Paper-based Analytical Devices Research Articles

Paper-based colorimetric loop-mediated isothermal amplification (LAMP) assay for the identification of latent Colletotrichum in harvested fruit

Paper-based analytical devices (PADs) have gained enormous attention because of their low-cost, simple fabrication, and portability. Here, we propose a paper-based device for performing reverse transcription loop-mediated isothermal amplification (RT-LAMP) with real-time simultaneous detection of C. gloeosporioides latent infections in tomatoes. RT-LAMP-based PAD platform comprises a paper substrate on which the DNA amplification reaction occurs. Among different types of tested papers, cellulose membrane (grade 4) enabled effective visualization of the amplification result. The assay was found highly selective for the latent stage of C. gloeosporioides with lower limit of detection (LOD) of 0.5 pg of total extracted RNA. The developed assay generated the results within 40 min and hence can be efficiently employed for identifying C. gloeosporioides in resource-limited settings.

A non-oxidation reduction strategy for highly selective detection of ascorbic acid by “on-off-on” photoluminescence switches

Great progress was made to detect ascorbic acid (AA) based on the photoluminescent nanoprobes/oxidant complexes by using the reducibility of AA. However, some reducing substances including cysteine and glutathione disturbed the accuracy of AA assay. Herein, a non-oxidation reduction regulation strategy was fabricated to construct a novel “on-off-on” photoluminescence model for highly selective determination of AA. Initially, copper nanoclusters capsulated into zeolitic imidazolate framework-8 (CuNCs@ZIF-8) was achieved. And then inorganic phosphate ions quenched the emission of CuNCs@ZIF-8 owing to the decomposition of ZIF-8 shell, rendering the signal in “turn-off” state. The linear range for phosphate ions was 1.0–60 μM with LOD of 0.35 μM. Interestingly, upon introducing AA into CuNCs@ZIF-8/phosphate, the emission of CuNCs@ZIF-8 recovered possibly due to the electronegativity improvement of metal core and surface rigidified through hydrogen bond interaction, and the system signal was “turned-on”. The LOD of AA was as low as 51 nM in a good linear range from 0.1 to 40 μM. Furthermore, CuNCs@ZIF-8 were used for AA detecting in tablets, beverages, and fresh fruits with the satisfactory recoveries from 96% to 104%. More importantly, a paper-based analytical device containing CuNCs@ZIF-8/phosphate were evaluated for the visual sensing of AA in food safety.

Nanozymes paper−based analytical device for the detection of organophosphate pesticides in fruits and vegetables

In this work, copper oxide nanoparticles (CuONPs) nanozymes paper-based analytical device was designed for the rapid detection of organophosphate pesticides in fruits and vegetables. The paper-based analytical device was modified with silica oxide nanoparticles to enhance the assay sensitivity. CuO nanozymes displayed peroxidase-like activity and catalyzed the oxidation of o-dianisidine in the presence of H2O2 from the hydrolysis of acetylthiocholine. This results in the formation of a brown−colored product. In the presence of organophosphate pesticides such as malathion, acetylcholinesterase activity was inhibited, resulting in reduced color intensity production, and which was measured with a smartphone. The proposed nanozymes paper-based analytical device exhibited a good linear detection range (0.1–5 mg L−1), a low detection limit of 0.08 mg L−1, and the analysis time was only about 10 min for malathion detection under optimal conditions. Moreover, the CuONPs had excellent catalytic activity and higher stability than peroxidase. Finally, this device can be applied to detect organophosphate pesticides in fruits and vegetables with rapidity, accuracy, portability, and ease of handling in the field.

