Microfluidic Paper-based Analytical Devices Research Articles

Microfluidic paper-based analytical device with a preconcentration system for the measurement of anionic surfactants using an optode detector.

The paper discusses the development of a low-cost, simple, and sensitive on-site measurement system for anionic surfactants (AS), specifically sodium alkylbenzene sulfonate (LAS), using a microfluidic paper-based analytical device (μPAD) with an optode as a detector. The need arises due to the discharge of AS-containing wastewater into natural environments, posing risks to aquatic organisms. Traditional methods for AS measurement have drawbacks like the use of harmful solvents, time-consuming procedures, and the need for expensive equipment, prompting the exploration of alternative approaches. The μPAD incorporates a sample solution preconcentration system using filter paper modified with chitosan. The optode, a chemical sensor detecting analytes optically, is employed for AS detection. When a large volume of AS is added to a positively charged modified filter paper with chitosan, the AS is adsorbed and concentrated on the filter paper. The concentrated AS is eluted with a small volume of alkaline solution, and the eluted AS is detected by the optode in the μPAD. The μPAD with preconcentration provides improved sensitivity and a broader range of detection compared to the method without preconcentration. In the present μPAD method, linear alkylbenzenesulfonate (LAS) in the concentration range of 0.1 to 10 μmol dm-3 was measured. The developed μPAD offers advantages such as portability, cost-effectiveness, and negligible interference from coexisting substances in environmental water samples. The μPAD method was applied for the determination of LAS in tap water and river water.

A paper-based point-of-care device for the detection of cysteine using gold nanoparticles from whole blood.

We present a colorimetric probe based on polyvinylpyrrolidone-capped gold nanoparticles (PVP-AuNPs) that is sensitive and selective for cysteine (Cys). A microfluidic paper-based analytical device (μ-PAD) with embedded dried PVP-AuNPs at the polyethersulfone (PES) paper surface is used for Cys detection. When thiol molecules attach to PVP-AuNPs in the presence of Cys, they clump together, and this causes the solution's color to shift from red to blue within 5 minutes. The device is capable of detecting Cys levels between 1.0 μM and 50.0 μM with a limit of detection (LOD) of 0.2 μM under optimized conditions. The stability of the μ-PAD was tested for 100 days, demonstrating re-dispersibility to detect Cys levels in blood. Dried PVP-AuNP-μPADs were integrated with blood plasma separation modules for point-of-care (POC) Cys detection. Consequently, the device shows potential as a self-sustaining, quantification platform with a recovery percentage ranging from 98.44 to 111.9 in clinical samples.

Flow control by circular cavities in lateral flow porous membranes.

This research explores the flow penetration in porous media by virtue of capillary action and geometric control of the liquid imbibition rate in microfluidic paper-based analytical devices (μPADs) having applications in food quality management, medical diagnostics, and environmental monitoring. We examine changes in flow resistance and membrane geometry, aiming to understand factors influencing capillary penetration rates for various practical applications. We conducted experiments and simulations using lateral porous membranes and altered the flow resistance by changing the liquids or the paper channel geometry by adding cavities. From experiments, it was revealed that by creating a circular cavity in the paper channel, the penetration rate was sufficiently altered. Moreover, increasing the cavity size and type of liquid (w.r.t. viscosity) also caused a decrease in the flow rate. Imbibition rates were also influenced by the position of the cavities in the paper channel. The maximum delay for water was almost 2 times with a 16 mm circular cavity located at 3 cm from strip bottom edge. Overall, we attained a maximum delay in the case of castor oil which was almost 85 times slower than water and 3.7 times slower than olive oil. A good agreement was observed with CFD analysis. We believe that this research would help in developing advance techniques to enhance the flow control strategies in μPADs and indicators.

A novel ratiometric design of microfluidic paper-based analytical device for the simultaneous detection of Cu2+ and Fe3+ in drinking water using a fluorescent MOF@tetracycline nanocomposite.

