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Microfluidic paper-based analytical devices (μPADs) have garnered significant interest as a promising analytical platform in the past decade. Compared with traditional microfluidics, μPADs present unique advantages, such as easy fabrication using established patterning methods, economical cost, ability to drive and manipulate flow without equipment, and capability of storing reagents for various applications. This Review aims to provide a comprehensive review of the field, highlighting fabrication methods available to date with their respective advantages and drawbacks, device designs and modifications to accommodate different assay needs, detection strategies, and the growing applications of μPADs. Finally, we discuss how the field needs to continue moving forward to realize its full potential.

Microfluidic magnetophoresis is a powerful technique that is used to separate and/or isolate cells of interest from complex matrices for analysis. However, mechanical pumps are required to drive flow, limiting portability and making translation to point-of-care (POC) settings difficult. Microfluidic paper-based analytical devices (μPADs) offer an alternative to traditional microfluidic devices that do not require external pumps to generate flow. However, μPADs are not typically used for particle analysis because most particles become trapped in the porous fiber network. Here we report the ability of newly developed fast-flow microfluidic paper-based analytical devices (ffPADs) to perform magnetophoresis. ffPADs use capillary action in a gap between stacked layers of paper and transparency sheets to drive flow at higher velocities than traditional μPADs. The multi-layer ffPADs allow particles and cells to move through the gap without being trapped in the paper layers. We first demonstrate that ffPADs enable magnetic particle separations in a μPAD with a neodymium permanent magnet and study key factors that affect performance. To demonstrate utility, E. coli was used as a model analyte and was isolated from human urine before detection with a fluorescently labeled antibody. A capture efficiency of 61.5% was then obtained of E. coli labeled magnetic beads in human urine. Future studies will look at the improvement of the capture efficiency and to make this assay completely off-chip without the need of a fluorescent label. The assay and device described here demonstrate the first example of magnetophoresis in a paper based, pump free microfluidic device.

The use of paper microfluidics to perform chemical measurements for various analytical applications has gained interest over the last decade. One of the growing applications of these platforms is for the qualitative and quantitative determination of drugs. The low cost and self-pumping ability of paper microfluidics are attractive for developing analytical tools capable of on-site drug screening. This review aims to present the unique features of microfluidic paper-based analytical devices (μPADs) that offer advantages to pharmaceutical analysis and evaluate the state-of-the-art technologies and applications of the platform for drug analysis in research and real-world settings. The current challenges and potential future directions of the field are also highlighted., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Microfluidic paper-based analytical devices (μPADs) allow user-friendly and portable chemical determinations, although they provide limited applicability due to insufficient sensitivity. Several approaches have been proposed to address poor sensitivity in μPADs, but they frequently require bulky equipment for power and/or read-outs. Universal serial buses (USB) are an attractive alternative to less portable power sources and are currently available in many common electronic devices. Here, USB-powered μPADs (USB μPADs) are proposed as a fusion of both technologies to improve performance without adding instrumental complexity. Two ITP USB μPADs were developed, both powered by a 5 V potential provided through standard USB ports. The first device was fabricated using the origami approach. Its operation was analyzed experimentally and numerically, yielding a two-order-of-magnitude sample focusing in 15 min. The second ITP USB μPAD is a novel design, which was numerically prototyped with the aim of handling larger sample volumes. The reservoirs were moved away from the ITP channel and capillary action was used to drive the sample and electrolytes to the separation zone, predicting 25-fold sample focusing in 10 min. USB μPADs are expected to be adopted by minimally-trained personnel in sensitive areas like resource-limited settings, the point-of-care and in emergencies., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Microfluidic paper-based analytical devices (μPADs) are simple but powerful analytical tools that are gaining significant recent attention due to their many advantages over more traditional monitoring tools. These include being inexpensive, portable, pump-free, and having the ability to store reagents. One major limitation of these devices is slow flow rates, which are controlled by capillary action in the hydrophilic pores of cellulosic paper. Recent investigations have advanced the flow rates in μPADs through the generation of a gap or channel between two closely spaced paper sheets. This multilayered format has opened up μPADs to new applications and detection schemes, where large gap sizes (>300 μm) provide at least 169× faster flow rates than single-layer μPADs, but do not conform to established mathematical models for fluid transport in porous materials, such as the classic Lucas-Washburn equation. In the present study, experimental investigations and analytical modeling are applied to elucidate the driving forces behind the rapid flow rates in these devices. We investigate a range of hypotheses for the systems fluid dynamics and establish a theoretical model to predict the flow rate in multilayered μPADs that takes into account viscous dissipation within the paper. Device orientation, sample addition method, and the gap height are found to be critical concerns when modeling the imbibition in multilayered devices.

