Microfluidic Paper-based Analytical Devices Research Articles

Modeling of Paper-Based Bi-Material Cantilever Actuator for Microfluidic Biosensors.

This research explores the dynamics of a fluidically loaded Bi-Material cantilever (B-MaC), a critical component of μPADs (microfluidic paper-based analytical devices) used in point-of-care diagnostics. Constructed from Scotch Tape and Whatman Grade 41 filter paper strips, the B-MaC's behavior under fluid imbibition is examined. A capillary fluid flow model is formulated for the B-MaC, adhering to the Lucas-Washburn (LW) equation, and supported by empirical data. This paper further investigates the stress-strain relationship to estimate the modulus of the B-MaC at various saturation levels and to predict the behavior of the fluidically loaded cantilever. The study shows that the Young's modulus of Whatman Grade 41 filter paper drastically decreases to approximately 20 MPa (about 7% of its dry-state value) upon full saturation. This significant decrease in flexural rigidity, in conjunction with the hygroexpansive strain and coefficient of hygroexpansion (empirically deduced to be 0.008), is essential in determining the B-MaC's deflection. The proposed moderate deflection formulation effectively predicts the B-MaC's behavior under fluidic loading, emphasizing the measurement of maximum (tip) deflection using interfacial boundary conditions for the B-MaC's wet and dry regions. This knowledge of tip deflection will prove instrumental in optimizing the design parameters of B-MaCs.

Use of scanners for colorimetric analysis of microfluidic paper-based analytical devices (µPADs): A practical guide

Office style scanners are often used by researchers for the colorimetric detection in the development of microfluidic paper-based analytical devices (µPADs). However, little attention has been paid to nuances in calibration curve responses caused by varying settings of a single image-capturing device. Hence, in this study, the effect of parameters, such as the scanner operation conditions, the method of data analysis used (e.g., choice of colour space system, suitable channel), and the influence of the physical characteristics of the µPAD (e.g., template colour, type of lamination pouches used) on the colorimetric detection were evaluated for the first time.Combinations of the scanning parameters were thoroughly examined and their impact on the quality of a mock calibration curve assessed. The most sensitive calibration curve was achieved only by the selection of the optimal scanning conditions combined with an appropriate choice of colorimetric system. A flatbed scanner, used in this study, showed greater variation of signal with varying scanning setup, whereas a portable scanner provided nearly identical responses under diverse scanning conditions. Gamma correction, scanning resolution, region of interest (ROI) surrounding colour (i.e., template colour), bit-depth, image file type, and the number of analysed pixels were found not to markedly affect the dynamics of the calibration curve when colour in the ROI is homogeneous. Parameters that had an effect on the colorimetric detection were the scanner settings, colour of the blank, image acquisition mode (colour vs greyscale), and µPAD covering (e.g., lamination). Based on these findings, generic guidelines for optimal data acquisition and µPAD manipulation with respect to µPAD calibration have been formulated.

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.

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

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.

Capillary Flow-Driven Microfluidics Combined with a Paper Device for Fast User-Friendly Detection of Heavy Metals in Water.

Human exposure to heavy metals is a concerning global problem because of its detrimental effect on our health and ecosystem. Assessing the levels of these metals is cost- and labor-intensive and nonuser friendly because current analysis approaches typically rely on heavy instrumentations like inductively coupled plasma-mass spectrometry, which is only possible in centralized labs. Hence, simple economical detection methods are in high demand in developing countries and areas with insufficient infrastructure, professional experts, and appropriate environmental treatment. Several microfluidic paper-based analytical devices have been reported as promising alternatives to conventional testing methods for on-site heavy metal detection. Paper-based microfluidics are advantageous because of their simple fabrication, biodegradability, low cost, and ability to operate without pumps. However, typical assay times for current platforms are slow, and they typically rely on pipetting a fixed volume into the assay cards. This adds complexity in actual field scenarios. Here, we report a novel, inexpensive, and straightforward capillary-driven microfluidic device combined with paper for rapid and user-friendly detection of Ni(II), Cu(II), and Fe(III) in water. A colorimetric approach was adopted to quantify these metals. The device was able to produce a homogeneous color signal within 8 s of sample insertion. The limit of detection and limit of quantification were calculated to be 2 and 6.67 ppm for nickel, 0.3 and 1 ppm for Cu, and 1.1 and 3.67 ppm for Fe, respectively. The majority (>90%) of the collected samples showed recovery in the 80-110% range with acceptable accuracy and precision (<15% RSD) for a colorimetric device. This technique can be beneficial for rapidly assessing heavy metal exposure in drinking and surface water at drastically reduced assay time and is the first example of capillary flow-driven microfluidic devices as a transport medium for heavy metal detection.

