Digital Microfluidic System Research Articles

Intelligent droplet manipulation in electrowetting devices via capacitance-based sensing and actuation for self-adaptive digital microfluidics

In this work, we report on an integrated digital microfluidic system for precise sensing and automatic actuating droplets (ISPSAA) based on electrowetting on dielectric (EWOD). A feedback controller in the system is capable of locating, judging and controlling individual droplet independent of liquid composition. An integrated position sensor is capable of tracking the continuous displacement of a droplet between electrodes through the feedback of capacitance difference in real time, and thus improves the droplet control precision. Real-time droplet allocation and actuation have been achieved in a digital and precise way by combining the droplet sensing and driving functions. A defect electrode on the droplet transportation path can also be sensed and avoided in an intelligent way by adjusting corresponding electrodes and parameters to actuate the droplet to reach destination position based on the designed route with minimum number of actuated electrodes. Because of its portability, low cost, and adaptive functions, ISPSAA are helpful for further understanding and implementation of digital microfluidic systems for practical applications.

Extraction of Cell-free Dna from An Embryo-culture Medium Using Micro-scale Bio-reagents on Ewod

As scientific and technical knowledge advances, research on biomedical micro-electromechanical systems (bio-MEMS) is also developing towards lab-on-a-chip (LOC) devices. A digital microfluidic (DMF) system specialized for an electrowetting- on-dielectric (EWOD) mechanism is a promising technique for such point-of-care systems. EWOD microfluidic biochemical analytical systems provide applications over a broad range in the lab-on-a-chip field. In this report, we treated extraction of cell-free DNA (cf-DNA) at a small concentration from a mouse embryo culture medium (2.5 days & 3.5 days) with electro-wetting on a dielectric (EWOD) platform using bio-reagents of micro-scale quantity. For such extraction, we modified a conventional method of genomic-DNA (g-DNA) extraction using magnetic beads (MB). To prove that extraction of cf-DNA with EWOD was accomplished, as trials we extracted designed-DNA (obtained from Chang Gung Memorial Hospital (CGMH), Taiwan which shows properties similar to that of cf-DNA). Using that designed DNA, extraction with both conventional and EWOD methods has been performed; the mean percentage of extraction with both methods was calculated for a comparison. From the cycle threshold (Ct) results with a quantitative polymerase chain reaction (q-PCR), the mean extraction percentages were obtained as 14.8 percent according to the conventional method and 23 percent with EWOD. These results show that DNA extraction with EWOD appears promising. The EWOD extraction involved voltage 100 V and frequency 2 kHz. From this analysis, we generated a protocol for an improved extraction percentage on a EWOD chip and performed cf-DNA extraction from an embryo-culture medium (KSOM medium) at 3.5 and 2.5 days. The mean weight obtained for EWOD-extracted cf-DNA is 0.33 fg from the 3.5-day sample and 31.95 fg from the 2.5-day sample. All these results will pave a new path towards a renowned lab-on-a-chip concept.

Using a Digital Microfluidic System to Evaluate the Stretch Length of a Droplet with a L-DEP and Varied Parameters

Digital microfluidics has become intensively explored as an effective method for liquid handling in lab-on-a-chip (LOC) systems. Liquid dielectrophoresis (L-DEP) has many advantages and exciting prospects in driving droplets. To fully realize the potential benefits of this technique, one must know the droplet volume accurately for its distribution and manipulation. Here we present an investigation of the tensile length of a droplet subjected to a L-DEP force with varied parameters to achieve precise control of the volume of a droplet. Liquid propylene carbonate served as a driving liquid in the L-DEP experiment. The chip was divided into two parts: an electrode of width fixed at 0.1 mm and a total width fixed at 1 mm. Each had a variation of six electrode spacings. The experimental results showed that the stretching length decreased with decreasing electrode width, but the stretching length did not vary with an increased spacing of the electrode. When the two electrodes were activated, the length decreased because of an increase in electrode spacing. The theory was based on the force balance on a droplet that involved the force generated by the electric field, friction force, and capillary force. The theory was improved according to the experimental results. To verify the theoretical improvement through the results, we designed a three-electrode chip for experiments. The results proved that the theory is consistent with the results of the experiments, so that the length of a droplet stretched with L-DEP and its volume can be calculated.

