Hybrid Microfluidic Research Articles

One Cell, One Drop, One Click: Hybrid Microfluidics for Mammalian Single Cell Isolation

Generating a stable knockout cell line is a complex process that can take several months to complete. In this work, a microfluidic method that is capable of isolating single cells in droplets, selecting successful edited clones, and expansion of these isoclones is introduced. Using a hybrid microfluidics method, droplets in channels can be individually addressed using a co-planar electrode system. In the hybrid microfluidics device, it is shown that single cells can be trapped and subsequently encapsulate them on demand into pL-sized droplets. Furthermore, droplets containing single cells are either released, kept in the traps, or merged with other droplets by the application of an electric potential to the electrodes that is actuated through an in-house user interface. This high precision control is used to successfully sort and recover single isoclones to establish monoclonal cell lines, which is demonstrated with a heterozygous NCI-H1299 lung squamous cell population resulting from loss-of-function eGFP and RAF1 gene knockout transfections.

Hybrid microfluidics platform for studying electrotaxis of glioblastoma

Equalizing hydrostatic pressures across a microchannel provides a high-throughput approach for studying the deadly disease and highlights the importance of voltage-gated ion channels.

Electrochemical Evaluation of Rhus Vernicifera Biocathodes for Oxygen Reduction Reaction in Neutral Media with Application in Air-Breathing Hybrid Microfluidic Fuel Cells

In this work, a Rhus vernicifera laccase electrodes (RvL-Vulcan/Toray and RvL-ABTS-Vulcan/Toray) were prepared by adsorption method and evaluated toward oxygen reduction reaction by cyclic voltammetry. The characterization was complemented by FTIR with ATR, these results indicates the presence of bands corresponding to the presence of enzymatic material for both electrodes and ABTS in the RvL-ABTS-Vulcan/Toray. The biocathodes were evaluated in an air-breathing microfluidic fuel cell reported previously by our group. For these experiments, a hybrid air-breathing microfluidic fuel cell configuration was constructed by an Au-Vulcan/Toray anode and 5 mM Glucose/0.3 M KOH as anolyte and the RvL-Vulcan/Toray and RvL-ABTS-Vulcan/Toray as biocathodes and PBS 7 as catholyte solution. The results showed that the RvL-ABTS-Vulcan/Toray 16 mWcm-2 performance was better than RvL 3.4 mWcm-2. This result it is attributed to the presence of ABTS in the material inmobilized.

Portable and automated fluorescence microarray biosensing platform for on-site parallel detection and early-warning of multiple pollutants

The portable and automated fluorescence microarray biosensing platform (FMB) that employed a compact hybrid optical structure, microfluidics, and microarray biosensors was constructed for on-site parallel detection of multiple analytes. In the FMB, a hybrid optical structure that composed of a 1 × 4 single mode fiber optic coupler, four fiber optic switches, a single-multi mode fiber optic bundle coupler was at the first time developed for the transmission of the excitation light and the collection and transmission of multi-channel fluorescence signals. Through the control of fiber optic switches, the parallel fluorescence assay of four channels could be achieved using only one excitation light and one photodiode detector on the basis of the time-resolved effect. This optical design not only greatly increased the efficiency of light transmission and fluorescence collection and detection sensitivity of the FMB, but also allows the miniaturization and portability of the whole system because of few optical separation elements used and no requirement of rigorous optical alignment. Taking Microcystin-LR (MC-LR), 2,4-D, atrazine (ATZ), and bisphenol A (BPA) for example, the application potential of the FMB to rapidly and parallelly detect four typical pollutants in real water with high sensitivity and specificity was demonstrated. The limits of detection of MC-LR, 2,4-D, ATZ, and BPA were 0.04 μg/L, 0.09 μg/L, 0.02 μg/L, and 0.03 μg/L, respectively. The FMB could also achieve early-warning of pollutants thanks to its ability of rapidity, high-frequency, and multiple-analyte detection. The FMB has significant implications as a multiplexable, portable, rapid, and quantitative detection platform for pollution accidents and water quality management to satisfy the increasing demands of alerting and protecting civilians.

Commentary on Some Recent Theses Relevant to Combating Aging: December 2019.

