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Chronic Kidney Disease (CKD) is a disorder that affects over 10% of the global population, and that would benefit from innovative methodologies that would provide early detection. Since it has been reported that there are high levels of urease in CKD patients' saliva, this sample is a promising non-invasive alternative to blood for CKD detection and monitoring. This work introduces a novel 3D µPAD for quantifying urease activity in saliva in a range of 0.041-0.750 U/mL, with limits of detection and quantification of 0.012 and 0.041 U/mL, respectively. The device uses the urease in the sample to convert urea into ammonia, causing a colorimetric change in the bromothymol blue. The accuracy of the developed device was evaluated by comparing the measurements of several saliva samples (#13) obtained with the μPAD and with a commercially available kit. Stability studies were also performed to assess its functionality as a point-of-care methodology, and the device was stable for 4 months when stored in a vacuum. After the sample placement, it could be scanned within 40 min without providing significantly different results. The developed device quantifies urease activity in saliva within 30 min, providing a simple, portable, lab-free alternative to existing methodologies.

Sildenafil is used to treat erectile dysfunction and pulmonary arterial hypertension but is often illicitly added to energy drinks and chocolates. This study introduces a lateral flow strip test using aptamers specific to sildenafil for detecting its illegal presence in food. The process involved using graphene oxide SELEX to identify high-affinity aptamers, which were then converted into molecular gate structures on mesoporous silica nanoparticles, creating a unique signaling system. This system was integrated into lateral flow chromatography strips and tested on buffers and chocolate samples containing sildenafil. The method simplifies the lateral flow assay (LFA) for small molecules and provides a tool for signal amplification. The detection limit for these strips was found to be 68.2 nM (31.8 µg/kg) in spiked food samples.

Escherichia coli (E. coli) detection suffers from slow analysis time and high costs, along with the need for specificity. While state-of-the-art electrochemical biosensors are cost-efficient and easy to implement, their sensitivity and analysis time still require improvement. In this work, we present a paper-based electrochemical biosensor utilizing magnetic core-shell Fe 2 O 3 @CdSe/ZnS quantum dots (MQDs) to achieve fast detection, low cost, and high sensitivity. Using electrochemical impedance spectroscopy (EIS) as the detection technique, the biosensor achieved a limit of detection of 2.7 × 10 2 CFU/mL for E. coli bacteria across a concentration range of 10 2 -10 8 CFU/mL, with a relative standard deviation (RSD) of 3.5781%. From an optical perspective, as E. coli concentration increased steadily from 10 4 to 10 7 CFU/mL, quantum dot fluorescence showed over 60% lifetime quenching. This hybrid biosensor thus provides rapid, highly sensitive E. coli detection with a fast analysis time of 30 min. This study, which combines the detection advantages of electrochemical and optical biosensor systems in a graphite-based paper sensor for the first time, has the potential to meet the needs of point-of-care applications. It is thought that future studies that will aim to examine the performance of the production-optimized, portable, graphite-based sensor system on real food samples, environmental samples, and especially medical clinical samples will be promising.

Crystal violet (CV) is a disinfectant and antifungal agent used in aquaculture that plays a vital role in treating aquatic diseases and sterilizing water. However, its potential for strong toxicity, including carcinogenicity and mutagenicity, upon accumulation in the body raises concerns regarding its safe use. Therefore, there is a growing need for the quantitative detection of CV in its early application stages to ensure human safety. Recently, Raman spectroscopy-based surface-enhanced Raman scattering (SERS) detection research has been actively conducted; consequently, an alveolar-mimicking SERS paper (AMSP) inspired by the structure of the human lungs was developed. The AMSP was optimized through various factors, including paper type, reducing agent, reducing agent concentration, and reaction time. This optimization enhanced the surface area of interaction with the target substances and promoted hotspot formation, resulting in enhanced SERS performance. The substrate exhibited exceptional uniformity, reproducibility, and reliability. CV was successfully detected at a concentration of 1 nM in laboratory settings. Furthermore, the AMSP detected CV at 1 nM in real-world environmental samples, including fish farm water and human serum, confirming its potential as a practical detection and monitoring platform for CV in real-world samples.