Silica quantum dots; an optical nanosensing approach for trace detection of pesticides in environmental and biological samples

Both the environment and human health have suffered as a result of excessive and irrational pesticide use. The human body is vulnerable to a wide range of illnesses brought on by prolonged exposure to or intake of food contaminated with pesticide residues, including immunological and hormonal abnormalities and the development of certain tumors. Sensors based on nanoparticles stand out from more conventional spectrophotometry analytical methods due to their low detection limits, high sensitivity, and ease of use; that is why the demand for simple, fast, and less expensive sensing methods increases daily and presents myriad uses. Such demands are fulfilled by employing paper-based analytical devices having intrinsic properties. The presented work reports an on-site, easy-to-handle, and disposable paper-based sensing device for performing fast screening along with readout from a smartphone. The fabricated device utilizes luminescent silica quantum dots, immobilized into a paper cellulose matrix, and the resonance energy transfer phenomenon is employed. The silica quantum dots probes were fabricated from citric acid and, by undergoing physical adsorption, were confined on the nitrocellulose substrate in small wax-traced spots. The silica quantum dots were excited by smartphone ultraviolet LED, acting as an energy source and for capturing the image. The obtained LOD is 0.054 μM, and the coefficient of variation is less than 6.1%, comparable to the result obtained by UV–Visible and fluorometric analysis under similar experimental conditions. In addition, high reproducibility (≥9.8%) and high recovery ≥90% were obtained in spiked blood samples. The fabricated sensor sensitively detected pesticides giving a LOD of 2.5 ppm along with the development of yellow color within a short period of 5 min. The sensor functions well when sophisticated instrumentation is not accessible. The presented work shows the potential of the paper strip for the on-site detection of pesticides in biological and environmental samples.

A Dye-Assisted Paper-Based Assay to Rapidly Differentiate the Stress of Chlorophenols and Heavy Metals on Enterococcus faecalis and Escherichia coli

Biological toxicity testing plays an essential role in identifying the possible negative effects induced by substances such as organic pollutants or heavy metals. As an alternative to conventional methods of toxicity detection, paper-based analytical device (PAD) offers advantages in terms of convenience, quick results, environmental friendliness, and cost-effectiveness. However, detecting the toxicity of both organic pollutants and heavy metals is challenging for a PAD. Here, we show the evaluation of biotoxicity testing for chlorophenols (pentachlorophenol, 2,4-dichlorophenol, and 4-chlorophenol) and heavy metals (Cu2+, Zn2+, and Pb2+) by a resazurin-integrated PAD. The results were achieved by observing the colourimetric response of bacteria (Enterococcus faecalis and Escherichia coli) to resazurin reduction on the PAD. The toxicity responses of E. faecalis-PAD and E. coli-PAD to chlorophenols and heavy metals can be read within 10 min and 40 min, respectively. Compared to the traditional growth inhibition experiments for toxicity measuring which takes at least 3 h, the resazurin-integrated PAD can recognize toxicity differences between studied chlorophenols and between studied heavy metals within 40 min.

Cover Feature: Enhanced Chemiluminescence of a Superior Luminol Derivative Provides Sensitive Smartphone‐Based Point‐of‐Care Testing with Enzymatic μPAD (Anal. Sens. 4/2023)

The cover image illustrates the principle of a newly developed chemiluminescence (CL) microfluidic paper-based analytical device (μPAD) that enables highly sensitive detection of any catalytic process in which H2O2 is produced. Through the use of a new, sensitive CL luminophore cell phone camera detection allows quantification on-site with the same sensitivity as afforded through a CCD camera in the lab. One-step reactions to time the catalytic reactions can easily be realized through a moveable barrier if desired. Lactate is detected with μM detection limits and three orders of magnitude dynamic range in sweat, the luminophore reaches a pM detection limit. Cover design by Simone Rink. More information can be found in the Research Article by Antje Baeumner and co-workers .

