Electrochemical Paper-based Analytical Device Research Articles

A new electrochemical paper platform for detection of 8-hydroxyquinoline in cosmetics using a cobalt phthalocyanine-modified screen-printed carbon electrode

An electrochemical paper-based analytical device (ePAD) using an electrocatalytic cobalt(II) phthalocyanine-modified screen-printed carbon electrode (CoPc-SPCE) was first developed for the determination of 8-hydroxyquinoline (8-HQ). The electrochemical behavior of 8-HQ on the CoPc-modified electrode was investigated by cyclic voltammetry (CV) and square-wave voltammetry (SWV). The obtained results demonstrated that the CoPc-SPCE shows high electrocatalytic activity toward the oxidation of 8-HQ. The oxidation potential was shifted to a negative value compared to those obtained from the unmodified electrode. Under the optimized experimental conditions, a good linear relationship between the 8-HQ concentration and the peak current was obtained within the range of 10–250μM with a detection limit of 0.89μM. Furthermore, the method was successfully applied for the determination of 8-HQ in cosmetic samples. According to a paired t-test, the results were in good agreement with results from the standard HPLC-UV method. The benefits of this proposed device are that it is easily fabricated, low cost, disposable, highly sensitive and reproducible.

Fast and flexible strategy to produce electrochemical paper-based analytical devices using a craft cutter printer to create wax barrier and screen-printed electrodes

This paper describes a simple, low-cost, and highly flexible rapid prototyping method to construct electrochemical paper-based analytical device (ePAD) for multiplexed analyte determinations. The ePAD was composed of two electrochemical cell (EC) compartments, separated by hydrophobic barriers of wax, and screen-printed electrodes (SPEs) deposited directly over the surface of the filter paper. The ePAD was entirely constructed using an inexpensive craft cutter printer with no needed of a wax printer. The rapid prototyping method involves two steps: the deposition of the SPEs and the creation of the wax barriers. In this case, the SPEs were screen-printed on filter paper by using adhesive tape as mask by cutting the electrodes pattern with the cutter printer. Following, the wax barriers were created using stamps made of filter paper also cut with the printer and impregnated with wax. In the ePAD, each ECs containing an array of 4-working electrodes, allowing up to 4 replicates in a single measurement. Both ECs shared one counter and one reference electrodes, permitting the simultaneous multianalysis. The ePAD was successfully applied to simultaneous detection of paracetamol (PAR), caffeine (CAF), and ascorbic acid (AA) in drugs. PAR and CAF were detected in a sample using one EC and AA was detected, in a different sample, on the other EC, both with no chemical modifications in the working electrodes. Limits of detection of 0.04 mmol L−1 for PAR, 0.22 mmol L−1 for CAF, and 0.40 mmol L−1 for AA were obtained. The construction process proposed provide an easy way to implement screen-printing electrodes and wax barriers in filter paper to create electrochemical devices for fast and simultaneous multianalysis.

Micro-patterning of single-walled carbon nanotubes and its surface modification with gold nanoparticles for electrochemical paper-based non-enzymatic glucose sensor

We developed a novel micro-patterning technique for single-walled carbon nanotubes (SWCNTs) on nitrocellulose (NC) membranes and characterized the mechanical and electrical properties of the patterned SWCNTs. The SWCNT patterns were successfully fabricated on wax-printed NC membrane by a vacuum filtration method within 15 min. The well-deposited SWCNT layer on the NC membrane exhibited strong adhesion and high conductivity due to the interconnection of the SWCNT layer in the porous structure of the NC membrane. The distinguishable patterns were resolved down to approximately 100 μm. The patterned SWCNT had an average resistivity of less than 100 Ω/sq and showed excellent mechanical properties in repeated bending tests. The suitability of the patterned SWCNT electrodes for electronic devices was demonstrated successfully via a simple light emitting diode array. Moreover, for the application of the patterned SWCNT to electrochemical paper-based analytical devices (ePADs), the gold nanoparticles (AuNPs) were directly deposited on the SWCNT electrode for non-enzymatic glucose determination. The oxidation peak had a linear response with glucose concentration in the range between 0.5 and 10 mM. The ePADs exhibited a sensitivity of 240 μA/mM·cm2 and a detection limit of 148 μM, which was derived from the electro-catalytic property of AuNPs toward glucose. Based on these results, we believe that the developed SWCNT patterning technique on paper substrates has great potential in ePADs.