Hollow microneedle microfluidic paper-based chip for biomolecules rapid sampling and detection in interstitial fluid

The interstitial fluid (ISF) contains rich bioinformation for disease diagnosis and healthcare monitoring. However, the efficient sampling and detection of the biomolecules in ISF is still challenging. Herein, we develop a facile but versatile ISF analysis platform by combining controllable hollow microneedles (HMNs) and elaborate microfluidic paper-based analytical devices (μPADs). The HMNs and μPADs was fixed in a bottom PDMS layer. A top PDMS layer containing a cylindrical cavity to produce negative pressure for sampling was packaged on the bottom PDMS layer. The HMNs enable efficient and swift sampling of sufficient ISF to the μPADs through one-touch finger operation without extra manipulations. The μPADs realized to simultaneously detect glucose and lactic acid in the detection area to produce chromogenic agents and analyzed by the self-programed RGB application (APP) in smartphones. The HMN microfluidic paper-based chip provides a point-of-care platform for accurate detection of biomolecules in ISF, holding great promise in the development of wearable device.

Biomarker Detection in Early Diagnosis of Cancer: Recent Achievements in Point-of-Care Devices Based on Paper Microfluidics

Microfluidics is very crucial in lab-on-a-chip systems for carrying out operations in a large-scale laboratory environment on a single chip. Microfluidic systems are miniaturized devices in which the fluid behavior and control can be manipulated on a small platform, with surface forces on the platform being greater than volumetric forces depending on the test method used. In recent years, paper-based microfluidic analytical devices (μPADs) have been developed to be used in point-of-care (POC) technologies. μPADs have numerous advantages, including ease of use, low cost, capillary action liquid transfer without the need for power, the ability to store reagents in active form in the fiber network, and the capability to perform multiple tests using various measurement techniques. These benefits are critical in the advancement of paper-based microfluidics in the fields of disease diagnosis, drug application, and environment and food safety. Cancer is one of the most critical diseases for early detection all around the world. Detecting cancer-specific biomarkers provides significant data for both early diagnosis and controlling the disease progression. μPADs for cancer biomarker detection hold great promise for improving cure rates, quality of life, and minimizing treatment costs. Although various types of bioanalytical platforms are available for the detection of cancer biomarkers, there are limited studies and critical reviews on paper-based microfluidic platforms in the literature. Hence, this article aims to draw attention to these gaps in the literature as well as the features that future platforms should have.

Fabrication of microfluidic paper-based channels by inkjet printing process for analytical applications

Microfluidic paper-based channels play an important role in microfluidic paper-based analytical devices (μPADs). There are some fabrication methods which could be utilised to fabricate microfluidic channels on paper substrate. Among these methods, inkjet printing process is considered as a promising fabrication method with many advantages such as low-cost, material saving, high precision, etc. The aim of this work is to apply inkjet printing technology to fabricate paper channels of μPADs. A new design of μPAD was proposed in this paper to demonstrate how to fabricate inkjet-printed hydrophobic lines to make paper-based biosensor. Biological target of our μPADs is human chOrionic gonadotropin (hCG). Colorimetric signals from μPADs were captured by digital camera and measured by ImageJ software, which showed that these μPADs can determine hCG in the range from 1,000 to 10,000 ng ml−1. These results showed that piezoelectric inkjet printing technology can fabricate 250 μm-width hydrophobic lines on paper substrate, helping in fabricating μPADs in next applications.

Rapid detection of three mycotoxins in animal feed materials using competitive ELISA-based origami microfluidic paper analytical device (μPAD).