This study describes the use of mass spectrometry imaging with matrix-assisted laser desorption/ionization (MALDI) and desorption electrospray ionization (DESI) to understand the color gradient generation commonly seen in microfluidic paper-based analytical devices (μPADs). The formation of color gradients significantly impacts assay sensitivity and reproducibility with μPADs but the mechanism for formation is poorly understood. The glucose enzymatic assay using potassium iodide (KI) as a chromogenic agent was selected to investigate the color gradient generated across a detection spot. Colorimetric measurements revealed that the relative standard deviation for the recorded pixel intensities ranged between 34 and 40%, compromising the analytical reliability. While a variety of hypotheses have been generated to explain this phenomenon, few studies have attempted to elucidate the mechanisms associated with its formation. Mass spectrometry imaging using MALDI and DESI was applied to understand the nonuniform color distribution on the detection zone. MALDI experiments were first explored to monitor the spatial distribution of the glucose oxidase and horseradish peroxidase mixture, before and after lateral flow assay with and without KI. MALDI(+)-TOF data revealed uniform enzyme distribution on the detection spots. On the other hand, after the complete assay DESI(-) measurements revealed a heterogeneous shape indicating the presence of iodide and triiodide ions at the zone edge. The reaction product (I 3 - ) is transported by lateral flow toward the zone edge, generating the color gradient. Mass spectrometry imaging has been used for the first time to prove that color gradient forms as result of the mobility small molecules and not the enzyme distribution on μPAD surface.

Coupling infrared (IR) spectroscopy to microfluidic devices provides a powerful tool for characterizing complex chemical and biochemical reactions. Examples of microfluidic devices coupled with infrared spectroscopy have been limited, however, largely due to the difficulties associated with fabricating systems in common infrared transparent materials like CaF 2 . Recent reports have shown that polydimethylsiloxane (PDMS) can be used as an IR transparent substrate when fabricated with thin layers. The use of soft lithography with PDMS expands the library of possible designs that can be achieved for IR measurements in microfluidics. In initial reports with thin PDMS, the target analytes were small molecules; however, IR spectroscopy offers a powerful tool to study protein structure and reactions. Here, a PDMS microfluidic device compatible with IR spectroscopy was fabricated by means of spin-coating of PDMS pre-polymer to obtain thin PDMS microfluidic features. The device was comprised of only PDMS and IR absorption of PDMS was significantly minimized due to the thickness (∼40 μm) of the PDMS layer. The use of thin PDMS allowed for measuring the amide I and II vibrational bands of proteins that have been difficult to measure in other microfluidic devices. To demonstrate the power of the system, the microfluidic device was successfully used to measure the enzyme kinetics as one class of important biochemical reactions with broad use in a variety of fields from medicine to biotechnology. As a model, the reaction of glucose oxidase with glucose was tracked by following the formation of gluconic acid. Michaelis-Menten kinetics from the device were compared with bulk solution measurements and found to be in good agreement., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Falsified and substandard antibiotics are a growing worldwide problem that leads to increased patient mortality and decreased trust in healthcare, and contributes to antimicrobial resistance. Monitoring falsified antibiotics is difficult because most falsified pharmaceuticals are most commonly found in developing countries, where detecting the active ingredient is difficult due to lack of access to complex instrumentation. Herein, we describe the development and optimization of a microfluidic paper-based analytical device (μPAD) to detect the active ingredient in the most falsified class of antibiotics, β-lactams. The assay is based on enzyme competition, making it the first demonstrated competitive enzyme assay reported in paper-based devices. The assay uses nitrocefin, a chromogenic substrate, to compete with β-lactam antibiotics in a reaction with β-lactamase. A yellow color indicates legitimate drugs, while a color change from yellow to red indicates falsified drugs. In addition to testing for the active ingredient, another section of the device was added to test the sample pH to further verify results and identify common falsified ingredients like aspirin or baking soda. Calibration curves for four different antibiotics, including cefazolin, have been generated making it the first paper-based device capable of detecting both cephalosporin and penicillin antibiotics. The μPAD has also been tested with common falsified ingredients and four antibiotics in tablet or injectable form, demonstrating its potential for in-field falsified antibiotic testing.