A novel photonic chemosensor for rapidly detecting synthetic dyes in orange juice using colorimetric and spectrophotometric methods

Synthetic dyes must be monitored and regulated. We aimed to develop a novel photonic chemosensor for rapidly monitoring synthetic dyes based on colorimetric (chemical interactions with optical probes using microfluidic paper-based analytical devices) and UV–Vis spectrophotometric methods. Various types of gold and silver nanoparticles were surveyed to identify the targets. In the presence of silver nanoprisms, the naked eye could visualize the unique and distinctive color changes of Tartrazine (Tar) to green and Sunset Yellow (Sun) to brown; UV–Vis spectrophotometry validated the results. The developed chemosensor showed linear ranges of 0.07–0.3 mM and 0.05–0.2 mM for Tar and Sun, respectively. Sources of interference had minimal effects, confirming the appropriate selectivity of the developed chemosensor. Our novel chemosensor demonstrated excellent analytical performance for measuring Tar and Sun in several types of orange juice as real samples, confirming its incredible potential for use in the food industry.

Mickey mouse-shaped laminated paper-based analytical device in simultaneous total cholesterol and glucose determination in whole blood

The microfluidic paper-based analytical device (μPAD) platform is gaining attention as a low-cost, portable, and disposable detection tool. However, the limitations of traditional fabrication methods include poor reproducibility and the use of hydrophobic reagents. In this study, an in-house computer-controlled X–Y knife plotter and pen plotter were used to fabricate μPADs, resulting in a simple, more rapid, reproducible process that consumes less volume of reagents. The μPADs were laminated to increase mechanical strength and reduce sample evaporation during analysis. The resulting laminated paper-based analytical device (LPAD) was used to simultaneously determine glucose and total cholesterol in whole blood using the LF1 membrane as a sample zone. The LF1 membrane selectively separates plasma from whole blood by size exclusion and yields plasma for further enzymatic reaction steps while retaining blood cells and larger proteins. The i1 Pro 3 mini spectrophotometer directly detected color on the LPAD. The results were clinically relevant and in agreement with hospital methods, with a detection limit of 0.16 mmol L⁻1 for glucose and 0.57 mmol L⁻1 for TC. The LPAD retained color intensity after 60 days of storage. The LPAD offers a low-cost, high-performance option for chemical sensing devices and expands the applicability of markers for diagnosing whole blood samples.

A label-free colorimetric 3D paper-based device for ochratoxin A detection using G-quadruplex/hemin DNAzyme with a smartphone readout

The colorimetric sensor usually depends on enzyme-mediated signal amplification to achieve trace analysis of ochratoxin A (OTA) residues in food samples. However, the enzyme labeling and manual addition of reagents steps increased assay time and operation complexity, restricting their application in point-of-care testing (POCT). Herein, we report a label-free colorimetric device integrating a 3D paper-based analytical device and a smartphone as handheld readout for rapid and sensitive detection of OTA. Using vertical-flow design, the paper-based analytical device enables the specific recognition of target and self-assembly of G-quadruplex (G4)/hemin DNAzyme to be performed, then employs DNAzyme for transducing the OTA binding event signal into a colorimetric signal. The design of independent functional units, including biorecognition unit, self-assembly unit and colorimetric units, which can address crowding and disorder of biosensing interfaces and improve the recognition efficiency of aptamer (apta). In addition, we eliminated signal losses and nonuniform coloring by introducing carboxymethyl chitosan (CMCS) to obtain perfectly focused signals on colorimetric unit. On the basis of parameter optimization, the device exhibited a detection range of 0.1–500 ng/mL and a detection limit of 41.9 pg/mL for OTA. Importantly, good results were obtained in spiked real samples, indicating applicability and reliability of developed device.

Semi-quantitative microfluidic paper-based analytical device for ionic silica detection.

Silicate ions (SiO32-), or ionic silica, are known to cause silica scaling in industrial water applications when excess amounts are present; hence, concentrations must be monitored and kept at a constant low level. Ionic silica is conventionally measured by spectrophotometry in the form of its silicomolybdic complex based on the molybdenum blue reaction, but the operation process is complicated and not suitable for on-site detection. To solve these issues, microfluidic paper-based analytical devices (µPADs) have been gaining attention as portable, low-cost analytical devices suitable for on-site detection. The foldable origami type device described in this work enabled silica detection based on the molybdenum blue reaction, in the range of 50-1000mg/L, with a practically detectable lowest concentration of 50mg/L. The device showed selectivity for silicate ions and stability over 21days when stored at 4°C. The semi-quantitative analytical performance makes the proposed paper-based device attractive for on-site industrial monitoring.