Numerical and experimental study of acoustic wave propagation in glass plate/water/128YX-LiNbO3 structure

There is a need for digital microfluidic systems that can integrate pretreatments and sensors on the same substrates in the medical and biotechnology fields. Surface acoustic waves can be used to manipulate and mix droplets. Sensors integrated on the droplet-manipulating surfaces can measure droplet properties. The adhesion of liquids to the substrate surfaces necessitates the development of disposable devices. We therefore propose a three-layer structure of glass plate/water/128YX-LiNbO3. A disposable device can be realized by replacing the inexpensive glass. However, the details of the propagation mode in these structures have not yet been studied. In this paper, numerical analysis and experiments were carried out. For the numerical analysis, the Rayleigh–Lamb equations for the glass plate immersed in liquid were used. Comparisons between the numerical analysis and experimental results indicate that the symmetric mode of the Lamb wave propagates in the glass plate.

Multicenter Evaluation of a PCR-Based Digital Microfluidics and Electrochemical Detection System for the Rapid Identification of 15 Fungal Pathogens Directly from Positive Blood Cultures.

Routine identification of fungal pathogens from positive blood cultures by culture-based methods can be time-consuming, delaying treatment with appropriate antifungal agents. The GenMark Dx ePlex investigational use only blood culture identification fungal pathogen panel (BCID-FP) rapidly detects 15 fungal targets simultaneously in blood culture samples positive for fungi by Gram staining. We aimed to determine the performance of the BCID-FP in a multicenter clinical study. Blood culture samples collected at 10 United States sites and tested with BCID-FP at 4 sites were compared to the standard-of-care microbiological and biochemical techniques, fluorescence in situ hybridization using peptide nucleic acid probes (PNA-FISH) and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Discrepant results were analyzed by bi-directional PCR/sequencing of residual blood culture samples. A total of 866 clinical samples, 120 retrospectively and 21 prospectively collected, along with 725 contrived samples were evaluated. Sensitivity and specificity of detection of Candida species (C. albicans, C. auris, C. dubliniensis, C. famata, C. glabrata, C. guilliermondii, C. kefyr, C. krusei, C. lusitaniae, C. parapsilosis, and C. tropicalis) ranged from 97.1 to 100% and 99.8 to 100%, respectively. For the other organism targets, sensitivity and specificity were as follows: 100% each for Cryptococcus neoformans and C. gattii, 98.6% and 100% for Fusarium spp., and 96.2% and 99.9% for Rhodotorula spp., respectively. In 4 of the 141 clinical samples, the BCID-FP panel correctly identified an additional Candida species, undetected by standard-of-care methods. The BCID-FP panel offers a faster turnaround time for identification of fungal pathogens in positive blood cultures that may allow for earlier antifungal interventions and includes C. auris, a highly multidrug-resistant fungus.

Composite Films of CsPbBr3 Perovskite Nanocrystals in a Hydrophobic Fluoropolymer for Temperature Imaging in Digital Microfluidics.

A composite film material that combines CsPbBr3 perovskite nanocrystals with a Hyflon AD 60 fluoropolymer was developed and utilized for high-resolution optical temperature imaging. It exhibited bright luminescence and, most importantly, long-term stability in an aqueous medium. CsPbBr3 nanocrystal-Hyflon films immersed in aqueous solutions showed stable luminescence over at least 4 months and exhibited a fully reversible pronounced temperature sensitivity of 1.2% K-1 between 20 and 80 °C. They were incorporated into a digital microfluidic (electrowetting on dielectric) platform and were used for spatially resolved temperature measurements during droplet movements. Thermal mapping with a CsPbBr3 nanocrystal-Hyflon sensing layer in a room temperature environment (22.0 °C) revealed an increase in local temperatures of up to 40.2 °C upon voltage-driven droplet manipulations in a digital microfluidic system, corresponding to a local temperature change of up to 18.2 °C.

Position and feedback for digital microfluidic system based on light intensity information

AbstractThe digital microfluidic system based on electrowetting on dielectric is a new technology that can control microliter droplets within the plane. Because of layout, fabrication, and driving uncertainties, feedback control schemes must be designed and implemented to ensure safety and reliability of digital microfluidic system for practical application. The basis for feedback is to realize the accurate droplet location. So, there is a strong need for the development of a digital microfluidic system integrated with position and feedback scheme for innovative biological research. In this paper, we propose a position and feedback scheme based on light intensity for digital microfluidic system. The verified experiment results show that the proposed position and feedback scheme can position multiple droplets accurately and improve the success rate of droplets transport application by feedback.