In response to growing environmental concerns, the scientific community is increasingly incorporating green chemistry principles into modern analytical techniques. Electrochemical paper-based analytical devices (ePADs) have emerged as a sustainable and efficient alternative to conventional analytical devices, offering robust applications in point-of-care testing, personalized healthcare, environmental monitoring, and food safety. ePADs align with green chemistry by minimizing reagent use, reducing energy consumption, and being disposable, making them ideal for eco-friendly and cost-effective analyses. Their user-friendly interface, alongside sensitive and selective detection capabilities, has driven their popularity in recent years. This review traces the evolution of ePADs from simple designs to complex multilayered structures that optimize analyte flow and improve detection. It also delves into innovative electrode fabrication methods, assessing key advantages, limitations, and modification strategies for enhanced sensitivity. Application-focused sections explore recent advancements in using ePADs for detecting diseases, monitoring environmental hazards like heavy metals and bacterial contamination, and screening contaminants in food. The integration of cutting-edge technologies, such as wearable wireless devices and the Internet of Things (IoT), further positions ePADs at the forefront of point-of-care testing (POCT). Finally, the review identifies key research gaps and proposes future directions for the field.

Determining the total antioxidant capacity (TAC) of biological samples is a valuable approach to measuring health status under oxidative stress conditions, such as infertility and type 2 diabetes. The Trolox equivalent antioxidant capacity (TEAC) assay is the most common approach to evaluating TAC in biological matrices. This assay is typically performed in clinical settings on a microtiter plate using a plate reader. However, the instrumentation and expertise requirements, and the resulting delay in the reporting of assay outcomes, make solution-based TEAC assays impractical for point-of-care or at-home testing, where individuals may want to monitor their health status during treatment. This work introduces the first microfluidic paper-based analytical device (µPAD) that measures TAC in human serum using TEAC assay chemistry. TAC was determined through a colorimetric image analysis of the degree of decolorization of 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) radical cations (ABTS ●+ ) by serum antioxidants. The µPAD showed a linear response to Trolox, ranging from 0.44 to 2.4 mM, (r  =  0.999). The performance of paper-based TEAC assays was validated through direct comparison to solution-based TEAC assays. There was a 0.04 mM difference in TAC values between the two platforms, well within one standard deviation of a standard solution-based assay conducted on an aliquot of the same serum sample (±0.25 mM). The µPAD had a limit of detection (LOD) of 0.20 mM, well below the TAC of normal human serum. The results suggest that the proposed device can be used for biological TAC determination and expands the field of TAC analysis in point-of-care health monitoring.

Cancer is the second leading cause of death globally, with 9.7 million fatalities in 2022. While routine screenings are vital for early detection, healthcare disparities persist, highlighting the need for equitable solutions. Recent advancements in cancer biomarker identification, particularly microRNAs (miRs), have improved early detection. MiR-21 is notably overexpressed in various cancers and can be a valuable diagnostic tool. Traditional detection methods, though accurate, are costly and complex, limiting their use in resource-limited settings. Paper-based electrochemical biosensors offer a promising alternative, providing cost-effective, sensitive, and rapid diagnostics suitable for point-of-care use. This study introduces an innovative electrochemical paper-based biosensor that leverages gold inkjet printing for the quantitative detection of miR-21. The biosensor, aimed at developing cost-effective point-of-care devices for low-resource settings, uses thiolated self-assembled monolayers to immobilize single-stranded DNA-21 (ssDNA-21) on electrodeposited gold nanoparticles (AuNPs) on the printed gold surface, facilitating specific miR-21 capture. The hybridization of ssDNA-21 with miR-21 increases the anionic barrier density, impeding electron transfer from the redox probe and resulting in a current suppression that correlates with miR-21 concentration. The biosensor exhibited a linear detection range from 1 fM to 1 nM miR-21 with a sensitivity of 7.69 fM µA -1 cm 2 and a rapid response time (15 min). With a low detection limit of 0.35 fM miR-21 in serum, the biosensor also demonstrates excellent selectivity against interferent species. This study introduces an electrochemical paper-based biosensor that uses gold inkjet printing to precisely detect miR-21, a key biomarker overexpressed in various cancers. This innovative device highlights the potential for cost-effective, accessible cancer diagnostics in underserved areas.