This Frontiers review analyzes the rapidly growing microfluidic strategies that have been employed in attempts to create physio relevant 'organ-on-chip' models using primary tissue removed from a body (human or animal). Tissue harvested immediately from an organism, and cultured under artificial conditions is referred to as ex vivo tissue. The use of primary (organotypic) tissue offers unique benefits over traditional cell culture experiments, and microfluidic technology can be used to further exploit these advantages. Defining the utility of particular models, determining necessary constituents for acceptable modeling of in vivo physiology, and describing the role of microfluidic systems in tissue modeling processes is paramount to the future of organotypic models ex vivo. Virtually all tissues within the body are characterized by a large diversity of cellular composition, morphology, and blood supply (e.g., nutrient needs including oxygen). Microfluidic technology can provide a means to help maintain tissue in more physiologically relevant environments, for tissue relevant time-frames (e.g., matching the natural rates of cell turnover), and at in vivo oxygen tensions that can be controlled within modern microfluidic culture systems. Models for ex vivo tissues continue to emerge and grow in efficacy as mimics of in vivo physiology. This review addresses developments in microfluidic devices for the study of tissues ex vivo that can serve as an important bridge to translational value.

An improved paper-based analytical device (PAD) using color screening to enhance device performance is described. Current detection methods for PADs relying on the distance-based signalling motif can be slow due to the assay time being limited by capillary flow rates that wick fluid through the detection zone. For traditional distance-based detection motifs, analysis can take up to 45 min for a channel length of 5 cm. By using a color screening method, quantification with a distance-based PAD can be achieved in minutes through a "dip-and-read" approach. A colorimetric indicator line deposited onto a paper substrate using inkjet-printing undergoes a concentration-dependent colorimetric response for a given analyte. This color intensity-based response has been converted to a distance-based signal by overlaying a color filter with a continuous color intensity gradient matching the color of the developed indicator line. As a proof-of-concept, Ni quantification in welding fume was performed as a model assay. The results of multiple independent user testing gave mean absolute percentage error and average relative standard deviations of 10.5% and 11.2% respectively, which were an improvement over analysis based on simple visual color comparison with a read guide (12.2%, 14.9%). In addition to the analytical performance comparison, an interference study and a shelf life investigation were performed to further demonstrate practical utility. The developed system demonstrates an alternative detection approach for distance-based PADs enabling fast (∼10 min), quantitative, and straightforward assays.

Microfluidic paper-based analytical devices (μPADs) are a versatile and inexpensive point-of-care (POC) technology, but their widespread adoption has been limited by slow flow rates and the inability to carry out complex in field analytical measurements. In the present work, we investigate multilayer μPADs as a means to generate enhanced flow rates within self-pumping paper devices. Through optical and electrochemical measurements, the fluid dynamics are investigated and compared to established flow theories within μPADs. We demonstrate a ∼145-fold increase in flow rate (velocity = 1.56 cm s -1 , volumetric flow rate = 1.65 mL min -1 , over 5.5 cm) through precise control of the channel height in a 2 layer paper device, as compared to archetypical 1 layer μPAD designs. These design considerations are then applied to a self-pumping sequential injection device format, known as a three-dimensional paper network (3DPN). These 3DPN devices are characterized through flow injection analysis of a ferrocene complex and anodic stripping detection of cadmium, exhibiting a 5× enhancement in signal compared to stationary measurements.