An Impedance Sensing Platform for Monitoring Heterogeneous Connectivity and Diagnostics in Lab-on-a-Chip Systems.

Reliable hardware connectivity is vital in heterogeneous integrated systems. For example, in digital microfluidics lab-on-a-chip systems, there are hundreds of physical connections required between a microelectromechanical fabricated device and the driving system that can be remotely located on a printed circuit board. Unfortunately, the connection reliability cannot be checked or monitored by vision-based detection methods that are commonly used in the semiconductor industry. Therefore, a sensing platform that can be seamlessly integrated into existing digital microfluidics systems and provide real-time monitoring of multiconnectivity is highly desired. Here, we report an impedance sensing platform that can provide fast detection of a single physical connection in timescales of milliseconds. Once connectivity is established, the same setup can be used to determine the droplet location. The sensing system can be scaled up to support multiple channels or applied to other heterogeneously integrated systems that require real-time monitoring and diagnostics of multiconnectivity systems.

Digital Microfluidics for the Detection of Selected Inorganic Ions in Aerosols.

A prototype aerosol detection system is presented that is designed to accurately and quickly measure the concentration of selected inorganic ions in the atmosphere. The aerosol detection system combines digital microfluidics technology, aerosol impaction and chemical detection integrated on the same chip. Target compounds are the major inorganic aerosol constituents: sulfate, nitrate and ammonium. The digital microfluidic system consists of top and bottom plates that sandwich a fluid layer. Nozzles for an inertial impactor are built into the top plate according to known, scaling principles. The deposited air particles are densely concentrated in well-defined deposits on the bottom plate containing droplet actuation electrodes of the chip in fixed areas. The aerosol collection efficiency for particles larger than 100 nm in diameter was higher than 95%. After a collection phase, deposits are dissolved into a scanning droplet. Due to a sub-microliter droplet size, the obtained extract is highly concentrated. Droplets then pass through an air/oil interface on chip for colorimetric analysis by spectrophotometry using optical fibers placed between the two plates of the chip. To create a standard curve for each analyte, six different concentrations of liquid standards were chosen for each assay and dispensed from on-chip reservoirs. The droplet mixing was completed in a few seconds and the final droplet was transported to the detection position as soon as the mixing was finished. Limits of detection (LOD) in the final droplet were determined to be 11 ppm for sulfate and 0.26 ppm for ammonium. For nitrate, it was impossible to get stable measurements. The LOD of the on-chip measurements for sulfate was close to that obtained by an off-chip method using a Tecan spectrometer. LOD of the on-chip method for ammonium was about five times larger than what was obtained with the off-chip method. For the current impactor collection air flow (1 L/min) and 1 h collection time, the converted LODs in air were: 0.275 μg/m3 for sulfate, 6.5 ng/m3 for ammonium, sufficient for most ambient air monitoring applications.

A digital microfluidic system with 3D microstructures for single-cell culture

Despite the precise controllability of droplet samples in digital microfluidic (DMF) systems, their capability in isolating single cells for long-time culture is still limited: typically, only a few cells can be captured on an electrode. Although fabricating small-sized hydrophilic micropatches on an electrode aids single-cell capture, the actuation voltage for droplet transportation has to be significantly raised, resulting in a shorter lifetime for the DMF chip and a larger risk of damaging the cells. In this work, a DMF system with 3D microstructures engineered on-chip is proposed to form semi-closed micro-wells for efficient single-cell isolation and long-time culture. Our optimum results showed that approximately 20% of the micro-wells over a 30 × 30 array were occupied by isolated single cells. In addition, low-evaporation-temperature oil and surfactant aided the system in achieving a low droplet actuation voltage of 36V, which was 4 times lower than the typical 150 V, minimizing the potential damage to the cells in the droplets and to the DMF chip. To exemplify the technological advances, drug sensitivity tests were run in our DMF system to investigate the cell response of breast cancer cells (MDA-MB-231) and breast normal cells (MCF-10A) to a widely used chemotherapeutic drug, Cisplatin (Cis). The results on-chip were consistent with those screened in conventional 96-well plates. This novel, simple and robust single-cell trapping method has great potential in biological research at the single cell level.