Lactate is a critical regulatory factor secreted by tumors, influencing tumor development, metastasis, and clinical prognosis. Precise analysis of tumor-cell-secreted lactate is pivotal for early cancer diagnosis. This study describes a paper-based microfluidic chip to enable the detection of lactate levels secreted externally by living cells. Under optimized conditions, the lactate biosensor can complete the assay in less than 30 min. In addition, the platform can be used to distinguish lactate secretion levels in different cell lines and can be applied to the screening of antitumor drugs. Through enzymatic chemical conversion, this platform generates fluorescent signals, enabling qualitative assessment under a handheld UV lamp and quantitative analysis via grayscale intensity measurements using ImageJ (Ver. 1.50i) software. The paper-based platform presented in this study is rapid and highly sensitive and does not necessitate other costly and intricate instruments, thus making it applicable in resource-constrained areas and serving as a valuable tool for investigating cell lactate secretion and screening various anti-cancer drugs.

This manuscript offers a concise overview of paper microfluidics, emphasizing its sustainable sensing applications in healthcare, environmental monitoring, and food safety. Researchers have developed innovative sensing platforms for detecting pathogens, pollutants, and contaminants by leveraging the paper's unique properties, such as biodegradability and affordability. These portable, low-cost sensors facilitate rapid diagnostics and on-site analysis, making them invaluable tools for resource-limited settings. This review discusses the fabrication techniques, principles, and applications of paper microfluidics, showcasing its potential to address pressing challenges and enhance human health and environmental sustainability.

Excessive emissions of heavy metals not only cause environmental pollution but also pose a direct threat to human health. Therefore, rapid and accurate detection of heavy metals in the environment is of great significance. Herein, we propose a method based on laser-induced breakdown spectroscopy (LIBS) combined with filter paper modified with bovine serum albumin-protected gold nanoclusters (LIBS-FP-AuNCs) for the rapid and sensitive detection of Cr 3+ and Mn 2+ . The filter paper modified with AuNCs was used to selectively enrich Cr 3+ and Mn 2+ . Combined with the multi-element detection capability of LIBS, this method achieved the simultaneous rapid detection of Cr 3+ and Mn 2+ . Both elements showed linear ranges for concentrations of 10-1000 μg L -1 , with limits of detection of 7.5 and 9.0 μg L -1 for Cr 3+ and Mn 2+ , respectively. This method was successfully applied to the determination of Cr 3+ and Mn 2+ in real water samples, with satisfactory recoveries ranging from 94.6% to 105.1%. This method has potential application in the analysis of heavy metal pollution.

The exploration into nanomaterial-based nonenzymatic biosensors with superb performance in terms of good sensitivity and anti-interference ability in disease marker monitoring has always attained undoubted priority in sensing systems. In this work, we report the design and synthesis of a highly active nanocatalyst, i.e., palladium and platinum nanoparticles (Pt&Pd-NPs) decorated ultrathin nanoporous gold (NPG) film, which is modified on a homemade graphene paper (GP) to develop a high-performance freestanding and flexible nanohybrid electrode. Owing to the structural characteristics the robust GP electrode substrate, and high electrochemically catalytic activities and durability of the permeable NPG support and ultrafine and high-density Pt&Pd-NPs on it, the resultant Pt&Pd-NPs-NPG/GP electrode exhibits excellent sensing performance of low detection limitation, high sensitivity and anti-interference capability, good reproducibility and long-term stability for the detection of small molecular biomarkers hydrogen peroxide (H 2 O 2 ) and glucose (Glu), and has been applied to the monitoring of H 2 O 2 in different types of live cells and Glu in body fluids such as urine and fingertip blood, which is of great significance for the clinical diagnosis and prognosis in point-of-care testing.

Herein, a cost-effective and portable microfluidic paper-based sensor is proposed for the simultaneous and rapid detection of glucose, free amino acids, and vitamin C in fruit. The device was constructed by embedding a poly(carboxybetaine acrylamide) (pCBAA)-modified cellulose paper chip within a hydrophobic acrylic plate. We successfully showcased the capabilities of a filter paper-based microfluidic sensor for the detection of fruit nutrients using three distinct colorimetric analyses. Within a single paper chip, we simultaneously detected glucose, free amino acids, and vitamin C in the vivid hues of cyan blue, purple, and Turnbull's blue, respectively, in three distinctive detection zones. Notably, we employed more stable silver nanoparticles for glucose detection, replacing the traditional peroxidase approach. The detection limits for glucose reached a low level of 0.049 mmol/L. Meanwhile, the detection limits for free amino acids and vitamin C were found to be 0.236 mmol/L and 0.125 mmol/L, respectively. The feasibility of the proposed sensor was validated in 13 different practical fruit samples using spectrophotometry. Cellulose paper utilizes capillary action to process trace fluids in tiny channels, and combined with pCBAA, which has superior hydrophilicity and anti-pollution properties, it greatly improves the sensitivity and practicality of paper-based sensors. Therefore, the paper-based colorimetric device is expected to provide technical support for the nutritional value assessment of fruits in the field of rapid detection.