Boron doped diamond (BDD) electrodes have exemplary electrochemical properties; however, widespread use of high-quality BDD has previously been limited by material cost and availability. In the present article, we report the use of a BDD paste electrode (BDDPE) coupled with microfluidic paper-based analytical devices (μPADs) to create a low-cost, high-performance electrochemical sensor. The BDDPEs are easy to prepare from a mixture of BDD powder and mineral oil and can be easily stencil-printed into a variety of electrode geometries. We demonstrate the utility and applicability of BDDPEs through measurements of biological species (norepinephrine and serotonin) and heavy metals (Pb and Cd) using μPADs. Compared to traditional carbon paste electrodes (CPE), BDDPEs exhibit a wider potential window, lower capacitive current, and are able to circumvent the fouling of serotonin. These results demonstrate the capability of BDDPEs as point-of-care sensors when coupled with μPADs.

We report a simple, low-cost, one-step fabrication method for microfluidic paper-based analytical devices (μPAD) using only polystyrene and a patterned screen. The polystyrene solution applied through the screen penetrates through the paper, forming a three-dimensional hydrophobic barrier, defining a hydrophilic analysis zone. The optimal polystyrene concentration and paper types were first investigated. Adjusting polystyrene concentration allows for various types of paper to be used for successful device fabrication. Using an optimized polystyrene concentration with Whatman#4 filter paper, a linear relationship was found to exist between the design width and the printed width. The smallest hydrophilic channel and hydrophobic barrier that can be obtained are 670 ± 50 μm and 380 ± 40 μm, respectively. High device-to-device fabrication reproducibility was achieved yielding a relative standard deviation (%RSD) in the range of 1.12-2.54% (n = 64) of the measured diameter of the well-shaped fabricated test zones with a designed diameter of 5 and 7 mm. To demonstrate the significance of the fabricated μPAD, distance-based and well-based paper devices were constructed for the analysis of H2O2 and antioxidant activity, respectively. The analysis of H2O2 in real samples using distance-based measurement with CeO2 nanoparticles as the colorimetric agent produced the same results at 95% confidence level, as those obtained using KMnO4 titration. A proof-of-concept antioxidant activity determination based on the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay was also demonstrated. The results verify that the polymer screen-printing method can be used as an alternative method for μPAD fabrication.

Occupational exposure to Cr is concerning because of its myriad of health effects. Assessing chromium exposure is also cost and resource intensive because the analysis typically uses sophisticated instrumental techniques like inductively coupled plasma-mass spectrometry (ICP-MS). Here, we report a novel, simple, inexpensive microfluidic paper-based analytical device (μPAD) for measuring total Cr in airborne particulate matter. In the μPAD, tetravalent cerium (Ce(IV)) was used in a pretreatment zone to oxidize all soluble Cr to Cr(VI). After elution to the detection zone, Cr(VI) reacts with 1,5-diphenylcarbazide (1,5-DPC) forming 1,5-diphenylcarbazone (DPCO) and Cr(III). The resulting Cr(III) forms a distinct purple colored complex with the DPCO. As proof-of-principle, particulate matter (PM) collected on a sample filter was analyzed with the μPAD to quantify the mass of total Cr. A log-linear working range (0.23-3.75 μg; r(2)=0.998) between Cr and color intensity was obtained with a detection limit of 0.12 μg. For validation, a certified reference containing multiple competing metals was analyzed. Quantitative agreement was obtained between known Cr levels in the sample and the Cr measured using the μPAD., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Electrochemical paper-based analytical devices (ePADs) with integrated plasma isolation for determination of glucose from whole blood samples have been developed. A dumbbell shaped ePAD containing two blood separation zones (VF2 membranes) with a middle detection zone was fabricated using the wax dipping method. The dumbbell shaped device was designed to separate plasma while generating homogeneous flow to the middle detection zone of the ePAD. The proposed ePADs work with whole blood samples with 24-60% hematocrit without dilution, and the plasma was completely separated within 4 min. Glucose in isolated plasma separated was detected using glucose oxidase immobilized on the middle of the paper device. The hydrogen peroxide generated from the reaction between glucose and the enzyme pass through to a Prussian blue modified screen printed electrode (PB-SPEs). The currents measured using chronoamperometry at the optimal detection potential for H2O2 (-0.1 V versus Ag/AgCl reference electrode) were proportional to glucose concentrations in the whole blood. The linear range for glucose assay was in the range 0-33.1 mM (r(2)=0.987). The coefficients of variation (CVs) of currents were 6.5%, 9.0% and 8.0% when assay whole blood sample containing glucose concentration at 3.4, 6.3, and 15.6mM, respectively. Because each sample displayed intra-individual variation of electrochemical signal, glucose assay in whole blood samples were measured using the standard addition method. Results demonstrate that the ePAD glucose assay was not significantly different from the spectrophotometric method (p=0.376, paired sample t-test, n=10)., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Paper-based analytical devices (PADs) represent a growing class of elegant, yet inexpensive chemical sensor technologies designed for point-of-use applications. Most PADs, however, still utilize some form of instrumentation such as a camera for quantitative detection. We describe here a simple technique to render PAD measurements more quantitative and straightforward using the distance of colour development as a detection motif. The so-called distance-based detection enables PAD chemistries that are more portable and less resource intensive compared to classical approaches that rely on the use of peripheral equipment for quantitative measurement. We demonstrate the utility and broad applicability of this technique with measurements of glucose, nickel, and glutathione using three different detection chemistries: enzymatic reactions, metal complexation, and nanoparticle aggregation, respectively. The results show excellent quantitative agreement with certified standards in complex sample matrices. This work provides the first demonstration of distance-based PAD detection with broad application as a class of new, inexpensive sensor technologies designed for point-of-use applications.