Development of an Electrochemical Paper-Based Analytical Device for Trace Detection of Virus Particles.

Viral pathogens are a serious health threat around the world, particularly in resource limited settings, where current sensing approaches are often insufficient and slow, compounding the spread and burden of these pathogens. Here, we describe a label-free, point-of-care approach toward detection of virus particles, based on a microfluidic paper-based analytical device with integrated microwire Au electrodes. The device is initially characterized through capturing of streptavidin modified nanoparticles by biotin-modified microwires. An order of magnitude improvement in detection limits is achieved through use of a microfluidic device over a classical static paper-based device, due to enhanced mass transport and capturing of particles on the modified electrodes. Electrochemical impedance spectroscopy detection of West Nile virus particles was carried out using antibody functionalized Au microwires, achieving a detection limit of 10.2 particles in 50 μL of cell culture media. No increase in signal is found on addition of an excess of a nonspecific target (Sindbis). This detection motif is significantly cheaper (∼$1 per test) and faster (∼30 min) than current methods, while achieving the desired selectivity and sensitivity. This sensing motif represents a general platform for trace detection of a wide range of biological pathogens.

Glucose biosensor based on disposable electrochemical paper-based transducers fully fabricated by screen-printing

This paper describes a new approach for the massive production of electrochemical paper-based analytical devices (ePADs). These devices are fully fabricated by screen-printing technology and consist of a lineal microfluidic channel delimited by hydrophobic walls (patterned with diluted ultraviolet screen-printing ink in chromatographic paper grade 4) and a three-electrode system (printed with carbon and/or Ag/AgCl conductive inks). The printing process was characterised and optimized for pattern each layer with only one squeeze sweep. These ePADs were used as transducers to develop a glucose biosensor. Ionic strength/pH buffering salts, electrochemical mediator (ferricyanide) and enzyme (glucose dehydrogenase FAD-dependent) were separately stored along the microfluidic channel in order to be successively dissolved and mixed after the sample dropping at the entrance. The analyses required only 10 µl and the biosensors showed good reproducibility (RSD = 6.2%, n = 10) and sensitivity (0.426 C/M cm2), wide linear range (0.5–50 mM; r2 = 0.999) and low limit of detection (0.33 mM). Furthermore, the new biosensor was applied for glucose determination in five commercial soft-drinks without any sample treatment before the analysis. These samples were also analysed with a commercial enzymatic-kit assay. The results indicated that both methods provide accurate results.

A new tactics for the detection of S. aureus via paper based geno-interface incorporated with graphene nano dots and zeolites

Staphylococcus aureus (S. aureus) is a pathogenic bacteria which causes infectious diseases and food poisoning. Current diagnostic methods for infectious disease require sophisticated instruments, long analysis time and expensive reagents which restrict their application in resource-limited settings. Electrochemical paper based analytical device (EPAD) was developed by integrating graphene nano dots (GNDs) and zeolite (Zeo) using specific DNA probe. The ssDNA/GNDs-Zeo modified paper based analytical device (PAD) was characterized using cyclic voltammetry (CV) and differential pulse voltammetry (DPV). The genosensor was optimized at pH7.4 and incubation temperature of 30°C. A linear current response with respect to target DNA concentrations was obtained. The limit of detection (LOD) of the proposed sensor was found out to be 0.1nM. The specificity was confirmed by introducing non-complimentary target DNA to ssDNA/GNDs-Zeo modified PAD. The suitability of the proposed EPAD genosensor was demonstrated with fruit juice samples mixed with S. aureus. The proposed EPAD genosensor is a low cost, highly specific, easy to fabricate diagnostic device for detection of S. aureus bacteria which requires very low sample volume and minimum analysis time of 10s.

Paper-Based Microfluidic Device with Integrated Sputtered Electrodes for Stripping Voltammetric Determination of DNA via Quantum Dot Labeling.