The regular and on-site monitoring of ions in drinking water is essential for safeguarding public health, ensuring high water quality, and preserving the ecological balance of aquatic ecosystems. Thus, developing a portable analytical device for the rapid, cost-effective, and visual on-site detection of multiple environmental pollutants is notably significant. In the present work, a novel ratiometric microfluidic paper-based analytical device (μPAD) was designed and developed for the simultaneous detection of Fe3+ and Cu2+ ions in water samples taking advantages from built-in masking zone. The μPAD was functionalized with a greenish-yellow fluorescent Zn-based metal-organic framework@tetracycline (FMOF-5@TC) nanocomposite, and the ratiometric design was based on the change in emission color from greenish yellow (FMOF-5@TC) to blue (FMOF-5). The μPAD consisted of one sample zone linked to two detection zones via two channels: the first channel was for the detection of both ions, while the second was intended for detecting only Cu2+ ions and comprised a built-in masking zone to remove Fe3+ ions prior to reaching the detection zone. The corresponding color changes were recorded with the aid of a smartphone and RGB calculations. The linear ranges were 0.1-80 μM for Cu2+ and 0.2-160 μM for Fe3+, with limits of detection of 0.027 and 0.019 μM, respectively. The simple μPAD design enabled the simultaneous detection of Cu2+ and Fe3+ ions in drinking water samples with excellent accuracy and precision, with spike recoveries of 81.28-96.36% and 83.01-102.33% for Cu2+ and Fe3+, respectively.

Detection of azadirachtin from neem kernels using a paper-based sandwich sensor.

In this work, a microfluidic paper-based analytical device (μPAD) was developed to detect the biopesticide azadirachtin (Aza) through a colorimetric assay. High precision estimation of Aza is classically carried out using high performance liquid chromatography (HPLC), which requires highly skilled personnel. Acidified vanillin is a commonly used colorimetric indicator in thin layer chromatography for detection of various phytochemicals. However, the assay involves concentrated acid, which limits the choice of paper substrates for paper-based sensors and raises safety concerns. In this work, we show how the assay can be extended from the liquid phase to a paper substrate. Glass microfiber (GMF) filter paper was found to be suitable paper as it was acid resistant; besides, its hydrophilicity enabled smooth flow of reagents. A microfluidic paper-based sensor (μPAD) was developed by sandwiching 5 mm sized GMF dots between two parafilm sheets. We demonstrate the use of colorimetric assay on the μPAD for on-site detection of Aza in neem kernels. The magenta color developed upon the reaction of acidified vanillin with Aza was captured using a smart-phone and analysed using RGB levels in the image. Calibration was established using neem kernel extract of known concentration. Linearity was seen in the concentration range of 5 to 25 mg L-1 Aza. A limit of detection of 2.3 mg L-1 was obtained using this method. The colorimetric assay showed a relative recovery of >85% when compared with the values obtained from HPLC. The stability of the reagents on the GMF sensor was investigated to understand the storage conditions and shelf life of the reagents and sensor. The present work demonstrates the development of a portable sensor for on-site detection of phytochemicals that can be an integral part of the agricultural supply chain.

Ion imprinted polymers integrated into a multi-functional microfluidic paper-based analytical device for trace cadmium detection in water.

A novel multi-functional microfluidic paper-based analytical device (μPAD) integrated with ion imprinted polymers (IIPs) was proposed for specific, portable and low-cost detection of cadmium (Cd(II)) in water. The IIP was grafted on paper and integrated into the μPAD for separation of Cd(II) through multi-layer design. The paper-based screen printed carbon electrode (pSPCE) modified with reduced graphene oxide was fabricated and combined with the μPAD for electrochemical sensing of the separated Cd(II). Reduced graphene oxide (rGO) was prepared via electroreduction on the working electrode surface of the pSPCE (rGO/pSPCE), which provided a sensitization effect with an improved signal for Cd(II) detection. The μPAD developed with the integrated IIP and combined with rGO/pSPCE is able to detect Cd(II) with a linear range from 1 ng ml-1 to 100 ng ml-1 and a detection limit of 0.05 ng ml-1. The accuracy of this μPAD was evaluated with spiked real water samples and compared with that of the inductively coupled plasma mass spectrometry (ICP-MS) method, from which the recovery values ranged from 96.5% to 114.2% with RSDs <10% between the two methods. This μPAD demonstrated its advantages of low-cost, portability, and suitability for highly sensitive detection of Cd(II), making it a valuable tool for on-site analysis.