Theses reviewed in this issue include "Atrophied Thymus, a Tumor Reservoir for Harboring Melanoma Cells," "Evolutionary Adaptations in Developmental Signaling Pathways Underlie Regenerative Scar-Free Wound Repair in African Spiny Mouse (Genus Acomys)," "Integrated Immunoassays on Paper/Polymer Hybrid Microfluidic Devices for Low-Cost Detection of Disease Biomarkers," "RNA Regulation in the Nervous System: CircRNA Expression Changes During Aging, and Function of the Calm1 Extended 3' UTR Isoform," "The Role of Amylin in Alzheimer's Disease," and "Therapeutic Monoclonal Antibodies to Detect and Halt ATTR Cardiac Amyloidosis and Neuropathy."

Rapid and Highly Controlled Generation of Monodisperse Multiple Emulsions via a One-Step Hybrid Microfluidic Device

Multiple Emulsions (MEs) contain a drop laden with many micro-droplets. A single-step microfluidic-based synthesis process of MEs is presented to provide a rapid and controlled generation of monodisperse MEs. The design relies on the interaction of three immiscible fluids with each other in subsequent droplet formation steps to generate monodisperse ME constructs. The design is within a microchannel consists of two compartments of cross-junction and T-junction. The high shear stress at the cross-junction creates a stagnation point that splits the first immiscible phase to four jet streams each of which are sprayed to micrometer droplets surrounded by the second phase. The resulted structure is then supported by the third phase at the T-junction to generate and transport MEs. The ME formation within microfluidics is numerically simulated and the effects of several key parameters on properties of MEs are investigated. The dimensionless modeling of ME formation enables to change only one parameter at the time and analyze the sensitivity of the system to each parameter. The results demonstrate the capability of highly controlled and high-throughput MEs formation in a one-step synthesis process. The consecutive MEs are monodisperse in size which open avenues for the generation of controlled MEs for different applications.

Synthesis of a Cyberphysical Hybrid Microfluidic Platform for Single-Cell Analysis

Single-cell genomics is used to advance our understanding of diseases, such as cancer. Microfluidic solutions have recently been developed to classify cell types or perform single-cell biochemical analysis on preisolated types of cells. However, new techniques are needed to efficiently classify cells and conduct biochemical experiments on multiple cell types concurrently. Nondeterministic cell-type identification, system integration, and design automation are major challenges in this context. To overcome these challenges, we present a hybrid microfluidic platform that enables complete single-cell analysis on a heterogeneous pool of cells. We combine this architecture with an associated design-automation and optimization framework, referred to as co-synthesis (CoSyn). The proposed framework employs real-time resource allocation to coordinate the progression of concurrent cell analysis. Besides this framework, a probabilistic model based on a discrete-time Markov chain is also deployed to investigate protocol settings, where experimental conditions, such as sonication time, vary probabilistically among cell types. Simulation results show that CoSyn efficiently utilizes platform resources and outperforms baseline techniques.

PDMS-Parylene Hybrid, Flexible Microfluidics for Real-Time Modulation of 3D Helical Inertial Microfluidics.

Inertial microfluidics has drawn much attention for its applications for circulating tumor cell separations from blood. The fluid flows and the inertial particle focusing in inertial microfluidic systems are highly dependent on the channel geometry and structure. Flexible microfluidic systems can have adjustable 3D channel geometries by curving planar 2D channels into 3D structures, which will enable tunable inertial separation. We present a poly(dimethylsiloxane) (PDMS)-parylene hybrid thin-film microfluidic system that can provide high flexibility for 3D channel shaping while maintaining the channel cross-sectional shape. The PDMS-parylene hybrid microfluidic channels were fabricated by a molding and bonding technique using initiated chemical vapor deposition (iCVD) bonding. We constructed 3D helical inertial microfluidic channels by coiling a straight 2D channel and studied the inertial focusing while varying radius of curvature and Reynolds number. This thin film structure allows for high channel curvature and high Dean numbers which leads to faster inertial particle focusing and shorter channel lengths than 2D spiral channels. Most importantly, the focusing positions of particles and cells in the microchannel can be tuned in real time by simply modulating the channel curvature. The simple mechanical modulation of these 3D structure microfluidic systems is expected to provide unique advantages of convenient tuning of cell separation thresholds with a single device.

Paper-Plastic Hybrid Microfluidic Device for Smartphone-Based Colorimetric Analysis of Urine.