We report the development of a competitive ELISA-based origami microfluidic paper-based analytical device (μPAD) for the detection of mycotoxins in animal feed material. The μPAD was patterned using the wax printing technique with the design of a testing pad in the middle and two absorption pads at the side. Anti-mycotoxin antibodies were effectively immobilized on chitosan-glutaraldehyde-modified sample reservoirs in the μPAD. The determination of zearalenone, deoxynivalenol, and T-2 toxin in corn flour was successfully achieved by performing competitive ELISA on the μPAD in 20min. Colorimetric results were easily distinguished by the naked eye with a detection limit of 1µg/mL for all three mycotoxins. The μPAD integrated with competitive ELISA holds potential for practical applications in thelivestock industry for rapid, sensitive, and cost-effective detection of different mycotoxins in animal feed materials.

Near-Field Communication Tag for Colorimetric Glutathione Determination with a Paper-Based Microfluidic Device

Here, we propose a microfluidic paper-based analytical device (µPAD) implemented with a near-field communication (NFC) tag as a portable, simple and fast colorimetric method for glutathione (GSH) determination. The proposed method was based on the fact that Ag+ could oxidize 3,3',5,5'-tetramethylbenzidine (TMB) into oxidized blue TMB. Thus, the presence of GSH could cause the reduction of oxidized TMB, which resulted in a blue color fading. Based on this finding, we developed a method for the colorimetric determination of GSH using a smartphone. A µPAD implemented with the NFC tag allowed the harvesting of energy from a smartphone to activate the LED that allows the capture of a photograph of the µPAD by the smartphone. The integration between electronic interfaces into the hardware of digital image capture served as a means for quantitation. Importantly, this new method shows a low detection limit of 1.0 µM. Therefore, the most important features of this non-enzymatic method are high sensitivity and a simple, fast, portable and low-cost determination of GSH in just 20 min using a colorimetric signal.

"Do it yourself" protocol to fabricate dual-detection paper-based analytical device for salivary biomarker analysis.

This paper describes the design and construction of dual microfluidic paper-based analytical devices (dual-μPADs) as a lab-on-paper platform involving a "do-it-yourself" fabrication protocol. The device comprises a colorimetric and electrochemical module to obtain a dual-mode signal readout sensing strategy. A 3D pen polymeric resin was used to prepare graphite carbon-based electrodes and hydrophobic barriers on paper substrates. The proposed carbon-based ink was employed to manufacture electrodes on paper based on a stencil-printing approach, which were further characterized by electrochemical and morphological analyses. The analytical performance of the dual-μPADs was simultaneously evaluated for lactate, pH, nitrite, and salivary amylase (sAA) analysis. To demonstrate the proof-of-concept, saliva samples collected from both healthy individuals and those with periodontitis were successfully tested to demonstrate the feasibility of the proposed devices. Samples collected from individuals previously diagnosed with periodontitis showed high levels of nitrite and sAA (> 94μmol L-1 and > 610 U mL-1) in comparison with healthy individuals (≤ 16μmol L-1 and 545 U mL-1). Moreover, periodontitis saliva resulted in acid solution and almost null lactate levels. Notably, this protocol supplies a simple way to manufacture dual-μPADs, a versatile platform for sensitive detecting of biomarkers in saliva playing a crucial role towards the point-of-care diagnosis of periodontal disease.

Construction of hydrophobic-hydrophilic microfluidic patterns with controlled wettability on PVDF filter paper surface using maskless microplasma scanning

Paper-based microfluidic analytical devices (μPADs) have attracted increasing interest due to their unique advantages. Localized hydrophilization on hydrophobic paper surfaces to form the microfluidic patterns and their wettability control are the key issues in μPADs preparation. This paper introduces a novel method to construct hydrophobic-hydrophilic microfluidic patterns on the surface of the Poly(vinylidene fluoride) (PVDF) filter paper by maskless microplasma writing. The hydrophilic performances and surface morphology of the patterns were characterized by water contact angle (WCA) and scanning electron microscopy (SEM). Fourier transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS) were used to analyze their chemical compositions. The results showed that paper surface changed from hydrophobic to hydrophilic and its wettability can be controlled through plasma parameters adjustment. PVDF layer was gradually removed from the filter paper and the fluorine-containing groups on paper surface of the micropatterns decreased with increasing microplasma treatment time. Besides, the polar functional groups (e.g. –C=O) also appeared and increased with longer treatment time. Finally, the mechanism of the microfluidic patterns construction by microplasma was discussed, and the microfluidic pattern with the pattern “H” were demonstrated and successfully constructed. This work shows great potential for simple, fast, environmental friendly and low-cost construction of μPADs.