The utilization of electrochemical detection techniques in paper-based analytical devices (PADs) has revolutionized point-of-care (POC) testing, enabling the precise and discerning measurement of a diverse array of (bio)chemical analytes. The application of electrochemical sensing and paper as a suitable substrate for point-of-care testing platforms has led to the emergence of electrochemical paper-based analytical devices (ePADs). The inherent advantages of these modified paper-based analytical devices have gained significant recognition in the POC field. In response, electrochemical biosensors assembled from paper-based materials have shown great promise for enhancing sensitivity and improving their range of use. In addition, paper-based platforms have numerous advantageous characteristics, including the self-sufficient conveyance of liquids, reduced resistance, minimal fabrication cost, and environmental friendliness. This study seeks to provide a concise summary of the present state and uses of ePADs with insightful commentary on their practicality in the field. Future developments in ePADs biosensors include developing novel paper-based systems, improving system performance with a novel biocatalyst, and combining the biosensor system with other cutting-edge tools such as machine learning and 3D printing.

Environmental toxic pollutants and pathogens that enter the ecosystem are major global issues. Detection of these toxic chemicals/pollutants and the diagnosis of a disease is a first step in efficiently controlling their contamination and spread, respectively. Various analytical techniques are available to detect and determine toxic chemicals/pathogens, including liquid chromatography, HPLC, mass spectroscopy, and enzyme-linked immunosorbent assays. However, these sensing strategies have some drawbacks such as tedious sample pretreatment and preparation, the requirement for skilled technicians, and dependence on large laboratory-based instruments. Alternatively, biosensors, especially paper-based sensors, could be used extensively and are a cost-effective alternative to conventional laboratory testing. They can improve accessibility to testing to identify chemicals and pollutants, especially in developing countries. Due to its low cost, abundance, easy disposal (by incineration, for example) and biocompatible nature, paper is considered a versatile material for the development of environmentally friendly electrochemical/optical (bio) sensor devices. This review presents an overview of sensing platforms constructed from paper, pointing out the main merits and demerits of paper-based sensing systems, their fabrication techniques, and the different optical/electrochemical detection techniques that they exploit.

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.

This article reports a simple and inexpensive leak-proof paper pad with an initial selection of a paper substrate on the grounds of surface morphology and fluid absorption time. Herein, a drying method is used for glucose detection on a paper pad through colorimetric analysis, and the spot detection of glucose is analyzed by optimizing the HRP concentration and volume to obtain accurate results. The rapid colorimetric method for the detection of glucose on the paper pad was developed with a limit of detection (LOD) of 2.92 mmol L -1 . Furthermore, the effects of the detection conditions were investigated and discussed comprehensively with the help of chemometric methods. Paper pads were developed for glucose detection with a range of 0.5-20 mM (apropos to the normal glucose level in the human body) and 0.1-0.5 M (to test the excessive intake of glucose). The developed concept has huge potential in the healthcare sector, and its extension could be envisioned to develop the reported paper pad as a point-of-care testing device for the initial screening of a variety of diseases.

The illegal use of β-adrenergic agonists during livestock growth poses a threat to public health; the long-term intake of this medication can cause serious physiological side effects and even death. Therefore, rapid detection methods for β-adrenergic agonist residues on-site are required. Traditional detection methods such as liquid chromatography have limitations in terms of expensive instruments and complex operations. In contrast, paper methods are low cost, ubiquitous, and portable, which has led to them becoming the preferred detection method in recent years. Various paper-based fluidic devices have been developed to detect β-adrenergic agonist residues, including lateral flow immunoassays (LFAs) and microfluidic paper-based analytical devices (μPADs). In this review, the application of LFAs for the detection of β-agonists is summarized comprehensively, focusing on the latest advances in novel labeling and detection strategies. The use of μPADs as an analytical platform has attracted interest over the past decade due to their unique advantages and application for detecting β-adrenergic agonists, which are introduced here. Vertical flow immunoassays are also discussed for their shorter assay time and stronger multiplexing capabilities compared with LFAs. Furthermore, the development direction and prospects for the commercialization of paper-based devices are considered, shedding light on the development of point-of-care testing devices for β-adrenergic agonist residue detection.