In situ biomimetic mineralization of a paper microfluidic device as a luminescent sensor for nitrite determination

Nucleation and growth of terbium-doped amorphous calcium phosphate (Tb-ACP) on cellulose fibers of a microfluidic paper-based analytical device (μPAD) towards an environmentally friendly design of a sensor for water pollutant.

One-drop chemosensing of dapoxetine hydrochloride using opto-analysis by multi-channel μPAD decorated silver nanoparticles: introducing a paper-based microfluidic portable device/sensor toward naked-eye pharmaceutical analysis by lab-on-paper technology.

Dapoxetine (DPX) belongs to the selective serotonin reuptake inhibitor (SSRI) class and functions by blocking the serotonin transporter and increasing serotonin activity, thereby delaying ejaculation. Therefore, monitoring of the concentration of DPX in human biofluids is important for clinicians. In this study, application of silver nanoparticles with the morphology of prisms (AgNPrs) for the sensitive measurement of DPX using colorimetric chemosensing and the spectrophotometric method was investigated. Also, DPX was determined in real samples using the spectrophotometry method. Based on the obtained results, all of the detection process in colorimetric assay is related to morphological reform of AgNPrs after it's specific electrostatic and covalent interaction with DPX as analyte. The UV-vis results indicate that the proposed AgNPrs-based chemosensing system has a wide range of linearity (0.01 μM to 1 mM) with a low limit of quantification of 0.01 μM in human urine samples, which is suitable for clinical analysis of this drug in human urine samples. It is important to point out that, this chemosensing strategy showed inappropriate analytical results for the detection of DPX in human urine samples which is a novelty of this platform. Finally, the optimized microfluidic paper-based analytical device (μPAD) was integrated with the colorimetric analysis of DPX to provide a time/color system for estimating analyte concentration by a portable substrate toward in situ and on-site biomedical analysis. Interestingly, the analytical validation tests showed appropriate results with great stability, which may facilitate commercialization of the engineered substrate. For the first time, in order to provide a simple and portable colorimetric/spectrophotometric recognition system to sensitive determination of DPX, an optimized pump-less microfluidic paper-based colorimetric device (μPCD) was introduced and validated for the real-time biomedical analysis of this analyte. According to the obtained results, this alternative approach is suitable for therapeutic drug monitoring (TDM) and biomedical analysis by miniaturized and cost-beneficial devices.

Distance-based detection of paracetamol in microfluidic paper-based analytical devices for forensic application.

Whisky adulteration is a prevalent practice driven by the high cost of these beverages. Counterfeiters commonly dilute whisky with less expensive alcoholic beverages, water, food additives, drugs or pharmaceuticals. Paracetamol (PAR), an analgesic drug that mitigates hangovers and headaches, is commonly used to adulterate whisky. Currently, the primary method for quantifying PAR levels is high-performance liquid chromatography, but this technique is both time consuming and usually generates more residues. In this context, the utilization of miniaturized and portable analytical devices becomes imperative for conducting point-of-care/need analyses. These devices offer several advantages, including portability, user-friendliness, low cost, and minimal material wastage. This study proposes the selective distance-based PAR quantification on whisky samples using a paper-based microfluidic analytical device (μPAD). Colorimetric detection on paper-based platforms offers great benefits such as affordability, portability, and the ability to detect PAR without complicated instrumentation. The optimal detection conditions were achieved by introducing 5 μL of a mixture containing 7.5 mmol L-1 of Fe(III) and K3[Fe(CN)6] into the detection zone, along with 12 μL of whisky samples into the sample zone. The method exhibited linear behavior within the concentration range from 15 to 120 mg L-1, with a determination coefficient of 0.998. PAR was quantified in adulterated samples. The results obtained with the paper-based devices were compared with a referenced method, and no significant differences were observed at a confidence level of 95%. The μPAD allowed to determine ca. 1 drop of pharmaceutical medicine PAR of 200 mg mL-1 in 1 L of solution, demonstrating excellent sensitivity. This method offers cost-effective and rapid analysis, reducing the consumption of samples, reagents, and wastes. Consequently, it could be considered a viable and portable alternative for analyzing beverages at criminal scenes, customs, and police operations, thereby enhancing the field of forensics.