Chemical gradients drive a diverse set of biological processes ranging from nerve transduction to ovulation. At present, the most common method for quantifying chemical gradients is microscopy. Here, a new concept for probing spatial and temporal chemical gradients is reported that uses a multi-layer microfluidic device to measure analyte concentration as a function of lateral position in a microfluidic channel using electrochemistry in a format that is readily adaptable to multi-analyte sensing.

Human exposure to particulate matter (PM) air pollution has been linked with respiratory, cardiovascular, and neurodegenerative diseases, in addition to various cancers. Consistent among all of these associations is the hypothesis that PM induces inflammation and oxidative stress in the affected tissue. Consequently, a variety of assays have been developed to quantify the oxidative activity of PM as a means to characterize its ability to induced oxidative stress. The vast majority of these assays rely on high-volume, fixed-location sampling methods due to limitations in assay sensitivity and detection limit. As a result, our understanding of how personal exposure contributes to the intake of oxidative air pollution is limited. To further this understanding, we present a microfluidic paper-based analytical device (μPAD) for measuring PM oxidative activity on filters collected by personal sampling. The μPAD is inexpensive to fabricate and provides fast and sensitive analysis of aerosol oxidative activity. The oxidative activity measurement is based on the dithiothreitol assay (DTT assay), uses colorimetric detection, and can be completed in the field within 30 min following sample collection. The μPAD assay was validated against the traditional DTT assay using 13 extracted aerosol samples including urban aerosols, biomass burning PM, cigarette smoke, and incense smoke. The results showed no significant differences in DTT consumption rate measured by the two methods. To demonstrate the utility of the approach, personal samples were collected to estimate human exposures to PM from indoor air, outdoor air on a clean day, and outdoor air on a wildfire-impacted day in Fort Collins, CO. Filter samples collected on the wildfire day gave the highest oxidative activity on a mass normalized basis, whereas typical ambient background air showed the lowest oxidative activity.