This work reports a microfabricated electrochemical paper-based analytical device (ePAD) for the voltammetric determination of DNA. The device is patterned by wax-printing on paper and features a circular assay zone connected to an inlet zone and a sink via grooved microfluidic channels for accelerated flow rate. An electrochemical cell with integrated electrodes is formed on the reverse side of the paper by sputtering of thin metal films (Sn, Pt and Ag as the working, counter and reference electrode, respectively). Proof-of-principle of the ePAD for biosensing is demonstrated for a DNA assay involving attachment of capture DNA, hybridization with biotinylated target oligonucleotide and labeling with streptavidin-conjugated CdSe/ZnS quantum dots (QDs). After the acidic dissolution of the QDs, the released Cd(II) is quantified by anodic stripping voltammetry (ASV) at the Sn-film working electrode. Thanks to the synergistic effects of QDs amplification, the inherent sensitivity of ASV and the excellent detection capabilities of the Sn-film working electrode for Cd(II), the target DNA can be detected at levels as low as 0.11 pmol L-1 using sample volumes as low as 1 μL. The developed microfluidic ePAD costs only 0.11$ and presents favorable fabrication and operational features that make it an excellent candidate biosensor for simple and ultrasensitive point-of-need testing.

A three-dimensional electrochemical paper-based analytical device for low-cost diagnostics.

Paper-based microfluidic devices with screen-printed electrodes (SPEs) for electrochemical sensing are popular for low-cost point-of-care diagnostics. The electroactive sensing area in these devices is always the irregular, bottom-SPE surface which is in contact with the analyte flowing within the paper substrate. Unfortunately, this results in an electroactive area which varies widely from sensor to sensor. In this paper, we present a three-dimensional paper-based analytical device with a hollow 3D fluid reservoir which allows for use of a more uniform top-SPE surface as the electroactive sensing area. The use of this isolated reservoir eliminates the need for dielectric inks used in conventional SPEs on paper. Our sensors are fabricated using a combination of wax-printing, screen-printing and simple folding via a cleanroom free process without the need for expensive equipment. Additionally, for the first time, we demonstrate an electrochemical paper-based analytical device with a custom designed potentiostat integrated circuit (IC) as a miniaturized reader. The versatility of the sensor is demonstrated through voltammetric, amperometric and potentiometric measurements of important biochemical analytes such as dopamine, glucose and pH. The 3D ePAD together with a custom CMOS potentiostat demonstrates a low-cost, versatile, self-contained system suitable for point-of-care diagnostic devices.

Determination of Ascorbic Acid in Commercial Tablets Using Pencil Drawn Electrochemical Paper-based Analytical Devices

This study describes the use of electrochemical paper-based analytical devices (ePADs) drawn with graphite pencil for the determination of ascorbic acid (AA) in commercial tablets. ePADs were fabricated using vegetal paper and graphite pencil. First, the three-electrode electrochemical cell drawn using a graphical software and toner lines were laser printed on the vegetal paper surface to delimit the electrode areas. Then, the electrode regions were manually painted with graphite pencil. Afterwards, the pseudo-reference electrode was defined with the deposition of silver ink over the graphite surface. Cyclic voltammetry and square wave voltammetry (SWV) experiments were performed to optimize the electroanalytical parameters as well as to quantitatively determine the AA concentration in two commercial tables. ePADs exhibited linear behavior for a concentration range between 0.5 and 3.0 mmol L-1. The achieved limit of detection and sensitivity were 70 μmol L-1 and 0.47 μA/mmol L-1, respectively. The AA concentration levels found by SWV experiments in both CenevitTM and Energil CTM were 2.80 ± 0.02 and 3.10 ± 0.01 mmol L-1, respectively. The accuracy of the proposed devices was investigated through recovery experiments in three concentration levels and it presented values between 95 and 115%.