Acoustofluidics-Assisted Multifunctional Paper-Based Analytical Devices.

Microfluidic paper-based analytical devices (μPADs) feature an economic and sensitive nature, while acoustofluidics displays contactless and versatile virtue, and both of them gained tremendous interest in the past decades. Integrating μPADs with acoustofluidic techniques provides great potential to overcome the inherent shortcomings and make appealing achievements. Here, we present acoustofluidics-assisted multifunctional paper-based analytical devices that leverage bulk acoustic waves to realize multiple applications on paper substrates, including uniform colorimetric detection, microparticle/cell enrichment, fluorescence amplification, homogeneous mixing, and nanomaterial synthesis. The glucose detection in the range of 5-15 mM was conducted to perform uniform colorimetric detection. Various types (brass powder, copper powder, diamond powder, and yeast cells) and sizes (5-200 μm) of solid particles and biological cells can be enriched on paper in a few seconds or minutes; thus, fluorescence amplification by 3 times was realized with the enrichment. The high-throughput and homogeneous mixing of two fluids can be achieved, and based on the mixing, nanomaterials (ZnO nanosheets) were synthesized on paper. We analyzed the underlying mechanisms of these applications in the devices, which are attributed to Faraday waves and Chladni patterns. With their simple fabrication and prominent effectiveness, the devices open up new possibilities for paper-based microfluidic devices.

Electrochemical microfluidic paper-based analytical device for label-free detection of SARS-CoV-2 antigen by LDH redox probe

In the offered study, the proposed sensor combined the innovative disposable microfluidic paper-based immunosensor and cost-effective plant-based anti-SARS-CoV-2 monoclonal antibody CR3022, expressed in Nicotiana benthamiana. An electrochemical microfluidic paper-based analytical device (µPAD) was introduced for quantitative detection of the SARS-CoV-2 antigen employing electrocatalytic activity of the Ni Fe layered double hydroxide (NiFe LDH). Its electrocatalytic activity asthe platform of proposed the analytical device enhanced the signal output via redox-recycling of the species the ferri/ferrocyanide [Fe(CN)6]3-/4- in solution and is applied to µPAD response signal. Layered NiFe double hydroxide (LDH) nanosheets with a suitable redox potential were prepared by incorporating Fe into the Ni-based LDH. With the help of Fe, the charge-transfer kinetics for the reduction of Ni3+ to Ni2+ was improved and the formation of unwanted Ni components with higher oxidation state was suppressed. The incorporated Fe as the electron transfer mediator enhanced the process of Ni(OH)2/NiOOH redox cycle. Under the optimum conditions, the dependency of impedance signal and SARS-CoV-2 concentration showed a linear region from 10 fg/mL-5 ng/mL with a detection limit of 0.23 fg/mL. This Paper-based immunosensor device also showed long stability and good reproducibility, which can be used for the quantitative assay of the SARS-CoV-2. The results of nasal samples analysis revealed the applicability of the resulting biosensor.