A microfluidic paper-based analytical device (μPAD) for the separation of blood plasma from whole blood is described. The device can separate plasma from whole blood and quantify plasma proteins in a single step. The μPAD was fabricated using the wax dipping method, and the final device was composed of a blood separation membrane combined with patterned Whatman No.1 paper. Blood separation membranes, LF1, MF1, VF1 and VF2 were tested for blood separation on the μPAD. The LF1 membrane was found to be the most suitable for blood separations when fabricating the μPAD by wax dipping. For blood separation, the blood cells (both red and white) were trapped on blood separation membrane allowing pure plasma to flow to the detection zone by capillary force. The LF1-μPAD was shown to be functional with human whole blood of 24-55% hematocrit without dilution, and effectively separated blood cells from plasma within 2 min when blood volumes of between 15-22 μL were added to the device. Microscopy was used to confirm that the device isolated plasma with high purity with no blood cells or cell hemolysis in the detection zone. The efficiency of blood separation on the μPAD was studied by plasma protein detection using the bromocresol green (BCG) colorimetric assay. The results revealed that protein detection on the μPAD was not significantly different from the conventional method (p > 0.05, pair t-test). The colorimetric measurement reproducibility on the μPAD was 2.62% (n = 10) and 5.84% (n = 30) for within-day and between day precision, respectively. Our proposed blood separation on μPAD has the potential for reducing turnaround time, sample volume, sample preparation and detection processes for clinical diagnosis and point-of care testing.

A microfluidic paper-based analytical device (μPAD) fabricated by wax printing was designed to assess occupational exposure to metal-containing aerosols. This method employs rapid digestion of particulate metals using microliters of acid added directly to a punch taken from an air sampling filter. Punches were then placed on a μPAD, and digested metals were transported to detection reservoirs upon addition of water. These reservoirs contained reagents for colorimetric detection of Fe, Cu, and Ni. Dried buffer components were used to set the optimal pH in each detection reservoir, while precomplexation agents were deposited in the channels between the sample and detection zones to minimize interferences from competing metals. Metal concentrations were quantified from color intensity images using a scanner in conjunction with image processing software. Reproducible, log-linear calibration curves were generated for each metal, with method detection limits ranging from 1.0 to 1.5 μg for each metal (i.e., total mass present on the μPAD). Finally, a standard incineration ash sample was aerosolized, collected on filters, and analyzed for the three metals of interest. Analysis of this collected aerosol sample using a μPAD showed good correlation with known amounts of the metals present in the sample. This technology can provide rapid assessment of particulate metal concentrations at or below current regulatory limits and at dramatically reduced cost.

The spatial and temporal distributions of an extensive number of diffusible molecules drive a variety of complex functions. These molecular distributions often possess length scales on the order of a millimeter or less; therefore, microfluidic devices have become a powerful tool to study the effects of these molecular distributions in both chemical and biological systems. Although there exist a number of studies utilizing microdevices for the creation of molecular gradients, there are few, if any, studies focusing on the measurement of spatial and temporal distributions of molecular species created within the study system itself. Here we present a microfluidic device capable of sampling multiple chemical messengers in a spatiotemporally resolved manner. This device operates through spatial segregation of nanoliter-sized volumes of liquid from a primary sample reservoir into a series of analysis microchannels, where fluid pumping is accomplished via a system of passive microfluidic pumps. Subsequent chemical analysis within each microchannel, achieved via optical or bioanalytical methods, yields quantitative data on the spatial and temporal information for any analytes of interest existing within the sample reservoir. These techniques provide a simple, cost-effective route to measure the spatiotemporal distributions of molecular analytes, where the system can be tailored to study both chemical and biological systems., (© 2011 American Chemical Society)

Wax screen-printing as a low-cost, simple, and rapid method for fabricating paper-based microfluidic devices (µPADs) is reported here. Solid wax was rubbed through a screen onto paper filters. The printed wax was then melted into the paper to form hydrophobic barriers using only a hot plate. We first studied the relationship between the width of a hydrophobic barrier and the width of the original design line. We also optimized the heating temperature and time and determined the resolution of structures fabricated using this technique. The minimum width of hydrophilic channel and hydrophobic barrier is 650 and 1300 µm, respectively. Next, our fabrication method was compared to a photolithographic method using the reaction between bicinchoninic acid (BCA) and Cu(1+) to demonstrate differences in background reactivity. Photolithographically defined channels exhibited a high background while wax printed channels showed a very low background. Finally, the utility of wax screen-printing was demonstrated for the simultaneous determination of glucose and total iron in control human serum samples using an electrochemical method with glucose oxidase and a colorimetric method with 1,10-phenanthroline. This study demonstrates that wax screen-printing is an easy-to-use and inexpensive alternative fabrication method for µPAD, which will be especially useful in developing countries.