Iodometric Titration on Microfluidic Paper-Based Analytical Devices for Determination of Ascorbic Acid: A Laboratory Experiment for Chemical Education Undergraduates

Iodometric Titration on Microfluidic Paper-Based Analytical Devices for Determination of Ascorbic Acid: A Laboratory Experiment for Chemical Education Undergraduates

Highly sensitive paper-based immunoassay: Combining nanomaterials modified cellulose with covalent and oriented immobilization of antibody

Among the analytical tools, paper-based analytical devices (PADs) have become a leading alternative for point-of care testing (POCT). In this study, PADs were fabricated using an office laser printer. Then, the paper zone was modified with graphene oxide (GO) and pyrene derivatives, which provide a sufficient amount of carboxylic groups for conjugating antibodies. At an optimal pH, antibodies were covalently bound onto carboxylated cellulose surface in an oriented manner through a two-step strategy: electrostatic adsorption was followed by EDC/NHS coupling. α-fetoprotein (AFP) as a detection model, we compared with cellulose powder modified and unmodified paper zone. The results showed the color intensity and color uniformity on GO modified paper was improved. The activity of immobilized antibodies on GO/1-pyrenebutyric acid (GO/PBA) modified was three times higher than that of GO modified and about 1.8-fold higher than that of GO/1-pyrenecarboxylic acid (GO/PCA) modified. The GO/PBA modified paper-based immunoassay has enhanced sensitivity and low detection limit. A linear correlation between color intensity and concentration of AFP in the range of 0.01～16.5 ng mL−1 with a detection limit of 9.0 pg mL−1 were achieved, respectively. The obtained results point towards rapid, sensitive, and specific early diagnosis of liver cancer at the point of care and other low-resource settings.

Paper-based analytical devices for point-of-need applications

Paper-based analytical devices (PADs) are powerful platforms for point-of-need testing sincethey are inexpensive devices fabricated in different shapes and miniaturized sizes, ensuring better portability. Additionally, the readout and detection systems can be accomplished with portable devices, allying with the features of both systems. These devices have been introduced as promising analytical platforms to meet critical demands involving rapid, reliable, and simple testing. They have been applied to monitor species related to environmental, health, and food issues. Herein, an outline of chronological events involving PADs is first reported. This work also introduces insights into fundamental parameters to engineer new analytical platforms, including the paper type and device operation. The discussions involve the main analytical techniques used as detection systems, such as colorimetry, fluorescence, and electrochemistry. It also showed recent advances involving PADs, especially combining optical and electrochemical detection into a single device. Dual/combined detection systems can overcome individual barriers of the analytical techniques, making possible simultaneous determinations, or enhancing the devices' sensitivity and/or selectivity. In addition, this review reports on distance-based detection, which is also considered a trend in analytical chemistry. Distance-based detection offers instrument-free analyses and avoids user interpretation errors, which are outstanding features for analyses at the point of need, especially for resource-limited regions. Finally, this review provides a critical overview of the practical specifications of the recent analytical platforms involving PADs, demonstrating their challenges. Therefore, this work can be a highlyuseful reference for new research and innovation.