Improving pesticide residue detection: Immobilized enzyme microreactor embedded in microfluidic paper-based analytical devices

Orientationally immobilized enzyme microreactors (OIMERs), embedded in microfluidic paper-based analytical devices (μPADs) were developed for improved detection of pesticide residues in food. Acetylcholinesterase (AChE) was orientationally immobilized on the reusable Part I of the μPADs, using the specific affinity binding of concanavalin A (Con A) to a glycosyl group on AChE. Using the disposable Part II, facile colorimetric quantification was performed with a smartphone and software, or qualitative detection by a naked-eye visual test. The AChE immobilized in OIMERs not only had improved activity and stability, but also high sensitivity, with a limit of detection as low as (0.007 ± 0.003) μg/mL. The method was used to detect pesticides residues in real vegetable samples; the recovery (88.6–102.7%) showed high reliability for pesticide residues detection in foods. A molecular docking study and an enzyme kinetic analysis were conducted to characterize the mechanism of action of the OIMERs.

Development of Microfluidic Paper-Based Analytical Devices (µPADs) for Determination of Cd2+, Pb2+, and Cu2+ Ions in Mineral Water

Microfluidic paper-based analytical devices (µPADs) have been successfully developed using dual detection: electrochemical and colorimetric systems. The µPADs have the potential to be used as Pb(II), Cd(II), and Cu(II) sensors to test the quality of water. The fabrication process uses hot embossing and screen-printing methods. The working electrode in the electrochemical zone was enhanced by the bismuth metal deposition process, while in the colorimetric zone, the gold nanoparticles modified with thioctic acid and dansylhydrazine (TA-Au-DNS) were used as a colorimetric sensor to detect Cu. The basic material of µPADs was characterized using a Fourier-transform infrared (FTIR) and a contact angle meter (CAM). In the electrochemical zone, the signals of square wave anodic stripping voltammetry (SWASV) resulted in good detection of Pb(II) and Cd(II) (from 0 to 100 ppb) with a limit of detection of 1.588 and 1.42 ppb, respectively. In the colorimetric zone, the performance of TA-Au-DNS for detecting Cu metal was obtained from readings through the red-green-blue (RGB) sensor as a miniature of µPADs reader. The LOD, LOQ, and average Vx0 (linearity values) in the detection of Cu(II) (from 58 to 100 ppb) are 8.51 ppb, 28.36 ppb, and 0.41%, respectively.

µPAD-based colorimetric nanogold aptasensor for CRP and IL-6 detection as sepsis biomarkers

Point-of-care testing is defined as medical diagnostic testing at or near the time and place of a patient. The integration of microfluidics with functionalized nanomaterials provides a rapid and sensitive detection method for biomarkers. Due to their optical characteristics, gold nanoparticles (GNPs) and aptamers can be used in a colorimetric technique. In this research, accurate and quick aptasensors were created using aptamers for two sepsis biomarkers, interleukin-6 (IL-6) and C-reactive protein (CRP). Aptamers were combined with GNPs to create aptasensors, which were then used to analyze various biomarker concentrations after aptamer-protein docking on the online HDOCK server to verify aptamer-protein interactions. To improve the results' stability and ease of use during the reaction time, microfluidic paper-based analytical devices (µPAD) was employed. The linear diagnostic ranges in this method were 50–1000 mg L−1 and 1–25 ng/L, and limit of detection values were 1.9 mg L−1 and 0.07 ng/L for CRP and IL-6, respectively; These values decreased when µPADs were used and became more suitable to diagnosis of sepsis. Two aptasensors were highly stable, reproducible, accurate, and recoverable. Our result suggests that colorimetric GNPs can be used to detect a specific analyte through color changes visible to the naked eye for monitoring purposes or as simple optical detectors for quantitative measurements.

Construction of fluorescent sensor array and three-dimensional microfluidic paper based analytical device for specific identification and visual determination of antibiotics in food.