In vitro diagnosis (IVD) has become a hot topic in laboratory research and achievement transformation. However, due to the high cost, and time-consuming and complex operation of traditional technologies, some new technologies are being introduced into IVD, to solve the existing problems. As a result, IVD has begun to develop toward point-of-care testing (POCT), a subdivision field of IVD. The pandemic has made governments and health institutions realize the urgency of accelerating the development of POCT. Microfluidic paper-based analytical devices (μPADs), a low-cost, high-efficiency, and easy-to-operate detection platform, have played a significant role in advancing the development of IVD. μPADs are composed of paper as the core material, certain unique substances as reagents for processing the paper, and sensing devices, as auxiliary equipment. The published reviews on the same topic lack a comprehensive and systematic introduction to μPAD classification and research progress in IVD segmentation. In this paper, we first briefly introduce the origin of μPADs and their role in promoting IVD, in the introduction section. Then, processing and detection methods for μPADs are summarized, and the innovative achievements of μPADs in IVD are reviewed. Finally, we discuss and prospect the upgrade and improvement directions of μPADs, in terms of portability, sensitivity, and automation, to help researchers clarify the progress and overcome the difficulties in subsequent μPAD research.

Over the past decades, paper-based lateral flow immunoassays (LFIAs) have been extensively developed for rapid, facile, and low-cost detection of a wide array of target analytes in a point-of-care manner. Conventional home pregnancy tests are the most significant example of LFAs, which detect elevated concentrations of human chorionic gonadotrophin (hCG) in body fluids to identify early pregnancy. In this work, we have upgraded these platforms to a higher version by developing a customized microfluidic paper-based analytical device (μPAD), as the new generation of paper-based point-of-care platforms, for colorimetric immunosensing. This will offer a cost-efficient and environmentally friendly alternative platform for paper-based immunosensing, eliminating the need for nitrocellulose (NC) membrane as the substrate material. The performance of the developed platform is demonstrated by detection of hCG (as a model case) in urine samples and subsequently indicating positive or negative pregnancy. A dual-functional silane-based composite was used to treat filter paper in order to enhance the colorimetric signal intensity in the detection zones of μPADs. In addition, microfluidic pathways were designed in a manner to provide the desired regulated fluid flow, generating sufficient incubation time (delays) at the designated detection zones, and consequently enhancing the obtained signal intensity. The presented approaches allow to overcome the existing limitations of μPADs in immunosensing and will broaden their applicability to a wider range of assays. Although, the application of the developed hCG μPAD assay is mainly in qualitative (i.e., positive or negative) detection of pregnancy, the semi-quantitative measurement of hCG was also investigated, indicating the viability of this assay for sensitive detection of the target hCG analyte within the related physiological range (i.e., 10-500 ng/mL) with a LOD value down to 10 ng/mL.

In this study, we describe a monolithic and fully integrated paper-based device for the simultaneous detection of three prognostic biomarkers in saliva. The pattern of the proposed multiplexed device is designed with a central sample deposition zone and three identical arms, each containing a pre-treatment and test zone. Its one-step fabrication is realized by CO 2 laser cutting, providing remarkable parallelization and rapidity (ca. 5 s/device). The colorimetric detection is based on the sensitive and selective target-induced reshaping of plasmonic multibranched gold nanoparticles, which exhibit a clear spectral shift (and blue-to-pink color change) in case of non-physiological concentrations of the three salivary biomarkers. A rapid and multiplexed naked-eye or smartphone-based readout of the colorimetric response is achieved within 10 min. A prototype kit for POCT testing is also reported, providing robustness and easy handling of the device.

In the last 10 years, paper-based electrochemical biosensors have gathered attention from the scientific community for their unique advantages and sustainability vision. The use of papers in the design the electrochemical biosensors confers to these analytical tools several interesting features such as the management of the solution flow without external equipment, the fabrication of reagent-free devices exploiting the porosity of the paper to store the reagents, and the unprecedented capability to detect the target analyte in gas phase without any sampling system. Furthermore, cost-effective fabrication using printing technologies, including wax and screen-printing, combined with the use of this eco-friendly substrate and the possibility of reducing waste management after measuring by the incineration of the sensor, designate these type of sensors as eco-designed analytical tools. Additionally, the foldability feature of the paper has been recently exploited to design and fabricate 3D multifarious biosensors, which are able to detect different target analytes by using enzymes, antibodies, DNA, molecularly imprinted polymers, and cells as biocomponents. Interestingly, the 3D structure has recently boosted the self-powered paper-based biosensors, opening new frontiers in origami devices. This review aims to give an overview of the current state origami paper-based biosensors, pointing out how the foldability of the paper allows for the development of sensitive, selective, and easy-to-use smart and sustainable analytical devices.