Distance-based paper microfluidics as environmentally friendly platforms for monitoring acid-base titrations

The goal of green chemistry is to reduce the impact of chemical processes on the environment. In this context, it is important to develop new analytical tools that comply with the principles of green analytical chemistry (GAC) to contribute to the sustainable development of the planet. This study describes the development of an environmentally friendly methodology using paper-based analytical devices (µPADs) to perform acid-base titrations by distance-based measurements. The devices were composed of a circular zone defined with 5 mm diameter connected to a rectangular channel of 40 mm long × 3 mm wide. The titrations of acidic (strong and weak) and alkaline (strong and weak) solutions were successfully demonstrated using 10 µL of titrant solution and 10 µL of sample solution. The solution added into the zone travels different distances depending on the concentration of the titrated, thus promoting the channel coloration or discoloration. The distance measurements covered by the titrated solutions were obtained recorded with a digital caliper. As proof-of-concept for quantitative purposes, the µPAD was then used to determine the acetic acid content in commercial vinegar. The average distances ranged from to 32.0 to 32.5 mm and the concentration values ranged from 4.3 to 4.6% (m/v). Conventional acid-base volumetry was used as a control method, and the obtained results demonstrated no statistical difference at 95% confidence level. Lastly, the 12 principles of GAC were analyzed to evaluate the suitability of the proposed method. Considering a unified scale between 0 and 1, the final green value was 0.72. Titrations with distance-based detection on µPADs were found to be a technique with fast responses and low cost that can be used as a replacement for conventional practices in teaching laboratories or adapted for the detection of different analytes.

Nanomaterials integrated with microfluidic paper-based analytical devices for enzyme-free glucose quantification

In this study, nanomaterials capable of enzyme-free glucose quantification and colorimetric readout are integrated into a microfluidic paper-based analytical devices (μPADs). Gold nanoparticles (AuNPs) and silver nanoparticles (AgNPs) were utilized as a peroxidase-like nanozyme and a colorimetric probe to achieve glucose monitoring. In this developed device, glucose is oxidized by AuNPs to generate hydrogen peroxide (H2O2), which flows in the paper microchannels toward detection zones. H2O2 then etches the immobilized AgNPs to induce a color change. The intensity of color change is easily monitored using a smartphone application. Following method optimization, we obtained a linear range from 0.50 to 10.0 mmol L−1 (R2 = 0.9921) and a detection limit (LOD) of 340.0 μmol L−1. This falls in the clinically relevant range for glucose monitoring and diabetes diagnosis in humans. In addition, the total analysis time is just 20 min, which is significantly less than the same experiment performed in the solution phase. Also, our method is markedly selective; other substrates do not interfere. The recovery test in human control samples was in the range of 98.47–102.34% and the highest relative standard deviation (RSD) was 3.58%. The enzyme-free approach for glucose sensing is highly desirable for diabetes diagnosis as it replaces the more expensive enzyme with cheaper nanomaterials. Furthermore, since nanomaterials are more environmentally stable compared to enzymes, it has the potential for widespread deployment as point-of-care diagnostics (POC) in resource-limited settings.

Deep learning-assisted ultra-accurate smartphone testing of paper-based colorimetric ELISA assays

Smartphone has long been considered as one excellent platform for disease screening and diagnosis, especially when combined with microfluidic paper-based analytical devices (μPADs) that feature low cost, ease of use, and pump-free operations. In this paper, we report a deep learning-assisted smartphone platform for ultra-accurate testing of paper-based microfluidic colorimetric enzyme-linked immunosorbent assay (c-ELISA). Different from existing smartphone-based μPAD platforms, whose sensing reliability is suffered from uncontrolled ambient lighting conditions, our platform is able to eliminate those random lighting influences for enhanced sensing accuracy. We first constructed a dataset that contains c-ELISA results (n=2048) of rabbit IgG as the model target on μPADs under eight controlled lighting conditions. Those images are then used to train four different mainstream deep learning algorithms. By training with these images, the deep learning algorithms can well eliminate the influences of lighting conditions. Among them, the GoogLeNet algorithm gives the highest accuracy (>97%) in quantitative rabbit IgG concentration classification/prediction, which also provides 4% higher area under curve (AUC) value than that of the traditional curve fitting results analysis method. In addition, we fully automate the whole sensing process and achieve the "image in, answer out" to maximize the convenience of the smartphone. A simple and user-friendly smartphone application has been developed that controls the whole process. This newly developed platform further enhances the sensing performance of μPADs for use by laypersons in low-resource areas and can be facilely adapted to the real disease protein biomarkers detection by c-ELISA on μPADs.

