focus mechanism is less than 0.069%. Using Flow-Check alignment beads for the counting experiment, the coefficient of variation (CV) of the SSC and the fluorescein isothiocyanate (FITC) channel can reach 8.37% and 2.46%, respectively. The gating of the lymphocytes according to CD45/SSC was also demonstrated successfully. It proves that the chip is a promising solution for point of care (POC) microflow cytometers.

A microfluidic droplet digital PCR for simultaneous detection of pathogenic Escherichia coli O157 and Listeria monocytogenes.

Sensitive and rapid identification of pathogenic bacterial is extremely important due to the serious threat of pathogens to human health. In this study, we demonstrate the simultaneous and sensitive detection of pathogenic Escherichia coli O157 and Listeria monocytogenes using a novel duplex droplet digital PCR (ddPCR) platform. The ddPCR platform, which uses a mineral oil-saturated polydimethylsiloxane (OSP) chip to overcome the problem of droplet evaporation, integrates the functions of droplet generation, on-chip amplification and end-point fluorescence readout. Simultaneous detection of two kinds of bacterial is achieved by the design of differentially labeled TaqMan-MGB fluorescent probes. Compared with a quantitative real-time PCR approach, the OSP chip-based duplex ddPCR platform exhibits high sensitivity, which is at the level of single molecule resolution without significant cross-assay interference. Moreover, the applicability of the proposed method is also evaluated in artificially contaminated drinking water sample, which displays a low detection limit down to 10 CFU/mL for both pathogenic bacterial within 2 h.

Easy-to-realise Polyvinylsiloxane Microfluidic Connectors for PDMS Chips

The connection of tubes to a polydimethylsiloxane (PDMS) device can usually not be realised easily. Tube connectors embedded in the PDMS structure have a large foot print. Thick layers of PDMS in which the tubes or needles can hold hinders the visualisation of sample within the device. In this contribution we present microfluidic connectors for PDMS chips realized by easy-to-use elastomeric impression material polyvinylsiloxane (PVS). PVS cylinders were added to a pre-cured PDMS chip and ready to use within five minutes without the need of exposure to high temperatures or the use of additional glues. The robustness of the PVS connectors has been demonstrated through pull-tests and liquid pressure tests.

Piezoelectric layer embedded-microdiaphragm sensors for the determination of blood viscosity and density

We introduce a lead zirconate titanate [PZT; Pb(Zr0.52Ti0.48)O3] microdiaphragm resonating sensor packaged in a polydimethylsiloxane chip. The proposed sensor can measure the density and viscosity of a liquid that is within the density and viscosity regime of blood (1.060 × 103 kg/m3, 3–4 cP). To verify the basic characteristics of the sensor, viscous solutions were prepared from glycerol and deionized water with a density in the range from 0.998 to 1.263 × 103 kg/m3 and a viscosity in the range from 1 to 1414 cP. We measured the frequency responses of the sensor before and after injecting the viscosity- and density-controlled liquid under the bottom of the microdiaphragm. The resonant frequencies in the (1,1) and (2,2) modes decreased linearly as a function of the liquid density in the range from 0.998 to 1.146 × 103 kg/m3 with a sensitivity of 28.03 Hz/kg·m−3 and 81.85 Hz/kg·m−3, respectively. The full width at half maximum had a logarithmic relationship with the liquid viscosity in the viscosity range from 1 to 8.4 cP. The quality factor (Q-factor) for the 50% glycerol/water mixture was determined to be greater than 20 for both the (1,1) and the (2,2) modes, indicating that the microdiaphragm resonating sensor is suitable for measuring the density and viscosity of a liquid within a density range from 0.998 to 1.1466 × 103 kg/m3 and a viscosity range from 1 to 8.4 cP. These density and viscosity ranges span the regime of possible changes of blood characteristics. The microdiaphragm resonating sensors were also tested with a real human serum to verify that the sensor is suitable for measuring the viscosity and density of blood. Therefore, the PZT microdiaphragm resonating sensor could be utilized for early diagnosis of diseases associated with changes in the physical properties of blood.

OncomiR detection in circulating body fluids: a PDMS microdevice perspective.

There is an increasing interest in circulating microRNAs (miRNAs) as potential minimally invasive diagnostic biomarkers in oncology. Considerable efforts are being made in the development of lab-on-a-chip devices for biomedical applications to purify and detect miRNAs from biological fluids. Here, we report the development of an innovative polydimethylsiloxane (PDMS)-based parallel device whose internal surface can opportunely be functionalized with positively charged 3-aminopropyltriethoxysilane (APTES) alone or mixed with two different neutral poly(ethylene glycol) silanes (PEG-s). The differently functionalized internal surfaces of the PDMS chip were characterized with s-SDTB (sulfosuccinimidyl-4-o-(4,4-dimethoxytrityl) butyrate) and the portion of the surface able to adsorb a synthetic fluorescently labeled miRNA was determined. Interestingly, the adsorbed miRNA (both synthetic and cell supernatant-derived) was found mainly on the bottom surface of the chip and could be reverse transcribed into cDNA directly on the same PDMS chip used for its purification, saving hours with respect to the use of standard purification kits. We identified 0.1% APTES/0.9% PEG-silane as the most efficient PDMS functionalization to capture both synthetic and extracellular miRNA. Moreover, the amount of captured miRNA was increased by treating the cell supernatant with a commercially available lysis buffer for RNA extraction. We assessed that the available miRNA binding sites on the functionalized surface were efficiently saturated with only one incubation, shortening the time and greatly simplifying the protocol for miRNA purification from biological samples. Finally, the extracellular miRNA purification efficiency of the PDMS functionalized multichip determined via real-time quantitative polymerase chain reaction (RT-qPCR) was confirmed by droplet digital PCR (ddPCR) quantification. This work shows an innovative, rapid and easy to use microdevice for the purification and reverse transcription of circulating miRNAs, approaching the realization of diagnostic and prognostic oncomiR-based assays.

Inner microvascular function assessment through an opto-mechanical polydimethylsiloxane chip

Inner microvascular function assessment through an opto-mechanical polydimethylsiloxane chip