The fast detection of trace amounts of hazardous contaminations can prevent serious damage to the environment. Paper-based sensors offer a new perspective on the world of analytical methods, overcoming previous limitations by fabricating a simple device with valuable benefits such as flexibility, biocompatibility, disposability, biodegradability, easy operation, large surface-to-volume ratio, and cost-effectiveness. Depending on the performance type, the device can be used to analyze the analyte in the liquid or vapor phase. For liquid samples, various structures (including a dipstick, as well as microfluidic and lateral flow) have been constructed. Paper-based 3D sensors are prepared by gluing and folding different layers of a piece of paper, being more user-friendly, due to the combination of several preparation methods, the integration of different sensor elements, and the connection between two methods of detection in a small set. Paper sensors can be used in chromatographic, electrochemical, and colorimetric processes, depending on the type of transducer. Additionally, in recent years, the applicability of these sensors has been investigated in various applications, such as food and water quality, environmental monitoring, disease diagnosis, and medical sciences. Here, we review the development (from 2010 to 2021) of paper methods in the field of the detection and determination of toxic substances.

In recent years, microfluidic lab-on-paper devices have emerged as a rapid and low-cost alternative to traditional laboratory tests. Additionally, they were widely considered as a promising solution for point-of-care testing (POCT) at home or regions that lack medical infrastructure and resources. This review describes important advances in microfluidic lab-on-paper diagnostics for human health monitoring and disease diagnosis over the past five years. The review commenced by explaining the choice of paper, fabrication methods, and detection techniques to realize microfluidic lab-on-paper devices. Then, the sample pretreatment procedure used to improve the detection performance of lab-on-paper devices was introduced. Furthermore, an in-depth review of lab-on-paper devices for disease measurement based on an analysis of urine samples was presented. The review concludes with the potential challenges that the future development of commercial microfluidic lab-on-paper platforms for human disease detection would face.

Paper-based analytical devices (PADs) and Electrospun Fiber-Based Biosensors (EFBs) have aroused the interest of the academy and industry due to their affordability, sensitivity, ease of use, robustness, being equipment-free, and deliverability to end-users. These features make them suitable to face the need for point-of-care (POC) diagnostics, monitoring, environmental, and quality food control applications. Our work introduces new and experienced researchers in the field to a practical guide for fibrous-based biosensors fabrication with insight into the chemical and physical interaction of fibrous materials with a wide variety of materials for functionalization and biofunctionalization purposes. This research also allows readers to compare classical and novel materials, fabrication techniques, immobilization methods, signal transduction, and readout. Moreover, the examined classical and alternative mathematical models provide a powerful tool for bioanalytical device designing for the multiple steps required in biosensing platforms. Finally, we aimed this research to comprise the current state of PADs and EFBs research and their future direction to offer the reader a full insight on this topic.

This review summarizes the state of the art of paper-based biosensors (PBBs) for coronavirus disease 2019 (COVID-19) detection. Three categories of PBB are currently being been used for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) diagnostics, namely for viral gene, viral antigen and antibody detection. The characteristics, the analytical performance, the advantages and drawbacks of each type of biosensor are highlighted and compared with traditional methods. It is hoped that this review will be useful for scientists for the development of novel PBB platforms with enhanced performance for helping to contain the COVID-19 outbreak, by allowing early diagnosis at the point of care (POC).