Paper-based device with a sputtered tin-film electrode for the voltammetric determination of Cd(II) and Zn(II)

This work describes a microfabricated electrochemical paper-based analytical device (ePAD) for the simultaneous voltammetric determination of Cd(II) and Zn(II) at trace levels. The ePAD integrates a wax-printed microfluidic channel and, on the reverse side of the paper, sputtered thin films of Sn, Pt and Ag serving as the working, counter and reference electrodes, respectively. The microfluidic channel ensures electrolytic contact between the electrodes with only 10 μL of working solution. The determination of Cd(II) and Zn(II) is performed at the 400 nm-thick sputtered Sn-film working electrode, in a pump-free and unstirred mode, by anodic stripping voltammetry. Under the selected conditions, the limits of detection were 0.9 μg L−1 Cd(II) and 1.1 μg L−1 Zn(II) and the between-sensor reproducibility was 5.7% for Cd(II) and 7.5% for Zn(II) at the 30 μg L−1 level (n = 5). The ePAD costs only 0.11$ and presents favorable fabrication and operational features, that make it suitable for simple, fast and on-site analysis.

Portable bioactive paper based genosensor incorporated with Zn-Ag nanoblooms for herpes detection at the point-of-care

The present work describes the fabrication of an electrochemical paper-based analytical device (EPAD) integrated with Zn-Ag nanoblooms for detection of herpes in human, caused by Herpes virus 5(HHV-5) DNA, at the point of care. The cyclic voltammetry(CV) was used for electrochemical detection of the HHV-5 DNA in infected patient samples. The EPAD exihibited optimum current response within 5s at pH7.0 and 35°C with two dynamic linear/working ranges, 113–103 and 3×105–106copies/mL and detection limit of 97copies/mL. The device showed high selectivity, repeatability, and sensitivity. The device had many advantageous features such as portable, facile approach, economical and potential for commercialization. The proposed sensing scheme is highly feasible for future clinical detection of Herpes virus.

Electrochemical Microfluidic Paper-Based Device for Virus-Sized Particle Detection

Over the last few years, the dramatic proliferation of emerging viruses such as Zika has resulted in a pressing need to develop inexpensive point-of-care diagnostic devices for rapid, sensitive, and accurate screening of patients. The ability to diagnose active infections on-site can improve patient compliance, allow physicians to tailor treatment options, and help prevent transmission to uninfected people. While commercially available point-of-care tests exist for bacterial infections, there are currently no approved point-of-care diagnostic assays for use in health clinics or in the field that detect viral infections. We are developing a point-of-care diagnostic device for viral detection that is inexpensive and can carry out all assay steps in an automated format. Our earlier work characterized electrochemical paper-based analytical devices (ePADs) coupled with metallic microwire electrodes as a disposable sensing system suitable for this application.1,2 Toward development of virus particle sensors, we have functionalized the electrodes with bioaffinity reagents to drive specific analyte binding, optimized electrode-integrated paper-based microfluidic platforms, and employed electrochemical impedance spectroscopy (EIS) to detect low levels of virus-sized particles. As shown in the figure, the functionalized electrode enables the binding of analyte to its surface and forms an insulating layer, increasing the impedance at the surface. In a proof of concept study, biotin-modified electrodes were incubated with streptavidin conjugated beads of size similar to viruses and investigated as a representative specific binding system. Two different electrode systems have been investigated. In the first, freestanding Au microwires were used because they can be easily modified with specific recognition elements using well-established coupling techniques prior to incorporation into the ePAD. As an alternative, Au nanoparticles were electrodeposited on screen-printed carbon electrodes, functionalized via the same chemistry, and compared for detection limits, sensitivities, and linear ranges. Preliminary optimization tests were achieved in a paper-based wax-confined static well fabricated as previously described.1 After demonstrating detection in static solutions, we tested the system using a flow-through ePAD system2 for improved sensitivity. To create steady-state flow, a fan-shaped passive pump was connected to the channel inlet and the microwire electrodes were embedded between two paper layers for electrochemical analysis. The two layers of paper create a channel and generate relatively high flow rates and more effective mass transport to the electrode surfaces. Key elements of flow rate, paper/electrode geometry, and number of electrodes were optimized for the method. Furthermore, multiple wires modified with the same affinity group were added in an overlapping, parallel configuration to increase surface area and enhance the signal. Sensor response was characterized with EIS using ferri/ferrocyanide as the mediator before and after analyte incubation. EIS provides significantly lower detection limits compared to traditional colorimetric lateral flow devices, while being inexpensive to manufacture and readily multiplexed for multiple analyte detection. The optimized electrochemical affinity assay makes ePAD particularly promising to rapidly detect virus-size particles in a multiplexed sample with high sensitivity, selectivity, and stability. (1) Adkins, J. A.; Henry, C. S. Analytica Chimica Acta 2015, 891, 247-254. (2) Adkins, J. A.; Noviana, E.; Henry, C. S. Anal Chem 2016, 88, 10639-10647. Figure 1

Development of a Quasi-Steady Flow Electrochemical Paper-Based Analytical Device.