For antibiotics misuse since the global outbreak of COVID 19, a novel strategy for discriminating and detecting antibiotics is proposed based on the graphene quantum dots with multi-doped heteroatoms including F, N and P (M-GQDs), which exhibit blue emission (419.0nm) under the excitation of 336.0nm. Specifically, the fluorescence of M-GQDs is quenched by tetracyclines (TCs) owing to inner filter effect (IFE) and enhanced by alkane-modified fluoroquinolones (AFQs), which is attributed to restricted conformational rotation based on π-π stacking, hydrogen-bonding and electrostatic interactions. Meanwhile, the electron-accepting property of oxazine ring in oxazine-modified fluoroquinolones (OFQs) increases emission peak at 498.0nm and decreases emission peak at 419.0nm as the color changes from blue to cyan. Moreover, a cascade system integrated with 3D microfluidic paper-based analytical device (3D-μPAD) is applied successfully for visually distinguishing three antibiotics, which shows great potential and versatility of M-GQDs for food safety monitoring.

A simple and low-cost paper-based device for simultaneous determination of hematocrit and hemoglobin levels in point-of-care settings

Hematocrit and hemoglobin levels are common blood tests used to assess the overall health status of patients as well as the oxygen-carrying capacity of the blood. The gold-standard technologies to determine the hematocrit and hemoglobin levels are precise and fast, but they are not pragmatic in extreme point-of-care settings due to the prohibitive cost, process complexity, and sophisticated facilities. Therefore, here, we report the development of a microfluidic paper-based analytical device to determine hematocrit and hemoglobin levels simultaneously. This device exploits the area of the stain formed by a spreading drop of 20 μl of whole blood on Whatman filter paper immobilized with sodium chloride and ethylenediaminetetraacetic acid for hematocrit level, whereas for hemoglobin concentration, it utilizes the gray color intensity of a 20 μl droplet of a mixture of blood and de-ionized water. The performance of the device is verified by comparing with gold-standard results of the automated hematology analyzer, showing a high degree of correlation (R2) of 0.9651 and 0.9701 for hematocrit and hemoglobin levels, respectively. The bias and standard deviation of the differences between the two measurements for hematocrit determination are 0.002 and 1.005, respectively, while the bias and standard deviation of differences between the two methods for hemoglobin determination are 0.065 and 0.398, respectively. This device is likely to provide a simple, fast, disposable, and inexpensive tool to determine the hematocrit and hemoglobin levels in resource-constraint settings.

Smartphone-integrated colorimetric and microfluidic paper-based analytical devices for the trace-level detection of permethrin.

Permethrin is a pyrethroid pesticide and insect repellent that prevents mosquito-borne infections like dengue and malaria in tropical areas. This work describes a new colorimetric sensor based on metronidazole-stabilized silver nanoparticles (MTZ-AgNPs) for the first rapid, sensitive, and selective permethrin detection. The MTZ-AgNPs-based colorimetric sensor has a limit of detection (LOD) of 0.0104µM and a limit of quantification (LOQ) of 0.0348µM, respectively. The sensor is further integrated with smartphone and microfluidic fabrication of paper-based analytical devices (µPADs) for real-time and on-site detection of permethrin. Under optimal settings, no potential environmental contaminants interfere with permethrin detection, confirming its high selectivity. Finally, the practical applicability of sensors is confirmed in real tomato and apple extract samples. The US environmental protection agency's recommended UPLC method validated the detection efficiency of the proposed colorimetric sensor. The % recoveries from UPLC and MTZ-AgNPs suggest that the present sensor can quantitatively analyze permethrin in real samples.