In this work, a disposable paper-plastic hybrid microfluidic lab-on-a-chip (LOC) has been developed and successfully applied for the colorimetric measurement of urine by the smartphone-based optical platform using a "UrineAnalysis" Android app. The developed device was cost-effectively implemented as a stand-alone hybrid LOC by incorporating the paper-based conventional reagent test strip inside the plastic-based LOC microchannel. The LOC device quantitatively investigated the small volume (40 μL) of urine analytes for the colorimetric reaction of glucose, protein, pH, and red blood cell (RBC) in integration with the finger-actuating micropump. On the basis of our experiments, the conventional urine strip showed large deviation as the reaction time goes by, because dipping the strip sensor in a bottle of urine could not control the reaction volume. By integrating the strip sensor in the LOC device for urine analysis, our device significantly improves the time-dependent inconstancy of the conventional dipstick-based urine strip, and the smartphone app used for image analysis enhances the visual assessment of the test strip, which is a major user concern for the colorimetric analysis in point-of-care (POC) applications. As a result, the user-friendly LOC, which is successfully implemented in a disposable format with the smartphone-based optical platform, may be applicable as an effective tool for rapid and qualitative POC urinalysis.

Editorial for Special Issue: Advances in Microfluidic Devices for Cell Handling and Analysis

Microfluidics is a technology that is expanding rapidly in many areas of research, especially in the biological areas of cell handling and analysis.[...]

Hybrid Integrated Microfluidic Channels on Multilayer Organic Substrate and on Copper for Tuning and Cooling an RF Reconfigurable S-/C-Band GaN-Based Power Amplifier

In this paper, a gallium nitride (GaN)-based power amplifier (PA) is cooled and tuned thanks to multifunctional hybrid microfluidic channels integrated on a multilayer organic substrate composed of liquid crystal polymer and ceramic-filled polytetrafluoroethylene. The tuning microchannels are integrated in the organic materials, while the cooling microchannels are micromachined in a copper carrier. The tuning allows the PA to switch the frequency of operation from 2.4 to 5.8 GHz and vice versa. Heat removal is achieved by the motion of distilled water below the GaN die, while tuning is achieved by swapping air and acetone inside the source and load matching network microchannels. The novel idea behind the design of such tunable PAs relies on the design of microfluidically tunable matching networks. For the first time, tunability parameters are introduced and a generic methodology is described for the design of microfluidically tunable matching networks for any dies, frequencies, fluids, or stackups. The measured power added efficiency (PAE) and output power (P out ) are: 1) PAE = 33.9% and P out = 37.1 dBm at 2.4 GHz with acetone and 2) PAE = 51.1% and P out = 37.9 dBm at 5.8 GHz with air. The liquid cooling seems to have no effect on P out for all measurements but shows a 1.5% increase in PAE for the 5.8-GHz measurements.

Hybrid microfluidics combined with active and passive approaches for continuous cell separation.

Microfluidics, which is classified as either active or passive, is capable of separating cells of interest from a complex and heterogeneous sample. Active methods utilise external fields such as electric, magnetic, acoustic, and optical to drive cells for separation, while passive methods utilise channel structures, intrinsic hydrodynamic forces, and steric hindrances to manipulate cells. However, when processing complex biological samples such as whole blood with rare cells, separation with a single module microfluidic device is difficult. Hybrid microfluidics is an emerging technique, which utilises active and passive methods whilst fulfilling higher requirements for stable performance, versatility, and convenience, including (i) the ability to process multi-target cells, (ii) enhanced ability for multiplexed separation, (iii) higher sensitivity, and (iv) tunability for a wider operational range. This review introduces the fundamental physics and typical formats for subclasses of hybrid microfluidic devices based on their different physical fields; presents current examples of cell sorting to highlight the advantage and usefulness of hybrid microfluidics on biomedicine, and then discusses the challenges and perspective of future development and the promising direction of research in this field.

Hybrid Microfluidic Platform for Multifactorial Analysis Based on Electrical Impedance, Refractometry, Optical Absorption and Fluorescence.

This paper describes the development of a novel microfluidic platform for multifactorial analysis integrating four label-free detection methods: electrical impedance, refractometry, optical absorption and fluorescence. We present the rationale for the design and the details of the microfabrication of this multifactorial hybrid microfluidic chip. The structure of the platform consists of a three-dimensionally patterned polydimethylsiloxane top part attached to a bottom SU-8 epoxy-based negative photoresist part, where microelectrodes and optical fibers are incorporated to enable impedance and optical analysis. As a proof of concept, the chip functions have been tested and explored, enabling a diversity of applications: (i) impedance-based identification of the size of micro beads, as well as counting and distinguishing of erythrocytes by their volume or membrane properties; (ii) simultaneous determination of the refractive index and optical absorption properties of solutions; and (iii) fluorescence-based bead counting.