An electrochemical paper-based analytical device (ePAD) was developed for quasi-steady flow detection at microwire electrodes, for the first time. The device implements a fan shaped geometry connected to an analysis channel whereby solution is pulled from an inlet, through a channel, and into the steadily increasing capillary network of the fan. The network counteracts the decrease in solution flow rate associated with increasing viscosity within the channel, generating quasi-steady flow within the analysis channel. Microwire electrodes were embedded between two paper layers within the analysis channel, such that solution flow occurred on both sides of the wire electrodes. The quasi-steady flow ePAD increased the current by 2.5 times and 0.7 times from a saturated channel with no flow and from a single-layer paper device with flow, respectively. Amperometric detection was used for flow injection analysis (FIA) of multiple analytes at both Au and Pt microwire working electrodes, both of which provided similar sensitivity (ca. 0.2 mM-1) when normalized to the same standard. The two-layer paper devices provided a detection limit of 31 μM for p-aminophenol (PAP) using Pt electrodes and was also used to detect enzyme activity for the reaction of β-galactosidase with p-aminophenyl-galactopyranoside (PAPG). Measured enzyme kinetics provided similar Vmax (0.079 mM/min) and Km (0.36 mM) values as those found in the literature. This device shows great promise toward use in enzyme-linked immunosorbent assays or other analytical techniques where flow or washing steps are necessary. The developed sensor provides a simple and inexpensive device capable of performing multiple injection analysis with steady-flow and online detection that would normally require an external pump to perform.

A Paper-Based "Pop-up" Electrochemical Device for Analysis of Beta-Hydroxybutyrate.

This paper describes the design and fabrication of a "pop-up" electrochemical paper-based analytical device (pop-up-EPAD) to measure beta-hydroxybutyrate (BHB)-a biomarker for diabetic ketoacidosis-using a commercial combination BHB/glucometer. Pop-up-EPADs are inspired by pop-up greeting cards and children's books. They are made from a single sheet of paper folded into a three-dimensional (3D) device that changes shape, and fluidic and electrical connectivity, by simply folding and unfolding the structure. The reconfigurable 3D structure makes it possible to change the fluidic path and to control timing; it also provides mechanical support for the folded and unfolded structures that enables good registration and repeatability on folding. A pop-up-EPAD designed to detect BHB shows performance comparable to commercially available plastic test strips over the clinically relevant range of BHB in blood when used with a commercial glucometer that integrates the ability to measure glucose and BHB (combination BHB/glucometer). With simple modifications of the electrode and the design of the fluidic path, the pop-up-EPAD also detects BHB in buffer using a simple glucometer-a device that is more available than the combination BHB/glucometer. Strategies that use a "3D pop-up"-that is, large-scale changes in 3D structure and fluidic paths-by folding/unfolding add functionality to EPADs (e.g., controlled timing, fluidic handling and path programming, control over complex sequences of steps, and alterations in electrical connectivity) and should enable the development of new classes of paper-based diagnostic devices.

Fully-drawn pencil-on-paper sensors for electroanalysis of dopamine

In this paper, a facile and low-cost “pencil-drawing” method is exploited for writing electrodes on filter paper to fabricate an electrochemical paper-based analytical device. Unlike other techniques using inks or pastes, the three-electrode system can be prepared directly by drawing with nothing more than pencils and paper. The optimization using ferricyanide as the redox probe showed that the device fully drawn with 6B grade pencil possesses excellent electrochemical properties and sensor-to-sensor reproducibility in spite of its truly simple fabrication process. Experimental results demonstrated that the proposed device could be utilized for quantification of dopamine with high sensitivity and no interference from ascorbic acid and uric acid, which are normally present in biological fluids. Human urine and blood serum samples were analyzed using the proposed device as proof of concept. Such fully-drawn sensors are flexible, portable and disposable, represent a cheap and environmentally friendly approach that can be rapidly and easily fabricated without any specialized materials, affording great convenience for practical use in resource-limited or emergency situations.