A simple and rapid distance paper-based analytical device (dPAD) for the detection of lead (Pb) in foods is proposed herein. The assay principle is based on competitive binding between carminic acid (CA) and polyethyleneimine (PEI) to Pb in a food sample. The paper channels were pre-immobilized with PEI, before reacting with a mixture of the sample and CA. Pb can strongly bind to the CA; hence, the length of the red color deposition on the flow channel decreased as a lower amount of free CA bound to PEI. The dPAD exhibited good linear correlation, with ranges of 5-100 µg·mL -1 ( R 2 = 0.974) of Pb. Although, the limit of detection (LOD) of this platform was rather high, at 12.3 µg·mL -1 , a series of standard additions (8.0, 9.0, and 10.0 µg·mL -1 ) can be used to interpret the cutoff of Pb concentrations at higher or lower than 2 µg·mL -1 . The presence of common metal ions such as calcium, magnesium, nickel, and zinc did not interfere with the color distance readout. The validity of the developed dPAD was demonstrated by its applicability to screen the contamination of Pb in century egg samples. The results obtained from the dPAD are in accordance with the concentration measured by atomic absorption spectroscopy (AAS) ( n = 9). In conclusion, this proposed dPAD, combined with the standard addition method, could be applied for screening Pb contamination in food matrices. This platform is, therefore, potentially applicable for field measurements of Pb in developing countries, because it is cheap and rapid, and it requires no significant laborious instruments.

This paper describes the use of fused deposition modeling (FDM) printing to fabricate paper-based analytical devices (PAD) with three-dimensional (3D) features, which is termed as 3D-PAD. Material depositions followed by heat reflow is a standard approach for the fabrication of PAD. Such devices are primarily two-dimensional (2D) and can hold only a limited amount of liquid samples in the device. This constraint can pose problems when the sample consists of organic solvents that have low interfacial energies with the hydrophobic barriers. To overcome this limitation, we developed a method to fabricate PAD integrated with 3D features (vertical walls as an example) by FDM 3D printing. 3D-PADs were fabricated using two types of thermoplastics. One thermoplastic had a low melting point that formed hydrophobic barriers upon penetration, and another thermoplastic had a high melting point that maintained 3D features on the filter paper without reflowing. We used polycaprolactone (PCL) for the former, and polylactic acid (PLA) for the latter. Both PCL and PLA were printed with FDM without gaps at the interface, and the resulting paper-based devices possessed hydrophobic barriers consisting of PCL seamlessly integrated with vertical features consisting of PLA. We validated the capability of 3D-PAD to hold 30 μL of solvents (ethanol, isopropyl alcohol, and acetone), all of which would not be retained on conventional PADs fabricated with solid wax printers. To highlight the importance of containing an increased amount of liquid samples, a colorimetric assay for the formation of dimethylglyoxime (DMG)-Ni (II) was demonstrated using two volumes (10 μL and 30 μL) of solvent-based dimethylglyoxime (DMG). FDM printing of 3D-PAD enabled the facile construction of 3D structures integrated with PAD, which would find applications in paper-based chemical and biological assays requiring organic solvents.

Screen-printed technology has helped considerably to the development of portable electrochemical sensors since it provides miniaturized but robust and user-friendly electrodes. Moreover, this technology allows to obtain very versatile transducers, not only regarding their design, but also their ease of modification. Therefore, in the last decades, the use of screen-printed electrodes (SPEs) has exponentially increased, with ceramic as the main substrate. However, with the growing interest in the use of cheap and widely available materials as the basis of analytical devices, paper or other low-cost flat materials have become common substrates for SPEs. Thus, in this revision, a comprehensive overview on paper-based SPEs used for analytical proposes is provided. A great variety of designs is reported, together with several examples to illustrate the main applications.

This paper presents a lateral flow assay (LFA) for the quantitative, fluorescence-based detection of the cardiac biomarker troponin I (cTnI) that features an analytical strip made of cellulose filter paper. The results show that the wicking and test time are comparable to those obtained with conventional nitrocellulose (NC)-based LFAs. Further, the cellulose paper provides an excellent background with no auto-fluorescence that is very adequate in detecting fluorescent lines. While fluorescence that was generated with cellulose strips was lower when compared to that generated in NC strips, signals could be improved by layering carbon nanofibers (CNF) on the cellulose. A nonlinear behavior of the concentration-response relationship was observed for the LFA architectures with NC, cellulose, and cellulose-CNF in the 0 to 200 ng/mL cTnI concentration range. The measurements were consistent and characterized by coefficients of variation lower than 2.5%. Detection and quantitation limits that were in the range 1.28-1.40 ng/mL and 2.10-2.75 ng/mL were obtained for LFA with cellulose and cellulose CNF strips that are equivalent to the limits obtained with the standard NC LFA. Overall, we showed that commercially available filter paper can be used in the analytical strip of LFA.