Abstract 2705: A computational algorithm to predict tumor growth and cancer stem cell proportion in-vitro and in-vivo from single-cell observations

Abstract We have developed a new mathematical algorithm and corresponding computer software that uses observations of single cell behavior to predict the growth of the cancer stem-like cell proportion in larger cancer cell groups. Cancer stem-like cells (CSCs) have been implicated in ovarian cancer tumor growth, chemotherapy resistance, and disease recurrence. Aldehyde dehydrogenase (ALDH) is a primary discriminator of CSCs in ovarian cancer as well as in other cancer types. Unfortunately, the rarity and ability of CSCs to rapidly differentiate makes them difficult to study in-vitro and in-vivo. Microfluidic capture devices now allow us to grow and evaluate single ovarian cancer cells in isolated culture. We deployed microfluidic devices to evaluate the different self-renewal and asymmetric division patterns of ALDH+ and ALDH(-) ovarian CSCs. We analyzed data gathered from arrays of single cell chamber observations and found that purified ALDH+ cells were more proliferative than ALDH(-) cells in both cell-line (n = 112, p < 0.001) and primary (n = 41, p = 0.008) ovarian cancer specimens. Importantly, ALDH+ cells could produce both ALDH+ and ALDH(-) cells, whereas ALDH- cells were only able to produce ALDH- cells. Based on this hierarchy-defining data, we developed an easy to use computer algorithm on the freely-available software R to predict cancer cell population growth in-vitro and in-vivo. In our algorithm, size changes of cell populations are simulated by drawing from observed events. We compared the predictions from our hybrid microfluidics chip and computational algorithm with validation experiments in-vivo and in-vitro for both cell line and primary ovarian cancer cells. We found a superb correlation between observed and predicted in-vitro total and CSC counts for both cell line and primary ovarian cancer cells (correlation r = 0.98, p < 0.0001). Furthermore, this approach appropriately predicted changes in cell growth in the presence of the CSC-promoting growth factor EGFL6 both in vitro and in vivo, over time frames of up to 28 days. The single cell division based sampling algorithm developed here allows for rapid and inexpensive means to predict in-vivo ovarian tumor growth based on microfluidic chip culture and can easily be adapted to evaluate new therapeutic options across other cancer subtypes. Citation Format: Alexander T. Pearson, Patrick Ingram, Shoumei Bai, Euisik Yoon, Trachette Jackson, Ronald J. Buckanovich. A computational algorithm to predict tumor growth and cancer stem cell proportion in-vitro and in-vivo from single-cell observations. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2705.

Evaluation of alcohol dehydrogenase and aldehyde dehydrogenase enzymes as bi-enzymatic anodes in a membraneless ethanol microfluidic fuel cell

Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (AldH) enzymes were immobilized by covalent binding and used as the anode in a bi-enzymatic membraneless ethanol hybrid microfluidic fuel cell. The purpose of using both enzymes was to optimize the ethanol electro-oxidation reaction (EOR) by using ADH toward its direct oxidation and AldH for the oxidation of aldehydes as by-products of the EOR. For this reason, three enzymatic bioanode configurations were evaluated according with the location of enzymes: combined, vertical and horizontally separated. In the combined configuration, a current density of 16.3 mA cm-2, a voltage of 1.14 V and a power density of 7.02 mW cm-2 were obtained. When enzymes were separately placed in a horizontal and vertical position the ocp drops to 0.94 V and to 0.68 V, respectively. The current density also falls to values of 13.63 and 5.05 mA cm-2. The decrease of cell performance of bioanodes with separated enzymes compared with the combined bioanode was of 31.7% and 86.87% for the horizontal and the vertical array.