Paper-based analytical device coupled with Bi-MOF: Electric field amplification and fluorescence sensing of glyphosate.

Glyphosate, a potent herbicide wildly used in the world, involves potential hazards to human health by accumulating in the food chain. Due to its absence of chromophores and fluorophores, the rapid visual detection of glyphosate has always been difficult. Herein, a paper-based geometric field amplification device visualized by the amino-functionalized bismuth-based metal-organic framework (NH2-Bi-MOF) was constructed for sensitive fluorescence determination of glyphosate. Fluorescence of the synthesized NH2-Bi-MOF was immediately enhanced by interaction with glyphosate. The field amplification of glyphosate was implemented by coordinating the electric field and the electroosmotic flow, which was orchestrated by the geometric configuration of paper channel and the concentration of polyvinyl pyrrolidone, respectively. Under the optimal conditions, the developed method exhibited a linear range of 0.80-200μmolL-1 with about 12500-fold signal enhancement achieved by just 100s electric field amplification. It was applied to soil and water with recoveries between 95.7% and 105.6%, holding great prospects in on-site analysis of hazardous anions for environment safety.

Direct Detection of Water-dissolved Ammonia Using Paper-based Analytical Devices

Direct Detection of Water-dissolved Ammonia Using Paper-based Analytical Devices

Smartphone and paper-based device for glucose monitoring using acetylene black-hemin nanozyme as catalyst

Glucose management is an important part of disease control for diabetes patients, thus the development of a rapid and real-time point of care testing (POCT) device for monitoring blood glucose is of great significance. In this work, a paper-based analytical device (PAD) is constructed by combining acetylene black (AB)-hemin complex modified filter paper as sensing platform with a smartphone as signal detector. Large specific surface area of AB decreases the self-associate and aggregate of hemin in aqueous solution, resulting in improved peroxidase-like activity of hemin. Compared with graphene oxide supported hemin, AB-hemin exhibits superior signal response on paper. Glucose oxidase (GOx) catalyzes the conversion of blood glucose to hydrogen peroxide, and then AB-hemin complex catalyzes the oxidation of colorless 3,3′,5,5′-tetramethylbenzidine (TMB) to blue TMB oxidized products (TMB+) in the presence of hydrogen peroxide, thus achieving the visual detection of blood glucose. In optimal conditions, PAD provides an applicable linear range from 0.2 mM to 30 mM and a low limit of detection (LOD) (0.06 mM). Notably, the detection accuracy of the developed paper-based sensor is in good agreement with that of the commercially available blood glucose meter (p > 0.05). Moreover, the proposed PAD presents high recoveries from 95.4% to 112% (RSD ≤ 3.2%), and therefore holds great potential for glucose monitoring and diabetes diagnosis.

UV photometric assays on paper-based analytical devices by contact printing photography through transparent cellophane

A limitation of paper-based devices that employ optical readout is that they can only be used for assays in the visible range of the electromagnetic spectrum. This work describes the first paper-based analytical devices that can be used for UV absorption photometric assays without scientific equipment. The devices consist of a transparent cellulose film (cellophane) as a reaction vessel and photometric cuvette and a photosensitive paper as a detector of UV irradiation. The analytes or their derivatives absorb part of the incident UV irradiation on the cellophane surface while the transmitted irradiation passes through the cellophane and photoreduces the surface of the photosensitive paper, producing an image. In this manner, the “colorimetrically invisible” UV absorption signaling is transformed into a visible image that can be captured and processed with common imaging devices. The analytical utility of these devices is demonstrated, as a proof-of-concept, for six UV-absorption model assays. The results show that cellophane-analytical devices are suitable for high-sensitivity measurements but exhibit a relatively large standard deviation, dictating multiple trials to correct the accuracy of the results. Technical aspects related to the improvement of the devices and the quality of analytical data are elucidated and discussed.