Application of nanomaterials in electrochemical paper-based biomedical microfluidic devices

Microfluidic paper-based analytical devices (μPADs), because of their mechanical flexibility, low cost, easy scale-up in manufacturing, small sample volumes and user-friendly properties, show great application prospect in bio-medicine, environmental monitor, food industry, drug industry and so on. As one kind of μPADs, the electrochemical microfluidic paper-based analytical devices (EμPADs) combine the advantages of μPADs with the advantages of electrochemical detection and keep attracting more and more attention. Recently, to improve the performance of EμPADs, applying nanomaterials for the fabrication of biosensors has become the research hotspot. This review presents some of the most prominent applications of nanomaterials in EμPADs over the past five years and also describes the challenges and future perspectives related to the recent application and development of paper-based electrochemical biosensors.

Electrochemical detection in paper-based analytical devices using microwire electrodes

Microwire electrodes are presented as an alternative to screen-printed electrodes for detection in electrochemical paper-based analytical devices (ePADs). Compared to carbon ink electrodes, microwire electrodes offer lower resistance and a significant increase in current density relative to carbon ink electrodes. Various microwire compositions and diameters, including 30 μm Pt, 25 μm Au, 18 μm Pt with 8% W, and 15 μm Pt with 20% Ir, were tested and compared to theoretically predicted behavior. The measured current in static solution was below predicted levels for cylindrical microelectrodes but greater than levels predicted for hemi-cylindrical electrodes most likely as a result of the proximity of the electrode to the paper surface. Furthermore, the current response was indicative of semi-thin layer behavior, likely due to the confined solution volume in the paper. After electrode characterization, a device was developed for the non-enzymatic detection of glucose, fructose, and sucrose using a Cu electrode in alkaline solution. The limits of detection for glucose, fructose, and sucrose were 270 nM, 340 nM, and 430 nM, respectively, which are significantly below sugar concentrations found in sweetened beverages or glucose levels in serum.

Electrochemical paper-based microfluidic devices.

Self-pumping porous microfluidic devices have attracted significant interest because of their low cost and broad applicability in point-of-care and low resource settings. One limitation of many of the devices is sensitivity and selectivity for detection. Electrochemistry can provide a sensitive, selective detection method while still using low cost, portable instrumentation as typified by handheld glucometers. Here, the development of electrochemical paper-based analytical devices (ePADs) is reviewed. Given the importance of electrode geometry and composition, fabrication methods are reviewed first. This is followed by a review of example applications demonstrated for ePADs. Finally, major accomplishments and future directions are summarized.

Recent developments in electrochemical paper-based analytical devices

This mini-review discusses recent advances in electrochemical paper-based analytical devices with an emphasis on applications of traditional electrochemical methods.

Paper-based potentiometric ion sensing.

This paper describes the design and fabrication of ion-sensing electrochemical paper-based analytical devices (EPADs) in which a miniaturized paper reference electrode is integrated with a small ion-selective paper electrode (ISPE) for potentiometric measurements. Ion-sensing EPADs use printed wax barriers to define electrochemical sample and reference zones. Single-layer EPADs for sensing of chloride ions include wax-defined sample and reference zones that each incorporate a Ag/AgCl electrode. In EPADs developed for other electrolytes (potassium, sodium, and calcium ions), a PVC-based ion-selective membrane is added to separate the sample zone from a paper indicator electrode. After the addition of a small volume (less than 10 μL) of sample and reference solutions to different zones, ion-sensing EPADs exhibit a linear response, over 3 orders of magnitude, in ranges of electrolyte concentrations that are relevant to a variety of applications, with a slope close to the theoretical value (59.2/z mV). Ion-selective EPADs provide a portable, inexpensive, and disposable way of measuring concentrations of electrolyte ions in aqueous solutions.