Dual-spot ratiometric microfluidic paper-based analytical device for accuracy and precision improvement

Instrumental and environmental fluctuations are common sources of error in smartphone-based optical detection, significantly affecting the accuracy of analytical measurements. In this regard, spotting the sample and reference simultaneously and in close proximity compensates for the fluctuations. This “dual-spot” design is similar to the double-beam technique used in spectrophotometry, which reduces fluctuations in the results. The underlying hypothesis is that any instrumental and/or environmental factors influencing the color intensity in the detection zones will similarly impact the color intensity in the control zone under the same conditions. To test our design, a ratiometric microfluidic paper-based analytical device (μPAD), functionalized with a mixture of green-emissive carbon dots (CDs) and red-emissive ethidium bromide, was developed for the selective detection of ascorbic acid (AA). The green emission of the CDs is quenched by both AA and Fe3+; NaF was thus loaded onto the 3D connector as a masking agent to remove the interference effect of the Fe3+ ions. The color variations were monitored under a UV lamp, using a smartphone to capture the images, and the RGB intensities were processed using the Color Grab application. The proposed double-spot method greatly enhanced the analytical precision and accuracy of the device. A linear working range from 0 to 125 μM was obtained, and the limit of detection was 2.71 μM. The μPAD was successfully used for the detection of AA in human serum, with recoveries from 87.27 to 98.52 %.

Self-cleaning paper-based microfluidic biosensor employing DNAzyme and semiconducting single-walled carbon nanotube for copper ion detection

Microfluidic paper-based analytical device (μPAD) offers a simple and efficient platform for point-of-care monitoring, which can be beneficial for copper determination in livestock feed and manure. However, common cellulose paper has excellent hydrophilicity, causing μPAD is accompanied by poor mechanical properties, short service life, and low sensitivity. Here, a self-cleaning paper-based microfluidic biosensor for Cu2+ determination was proposed to overcome the mentioned shortages in the application. Polymeric octadecyl trichlorosilane was synthesized and decorated on cellulose paper to form hydrophobic paper, which can improve the hydrophobicity, self-cleaning, and pollution ability. In addition, hydrophobic paper, semiconducting single-walled carbon nanotube, and DNAzyme through the chemical bond were employed to fabricate a self-cleaning paper-based microfluidic biosensor. The properties were investigated using scanning electron microscopy, Raman, and electrochemical methods. The detecting parameters were also optimized. It could measure the Cu2+ concentration from 1 nM to 100 μM, and the detection limit was 0.65 nM. The self-cleaning paper-based microfluidic biosensor was applied to detect Cu2+ concentration in livestock feed and manure that can meet the requirements for fast screening and detection.

A Microfluidic Paper-based Analytical Device for Simultaneous Measurement of Albumin, Creatinine and Uric Acid in Urine based on Standard Addition Method

This work describes a simple method for the simultaneous measurement of urinary albumin, creatinine and uric acid using a standard addition approach on a microfluidic paper-based analytical device (µPAD). Hydrophobic barrier of the µPAD was created by stamping indelible ink onto a filter paper. The µPAD was designed with a flower-liked configuration. After aliquoting urine to a central sample zone, the sample flowed to ten surrounding channels, namely “the inner channels”, where the blank (water) or standard solutions had been added. The inner channels were connected to circular reagent zones where reagents had been immobilized. Tetrabromophenolphthalein ethyl ester, alkaline picrate, and a mixed solution of ferric chloride and ferric cyanide were used as the chromogenic reagents for the colorimetric detections of albumin, creatinine, and uric acid, respectively. Each reagent zone was linked with a circular detection zone through a second channel, namely “the outer channels”. The blue-, orange- and greenish-blue colored products were observed in the detection zones for the measurement of albumin, creatinine, and uric acid, correspondingly. A digital image of the µPAD was captured with a mobile phone. The color intensities were evaluated by ImageJTM and were employed for the quantitative analyses. The standard addition calibrations were found to be linear (r2 &gt; 0.99) for the spiked analyte concentrations up to 100 mg L-1 albumin, 1000 mg L-1 creatinine, and 50 mg dL-1 uric acid. The paper platform provided high precision (RSD &lt; 5 %) and good analytical recovery (91.8-109.7 %). Under paired t-test, the results obtained by the developed µPAD and the validating methods were not significantly different at 95 % confidence (albumin: tstat = -0.130, tcri = 3.182, creatinine: tstat = -0.133, tcri = 3.182, and uric acid: tstat = 1.119, tcri = 3.182).