Paper-based microfluidic analysis devices (μPADs) have attracted attention as a cost-effective platform for point-of-care testing (POCT), food safety, and environmental monitoring. Recently, three-dimensional (3D)-μPADs have been developed to improve the performance of μPADs. For accurate diagnosis of diseases, however, 3D-μPADs need to be developed to simultaneously detect multiple biomarkers. Here, we report a 3D-μPADs platform for the detection of multiple biomarkers that can be analyzed and diagnosed with a smartphone. The 3D-μPADs were fabricated using a 3D digital light processing printer and consisted of a sample reservoir (300 µL) connected to 24 detection zones (of 4 mm in diameter) through eight microchannels (of 2 mm in width). With the smartphone application, eight different biomarkers related to various diseases were detectable in concentrations ranging from normal to abnormal conditions: glucose (0-20 mmol/L), cholesterol (0-10 mmol/L), albumin (0-7 g/dL), alkaline phosphatase (0-800 U/L), creatinine (0-500 µmol/L), aspartate aminotransferase (0-800 U/L), alanine aminotransferase (0-1000 U/L), and urea nitrogen (0-7.2 mmol/L). These results suggest that 3D-µPADs can be used as a POCT platform for simultaneous detection of multiple biomarkers.

Screen-printing technology has revolutionized many fields, including that of electrochemical biosensing. Due to their current relevance, this review, unlike other papers, discusses the relevant aspects of electrochemical biosensors manufactured using this technology in connection to both paper substrates and wearable formats. The main trends, advances, and opportunities provided by these types of devices, with particular attention to the environmental and biomedical fields, are addressed along with illustrative fundamentals and applications of selected representative approaches from the recent literature. The main challenges and future directions to tackle in this research area are also pointed out.

Given their photoluminescent character, portable quantum dot readers are often sophisticated and relatively expensive. In response, we engineered a "do it yourself" fluorescence reader employing paper materials and a mid-range smartphone camera. Black paperboard facilitated a versatile, lightweight and foldable case; whereas cellophane paper was observed to behave as a simple, yet effective, optical bandpass filter leading to an advantageous device for the quantitative interrogation of quantum dot nanocrystals concentrations (from 2.5 to 20 nM), which are suitable for optical point-of-care biosensing. The streptavidin-coated nanocrystals employed are commercially available and the developed reader was benchmarked with a standard portable quantum dot reader, thereby demonstrating advantages in terms of cost and linear analytical range.

Paper-based carbon working electrodes were modified with mercury or bismuth films for the determination of trace metals in aqueous solutions. Both modification procedures were optimized in terms of selectivity and sensitivity for the determination of different heavy metals, aiming their simultaneous determination. Cd (II), Pb (II) and In (III) could be quantified with both films. However, Cu (II) could not be determined with bismuth films. The modification with mercury films led to the most sensitive method, with linear ranges between 0.1 and 10 µg/mL and limits of detection of 0.4, 0.1, 0.04 and 0.2 µg/mL for Cd (II), Pb (II), In (III) and Cu (II), respectively. Nevertheless, the bismuth film was a more sustainable alternative to mercury. Tap-water samples were analyzed for the determination of metals by standard addition methodology with good accuracy, by using a low-cost and easily disposable paper-based electrochemical platform. This system demonstrated its usefulness for monitoring heavy metals in water., Competing Interests: The authors declare no conflict of interest.

The excessive use of antibiotics in food-producing animals causes a steady rise of multiple antibiotic resistance in foodborne bacteria. Next to sulfonamides, the most common antibiotics groups are fluoroquinolones, aminoglycosides, and ß-lactams. Therefore, there is a need for a quick, efficient, and low-cost detection procedure for antibiotics. In this study, we propose an inkjet-printed aptamer-based biosensor developed for the detection of the fluoroquinolone ciprofloxacin. Due to their extraordinary high affinity and specificity, aptamers are already widely used in various applications. Here we present a ciprofloxacin-binding RNA aptamer developed by systematic evolution of ligands by exponential enrichment (SELEX). We characterized the secondary structure of the aptamer and determined the KD to 36 nM that allow detection of antibiotic contamination in a relevant range. We demonstrate that RNA aptamers can be inkjet-printed, dried, and resolved while keeping their functionality consistently intact. With this proof of concept, we are paving the way for a potential range of additional aptamer-based, printable biosensors.