Perspective use of direct human blood as an energy source in air-breathing hybrid microfluidic fuel cells

This work presents a flexible and light air-breathing hybrid microfluidic fuel cell (HμFC) operated under biological conditions. A mixture of glucose oxidase, glutaraldehyde, multi-walled carbon nanotubes and vulcan carbon (GOx/VC-MWCNT-GA) was used as the bioanode. Meanwhile, integrating an air-exposed electrode (Pt/C) as the cathode enabled direct oxygen delivery from air. The microfluidic fuel cell performance was evaluated using glucose obtained from three different sources as the fuel: 5 mM glucose in phosphate buffer, human serum and human blood. For the last fuel, an open circuit voltage and maximum power density of 0.52 V and 0.20 mW cm−2 (at 0.38 V) were obtained respectively; meanwhile the maximum current density was 1.1 mA cm−2. Furthermore, the stability of the device was measured in terms of recovery after several polarization curves, showing excellent results. Although this air-breathing HμFC requires technological improvements before being tested in a biomedical device, it represents the best performance to date for a microfluidic fuel cell using human blood as glucose source.

Developing an ultra non-fouling SU-8 and PDMS hybrid microfluidic device by poly(amidoamine) engraftment

A new strategy to prepare an ultra non-fouling SU-8/poly(dimethylsiloxane) (PDMS) hybrid microfluidic system is presented. We report a simple method to bond both SU-8 and PDMS surfaces to prepare the hybrid microfluidic device and then simultaneously modify both SU-8 and PDMS surfaces using poly(amidoamine) (PAMAM), a highly hydrophilic dendrimer to enhance non-fouling properties of the hybrid microfluidic channel. Water contact angle, atomic force microscopy (AFM) and X-ray photoelectron spectroscopy (XPS) are used to characterize and confirm surface modifications. Finally the non-fouling performance of the resulting microfluidic system is tested using both microbeads and Escherichia coli O157:H7 bacterial cells. We demonstrate that the obtained hybrid microfluidic system shows a significant microbead adsorption suppression of 99.7%, an ultra non-fouling performance that has not been reported in the literature before; in addition, 95% bacteria surface adsorption suppression is also obtained. Considering the significantly improved non-fouling performance and the ease of preparing the hybrid microfluidic system, we anticipate that this strategy will find wide applications in biosensing and detection.

A versatile PDMS/paper hybrid microfluidic platform for sensitive infectious disease diagnosis.

Bacterial meningitis is a serious health concern worldwide. Given that meningitis can be fatal and many meningitis cases occurred in high-poverty areas, a simple, low-cost, highly sensitive method is in great need for immediate and early diagnosis of meningitis. Herein, we report a versatile and cost-effective polydimethylsiloxane (PDMS)/paper hybrid microfluidic device integrated with loop-mediated isothermal amplification (LAMP) for the rapid, sensitive, and instrument-free detection of the main meningitis-causing bacteria, Neisseria meningitidis (N. meningitidis). The introduction of paper into the microfluidic device for LAMP reactions enables stable test results over a much longer period of time than a paper-free microfluidic system. This hybrid system also offers versatile functions, by providing not only on-site qualitative diagnostic analysis (i.e., a yes or no answer), but also confirmatory testing and quantitative analysis in laboratory settings. The limit of detection of N. meningitidis is about 3 copies per LAMP zone within 45 min, close to single-bacterium detection sensitivity. In addition, we have achieved simple pathogenic microorganism detection without a laborious sample preparation process and without the use of centrifuges. This low-cost hybrid microfluidic system provides a simple and highly sensitive approach for fast instrument-free diagnosis of N. meningitidis in resource-limited settings. This versatile PDMS/paper microfluidic platform has great potential for the point of care (POC) diagnosis of a wide range of infectious diseases, especially for developing nations.

Hybrid Microfluidic Systems: Fabrication of a Hybrid Microfluidic System Incorporating both Lithographically Patterned Microchannels and a 3D Fiber‐Formed Microfluidic Network (Adv. Healthcare Mater. 2/2012)

Hybrid Microfluidic Systems: Fabrication of a Hybrid Microfluidic System Incorporating both Lithographically Patterned Microchannels and a 3D Fiber‐Formed Microfluidic Network (Adv. Healthcare Mater. 2/2012)

Fabrication of a Hybrid Microfluidic System Incorporating both Lithographically Patterned Microchannels and a 3D Fiber‐Formed Microfluidic Network

A device containing a 3D microchannel network (fabricated using sacrificial melt-spun microfibers) sandwiched between lithographically patterned microfluidic channels offers improved delivery of soluble compounds to a large volume compared to a simple stack of two microfluidic channel layers. With this improved delivery ability comes an increased fluidic resistance due to the tortuous network of small-diameter channels.