We report here the use of multiple indicators for a single analyte for paper-based microfluidic devices (microPAD) in an effort to improve the ability to visually discriminate between analyte concentrations. In existing microPADs, a single dye system is used for the measurement of a single analyte. In our approach, devices are designed to simultaneously quantify analytes using multiple indicators for each analyte improving the accuracy of the assay. The use of multiple indicators for a single analyte allows for different indicator colors to be generated at different analyte concentration ranges as well as increasing the ability to better visually discriminate colors. The principle of our devices is based on the oxidation of indicators by hydrogen peroxide produced by oxidase enzymes specific for each analyte. Each indicator reacts at different peroxide concentrations and therefore analyte concentrations, giving an extended range of operation. To demonstrate the utility of our approach, the mixture of 4-aminoantipyrine and 3,5-dichloro-2-hydroxy-benzenesulfonic acid, o-dianisidine dihydrochloride, potassium iodide, acid black, and acid yellow were chosen as the indicators for simultaneous semi-quantitative measurement of glucose, lactate, and uric acid on a microPAD. Our approach was successfully applied to quantify glucose (0.5-20 mM), lactate (1-25 mM), and uric acid (0.1-7 mM) in clinically relevant ranges. The determination of glucose, lactate, and uric acid in control serum and urine samples was also performed to demonstrate the applicability of this device for biological sample analysis. Finally results for the multi-indicator and single indicator system were compared using untrained readers to demonstrate the improvements in accuracy achieved with the new system., (2010 Elsevier B.V. All rights reserved.)

We report the first demonstration of electrochemical detection for paper-based microfluidic devices. Photolithography was used to make microfluidic channels on filter paper, and screen-printing technology was used to fabricate electrodes on the paper-based microfluidic devices. Screen-printed electrodes on paper were characterized using cyclic voltammetry to demonstrate the basic electrochemical performance of the system. The utility of our devices was then demonstrated with the determination of glucose, lactate, and uric acid in biological samples using oxidase enzyme (glucose oxidase, lactate oxidase, and uricase, respectively) reactions. Oxidase enzyme reactions produce H2O2 while decomposing their respective substrates, and therefore a single electrode type is needed for detection of multiple species. Selectivity of the working electrode for H2O2 was improved using Prussian Blue as a redox mediator. The determination of glucose, lactate, and uric acid in control serum samples was performed using chronoamperometry at the optimal detection potential for H2O2 (0 V versus the on-chip Ag/AgCl reference electrode). Levels of glucose and lactate in control serum samples measured using the paper devices were 4.9 +/- 0.6 and 1.2 +/- 0.2 mM (level I control sample), and 16.3 +/- 0.7 and 3.2 +/- 0.3 mM (level II control sample), respectively, and were within error of the values measured using traditional tests. This study shows the successful integration of paper-based microfluidics and electrochemical detection as an easy-to-use, inexpensive, and portable alternative for point of care monitoring.

The development of a simple method to measure reaction rates using magnetic microparticles in a microfluidic device is explored.

Microfluidic devices have gained significant scientific interest due to the potential to develop portable, inexpensive analytical tools capable of quick analyses with low sample consumption. These qualities make microfluidic devices attractive for point-of-use measurements where traditional techniques have limited functionality. Many samples of interest in biological and environmental analysis, however, contain insoluble particles that can block microchannels, and manual filtration prior to analysis is not desirable for point-of-use applications. Similarly, some situations involve limited control of the sample volume, potentially causing unwanted hydrodynamic flow due to differential fluid heads. Here, we present the successful inclusion of track-etched polycarbonate membrane filters into the reservoirs of poly(dimethylsiloxane) capillary electrophoresis microchips. The membranes were shown to filter insoluble particles with selectivity based on the membrane pore diameter. Electrophoretic separations with membrane-containing microchips were performed on cations, anions, and amino acids and monitored using conductivity and fluorescence detection. The dependence of peak areas on head pressure in gated injection was shown to be reduced by up to 92%. Results indicate that separation performance is not hindered by the addition of membranes. Incorporating membranes into the reservoirs of microfluidic devices will allow for improved analysis of complex solutions and samples with poorly controlled volume.