Minimal microfabrication required digital microfluidic system toward point-of-care nucleic acid amplification test application for developing countries

Digital Microfluidics (DMF) has potentially been a favorable platform for point-of-care molecular diagnostics. However, its fabrication process normally relies heavily on expensive microfabrication facilities and equipment. In this work, we developed a portable, small footprint DMF device that requires minimal microfabrication work (only a small clean bench and a DIY spin coater were required) but is still capable of performing nucleic acid amplification tests. The DMF system implemented in this work could perform successfully the Loop-mediated isothermal Amplification (LAMP) reaction with samples containing extracted DNA of Plasmodium falciparum that causes Malaria in human. The LAMP reaction was conducted at the optimal temperature of 65 °C in 45 min with the total reaction volume of 1.5 µL. The results of LAMP were designed to be observable to naked eyes via the color change from pink to yellowish orange, which simplifies the readout step and eliminates the necessity for external and bulky result-reading equipment.

P.325 Reward anticipation processing in major depressive disorder and prediction of treatment response - a functional magnetic resonance imaging study

DNA hybridization kinetics has been playing a critical role in molecular diagnostics for binding discrimination, but its study on digital microfluidic (DMF) systems is ultimately restrained by the laminar flow condition. The kinetic mixing technique is widely employed to ensure a fast reaction rate, but poses intrinsic risk in cross contamination and exhibits instable fluorescence intensity during the droplet transportation. While the electrothermal technique can provide stationary droplet mixing through the established thermal gradient within the hybridization solution, the significant increase in the droplet temperature will inevitably undermine the hybridization equilibrium and jeopardize the binding discrimination. To enhance the hybridization efficiency while ensuring a stable droplet temperature (within ±0.1℃), this paper presents a DMF platform that can perform isothermal hydrodynamic-flow-enhanced droplet mixing. Specifically, with a single electrode, droplet-boundary oscillation under a slow AC actuation is studied for improving the reaction rate. The dependencies between the mixing efficiency and the actuation voltage, actuation frequency and the spacer thickness are also systematically studied. Reliable mixing efficiency improvement is further validated over a wide range of solute concentrations. The results from real-time on-chip DNA hybridization kinetics with stationary droplets using the complete sandwiched DMF system shows that the proposed rapid mixer can achieve the same hybridization equilibrium with >13 times faster reaction rate when compared to the reference one through pure diffusion, while preventing biased hybridization kinetics as demonstrated in the electrothermal technique.

Quantitative measurements of inorganic analytes on a digital microfluidics platform

Two methods were studied for selectively measuring the on-chip absorbance of trace sulfate analytes in droplets on a digital microfluidics (DMF) platform. In one method, the direction of measurement was perpendicular to the flat upper and lower surfaces of the DMF platform (vertical), and in the second method, the measurement direction was parallel to the DMF platform surfaces (horizontal). The channel height or the vertical light path length was 0.24 mm, and the droplet diameter was 1 mm. The DMF system employed a silicone oil transport medium whereby a thin, non-uniform oil layer formed between the droplet and the upper/lower plates which was unstable, resulting in randomly formed local oil lenses. The mobile oil lenses caused vertical absorbance measurement errors and uncertainties. The effects of the oil lenses were verified by simulation. Horizontal absorbance measurements were taken with embedded optical fibers (0.2 mm in diameter) aligned over the bottom chip surface in contact with the sides of the droplet, resulting in a horizontal light path length approximately three times that of the vertical light path. Because no oil lenses could form on the droplet’s sides, the stability of repeated horizontal measurements outperformed repeated vertical measurements made on the same droplet and on multiple droplets actuated into the measurement positions. Comparisons were based on measurement standard deviations and limits of detection (LOD). The following LODs and measurement standard deviations were achieved for horizontal measurements of multiple sulfate concentrations in 1.5 µl droplets: 7 ppm for sulfate (0.3–2.7%) and an R2 value of 0.957 from a least square data fit. Measurements on a commercial plate reader gave comparable results (200 µl liquid in each well, LOD equals 11 ppm, CV equals to 0.2–4%), even though the absorbance path was larger (0.7 mm). This LOD value means that the chip could detect 10.5 ng of sulfate. LOD values on vertical measurements were also similar, but large measurement errors from numerous outlier points yielded an R2 value of 0.735 and large average measurement standard deviations (36%).