Iron Determination in Natural Waters Using a Synthesised 3-Hydroxy-4-Pyridione Ligand in a Newly Developed Microfluidic Paper-Based Device

This work describes the development of an iron sensor based on a microfluidic paper-based technique, to attain iron quantification in natural waters. A new water-soluble naphthalene-3-hydroxy-4-pyridione ligand was used as a colour reagent, as it formed an orange complex with iron. As a newly described ligand, several chemical and physical parameters, namely, the sample and reagent volumes and reagent concentrations, were studied related to the formation of the coloured complex. The microfluidic paper-based analytical device (μPAD) assembly, namely, the use of different types of filter paper and different numbers of layers, was developed to obtain the best performance. Under the optimal conditions, a linear correlation was obtained in the range of 0.25–2.00 mg/L of iron, with a minimum detectable value of 0.07 mg/L. The proposed μPAD method was validated by an analysis of the certified samples and by a comparison of the tested water samples with the inductively coupled plasma (ICP) results (RE &lt; 10%). Then, the μPAD device was successfully applied to the determination of iron in tap water, well water, river water, and seawater, with no need for any prior sample pre-treatment; recovery studies were also performed (average = 100.3% with RSD = 4.2%).

Targeting iron speciation in wines: Design of a microfluidic paper-based device for determination of iron(II) and iron(III)

During the winemaking process, wine is frequently exposed to oxygen which can affect the organoleptic characteristics and therefore the quality of the finished wine. Most phenolic molecules in wine serve as oxidation substrates affecting colour and flavour. Transition metal ions namely copper and iron act as electron pumps; in fact, Fe(III) is reduced to Fe(II) as a consequence of the oxidation of phenols to quinones and, in the presence of oxygen, Fe(II) is oxidised back to Fe(III). Consequently, speciation analysis by the oxidation state is critical and even more relevant than the total ion concentration when one wants to evaluate the risk for wine oxidation. Interestingly the different reactivity of Fe(III) and Fe(II) makes spectrophotometric methods, namely colorimetric reactions, ideally suited for speciation analysis. In this work, a microfluidic paper-based analytical device (µPAD) was designed for the iron speciation in wine samples. The colorimetric reaction between bathophenanthroline and iron(II) was chosen and the oxidation of Fe(III) to Fe(II), using hydroxylamine, was carried out within the same device. Aiming for application to different types of wine, the developed device also enabled to account for the potential sample colour (subtraction of interfering signal). The designed µPAD enabled the iron speciation in both white and red wines.

MagnEtophoretic Slider Assay (MeSA): A simple platform for point-of-care diagnostics

Infectious diseases account for millions of deaths each year. To reduce the number of infectious disease related deaths, diagnostic testing needs to be more accessible to patients in low-income countries as well as developed countries. Current diagnostic methods involve centralized laboratories, trained personnel, and are time-intensive, limiting translation to the point-of-care (POC). Microfluidic devices are a popular alternative for diagnostics due to reduced assay times, reduced sample volume, and lower cost. Microfluidic devices are small (&amp;lt;10 cm) and can perform complex assays. Microfluidic paper-based analytical devices (µPADs) are a popular approach to help translate diagnostics to the POC but historically suffer from poor sensitivity when compared to established laboratory methods. Magnetically labeling analytes allows samples to be sorted resulting in improved sensitivity and specificity. Microfluidic magnetophoresis is the process of manipulating magnetic particles in a magnetic field and offers the ability to wash and concentrate a sample during flow. However, until recently, magnetophoresis has not been used in conjunction with µPADs because magnetophoresis requires complex and expensive instrumentation to control flow. Coupling magnetophoresis with µPADs enables pump-free flow control, simple operation, and low cost. Early magnetophoresis µPADs showed detection limits similar to traditional methods but higher than targets for clinical use. In this work, we demonstrate a novel, simple MagnEtophoretic Slider Assay (MeSA) that is free of any external instrumentation and offers a new platform for POC diagnostics. We demonstrate the assay’s capability through biotin competitive assays and a sandwich immunoassay for E. coli detection. The calculated limit of detection for E. coli was 1.62 × 103 Colony Forming Units per mL (CFU/ml). The work described is a novel and simple microfluidic platform that has potential for a wide range of future applications.