Deformation, speed, and stability of droplet motion in closed electrowetting-based digital microfluidics

Electrowetting-based microdrop manipulation has received considerable attention owing to its wide applications in numerous scientific areas based on the digital microfluidics (DMF) technology. However, the techniques for highly precise droplet handling in such microscopic systems are still unclear. In this work, the deformation, speed, and stability of droplet transporting in closed electrowetting-based DMF systems are comprehensively investigated with both theoretical and numerical analyses. First, a theoretical model is derived which governs the droplet motion and includes the influences of the key electrowetting system parameters. After that, a finite volume formulation with a two-step projection method is used for solving the microfluidic flow on a fixed numerical domain. The liquid-gas interface of the droplet is tracked by a coupled level-set and volume-of-fluid method, and the surface tension at the interface is computed by the continuum surface force scheme. A parametric study has been carried out to examine the effects of the static contact angles (θs,ON and θs,OFF), hysteresis effect (Δθ), channel height (H), and electrode size (LE) on droplet shape, speed, and deformation during transport, which unanimously shows that droplet length, neck width, and transport stability are directly related to a dimensionless parameter κ* that only comprises θs,ON, θs,OFF, H, LE, and the hysteresis angle Δθ. Based on the results, the scaling laws for estimating droplet shape and stability of the transport process have been developed, which can be used for promoting the accuracy and efficiency of droplet manipulation in a large variety of droplet-based DMF applications.

A fucosyltransferase inhibition assay using image-analysis and digital microfluidics.

Sialyl-LewisX and LewisX are cell-surface glycans that influence cell-cell adhesion behaviors. These glycans are assembled by α(1,3)-fucosyltransferase enzymes. Their increased expression plays a role in inflammatory disease, viral and microbial infections, and cancer. Efficient screens for specific glycan modifications such as those catalyzed by fucosyltransferases are tended toward costly materials and large instrumentation. We demonstrate for the first time a fucosylation inhibition assay on a digital microfluidic system with the integration of image-based techniques. Specifically, we report a novel lab-on-a-chip approach to perform a fluorescence-based inhibition assay for the fucosylation of a labeled synthetic disaccharide, 4-methylumbelliferyl β-N-acetyllactosaminide. As a proof-of-concept, guanosine 5'-diphosphate has been used to inhibit Helicobacter pylori α(1,3)-fucosyltransferase. An electrode shape (termed "skewed wave") is designed to minimize electrode density and improve droplet movement compared to conventional square-based electrodes. The device is used to generate a 10 000-fold serial dilution of the inhibitor and to perform fucosylation reactions in aqueous droplets surrounded by an oil shell. Using an image-based method of calculating dilutions, referred to as "pixel count," inhibition curves along with IC50 values are obtained on-device. We propose the combination of integrating image analysis and digital microfluidics is suitable for automating a wide range of enzymatic assays.

Additively Manufactured Digital Microfluidic Platforms for Ion-Selective Sensing.

Digital microfluidic (DMF) sensors integrated with circuit systems have been applied to a broad range of applications including biology, medicine, and chemistry. Compared with the conventional microfluidic devices that require extra liquid as a carrier and a complex pumping system to operate, DMF is an ideal platform for ion-selective sensing as it enables the droplet operation in a discrete, accurate, and automatic way. However, it is quite rare that DMF platform is utilized for the ion-selective detection. In this paper, we report an integrated DMF system which combines DMF and ion-selective sensing for facile blending of multiple ions, and detection of targeted primary ion. The platform is fabricated through an additive manufacturing method, together with the real-time droplet's motion monitoring feedback system. Thus, the fabricated system demonstrates controlled droplet manipulation ability including droplet actuation, mixing, and speed control. Targeted primary ion is selectively detected under concentration range from 10-6 to 1 M. The interference study with blended ions has been investigated through on-chip ion selective membranes.

An Efficient Multiple Fault Detection Technique in Digital Microfluidic Biochips

ABSTRACT The involvement of Digital Microfluidic Biochips (DMFBs) in the field of disease detection, automated drug discovery, on-chip DNA (Deoxyribonucleic acid) analysis has become well-accepted during last decades. Though trustworthiness of Digital Microfluidic Biochip system plays a vital role in point-of-care diagnosis, defective electrodes are the main reasons for various misleading assay performances. It also affects overall assay completion time; even it leads to discard a chip in the worst scenario. Hence, fault detection is a crucial need. In this paper, an efficient fault detection mechanism is formulated to identify multiple numbers of defective/faulty electrodes on an m × n biochip array, where m and n can be of any positive number. Test droplets are strategically routed throughout an arbitrary sized electrode array and capacitive sensing circuit is deployed to take decision about existence of faults. The basic idea of the proposed technique is to identify actual positions of faulty electrodes through boundary test, row test, and diagonal test. In some complicated cases, instead of finding exact faulty location(s), through an extended testing our proposed technique identifies faulty zone(s) (it is a bounded region where one or more defective electrodes reside) to ensure safe assay performance on a chip. A detailed study evaluates the proposed approach considering different faulty environments.

Hydrodynamic-flow-enhanced rapid mixer for isothermal DNA hybridization kinetics analysis on digital microfluidics platform

DNA hybridization kinetics has been playing a critical role in molecular diagnostics for binding discrimination, but its study on digital microfluidic (DMF) systems is ultimately restrained by the laminar flow condition. The kinetic mixing technique is widely employed to ensure a fast reaction rate, but poses intrinsic risk in cross contamination and exhibits instable fluorescence intensity during the droplet transportation. While the electrothermal technique can provide stationary droplet mixing through the established thermal gradient within the hybridization solution, the significant increase in the droplet temperature will inevitably undermine the hybridization equilibrium and jeopardize the binding discrimination. To enhance the hybridization efficiency while ensuring a stable droplet temperature (within ±0.1℃), this paper presents a DMF platform that can perform isothermal hydrodynamic-flow-enhanced droplet mixing. Specifically, with a single electrode, droplet-boundary oscillation under a slow AC actuation is studied for improving the reaction rate. The dependencies between the mixing efficiency and the actuation voltage, actuation frequency and the spacer thickness are also systematically studied. Reliable mixing efficiency improvement is further validated over a wide range of solute concentrations. The results from real-time on-chip DNA hybridization kinetics with stationary droplets using the complete sandwiched DMF system shows that the proposed rapid mixer can achieve the same hybridization equilibrium with >13 times faster reaction rate when compared to the reference one through pure diffusion, while preventing biased hybridization kinetics as demonstrated in the electrothermal technique.

Self-Cleaning: From Bio-Inspired Surface Modification to MEMS/Microfluidics System Integration.

This review focuses on self-cleaning surfaces, from passive bio-inspired surface modification including superhydrophobic, superomniphobic, and superhydrophilic surfaces, to active micro-electro-mechanical systems (MEMS) and digital microfluidic systems. We describe models and designs for nature-inspired self-cleaning schemes as well as novel engineering approaches, and we discuss examples of how MEMS/microfluidic systems integrate with functional surfaces to dislodge dust or undesired liquid residues. Meanwhile, we also examine “waterless” surface cleaning systems including electrodynamic screens and gecko seta-inspired tapes. The paper summarizes the state of the art in self-cleaning surfaces, introduces available cleaning mechanisms, describes established fabrication processes and provides practical application examples.

Electrowetting-on-dielectric chip with integrated screen-printed electrochemical sensor for rapid chemical analysis

In this work, a screen-printed electrochemical detector was fabricated on a digital electrowetting-on- dielectric (EWOD) microfluidic chip for the first time and employed for rapid chemical analysis. The digital microfluidic system consisted of a T-junction EWOD platform for merging buffer and analyte droplets and an electrochemical detector at the end of T-junction. The detector comprised three electrodes including carbon working, carbon counter and silver/silver chloride reference electrodes. The system was successfully fabricated by microfabrication and screen-printing processes and tested for electrochemical analyses of potassium hexacyanoferrate and hydrogen peroxide. The analyte and buffer droplets were controlled to mix with different concentrations and detected at the electrochemical detector using cyclic voltammetry. The results showed good detection performances with medium sensitivity, moderate detection limits, high stability and good reproducibility. In addition, the total analysis time including droplet mixing and CV measurement was shorter than 14 s. Therefore, the EWOD chip with integrated screen-printed carbon electrodes was a promising candidate for rapid automated chemical analysis.