Microfluidic Biosensors for Point-of-Care Nucleic Acid Amplification Tests

This Special Issue of Biosensors, “Microfluidic Biosensors for Point-of-Care Nucleic Acid Amplification Tests” aims to gather original research papers and comprehensive reviews detailing novel research, fabrication methods, and applications, as well as the challenges and prospects of developing microfluidics for improved biosensing and diagnostics [...

Recent Methods for the Viability Assessment of Bacterial Pathogens: Advances, Challenges, and Future Perspectives

Viability assessment is a critical step in evaluating bacterial pathogens to determine infectious risks to public health. Based on three accepted viable criteria (culturability, metabolic activity, and membrane integrity), current viability assessments are categorized into three main strategies. The first strategy relies on the culturability of bacteria. The major limitation of this strategy is that it cannot detect viable but nonculturable (VBNC) bacteria. As the second strategy, based on the metabolic activity of bacteria, VBNC bacteria can be detected. However, VBNC bacteria sometimes can enter a dormant state that allows them to silence reproduction and metabolism; therefore, they cannot be detected based on culturability and metabolic activity. In order to overcome this drawback, viability assessments based on membrane integrity (third strategy) have been developed. However, these techniques generally require multiple steps, bulky machines, and laboratory technicians to conduct the tests, making them less attractive and popular applications. With significant advances in microfluidic technology, these limitations of current technologies for viability assessment can be improved. This review summarized and discussed the advances, challenges, and future perspectives of current methods for the viability assessment of bacterial pathogens

Fabrication of a foldable all-in-one point-of-care molecular diagnostic microdevice for the facile identification of multiple pathogens

In the present study, we fabricated a foldable all-in-one microdevice for the nucleic acid test (NAT) performing DNA extraction, polymerase chain reaction (PCR), and on-site colorimetric detection for point‐of‐care (POC) molecular diagnosis of multiple pathogens. The microdevice is composed of three layers. The top layer contains six filter paper discs pre-stored with PCR reagent. The middle layer contains six through-holes corresponding to the positions of six paper discs on the top layer, and was sealed by a flat bottom layer, forming six main chambers where commercially available FTA cards are stored. The intact bacteria samples were introduced into six main chambers for sample treatment – specifically, the isolation, purification, and storage of nucleic acids inside the FTA cards. After the main chambers were filled with water, the top layer was folded towards the middle layer. After 30 thermal cycles for PCR, targets were colorimetrically detected on chip by loading silver nitrate solution into the main chambers and illuminated by UV at 234 nm for 30 s. To examine the versatile applicability, the microdevice was used for the direct detection of three major foodborne pathogens: Salmonella spp., Staphylococcus aureus, and Escherichia coli O157:H7. The feasibility of the microdevice was also tested for detecting multidrug-resistant bacteria, Acinetobacter baumannii. Using the microdevice, multiple pathogens were detected with high specificity and sensitivity of 3.0×102 and 3.0×103 CFU/sample for Gram-negative (Salmonella spp. and Escherichia coli O157:H7) and Gram-positive (Staphylococcus aureus) bacteria, respectively, in less than 2 h, making the microdevice highly suitable in low-resource settings

Pressure-Free Assembling of Poly(methyl methacrylate) Microdevices via Microwave-Assisted Solvent Bonding and Its Biomedical Applications

Poly(methyl methacrylate) (PMMA) has become an appealing material for manufacturing microfluidic chips, particularly for biomedical applications, because of its transparency and biocompatibility, making the development of an appropriate bonding strategy critical. In our research, we used acetic acid as a solvent to create a pressure-free assembly of PMMA microdevices. The acetic acid applied between the PMMA slabs was activated by microwave using a household microwave oven to tightly merge the substrates without external pressure such as clamps. The bonding performance was tested and a superior bond strength of 14.95 ± 0.77 MPa was achieved when 70% acetic acid was used. Over a long period, the assembled PMMA device with microchannels did not show any leakage. PMMA microdevices were also built as a serpentine 2D passive micromixer and cell culture platform to demonstrate their applicability. The results demonstrated that the bonding scheme allows for the easy assembly of PMMAs with a low risk of clogging and is highly biocompatible. This method provides for a simple but robust assembly of PMMA microdevices in a short time without requiring expensive instruments

Bonding Strategies for Thermoplastics Applicable for Bioanalysis and Diagnostics

Microfluidics is a multidisciplinary science that includes physics, chemistry, engineering, and biotechnology. Such microscale systems are receiving growing interest in applications such as analysis, diagnostics, and biomedical research. Thermoplastic polymers have emerged as one of the most attractive materials for microfluidic device fabrication owing to advantages such as being optically transparent, biocompatible, cost-effective, and mass producible. However, thermoplastic bonding is a key challenge for sealing microfluidic devices. Given the wide range of bonding methods, the appropriate bonding approach should be carefully selected depending on the thermoplastic material and functional requirements. In this review, we aim to provide a comprehensive overview of thermoplastic fabricating and bonding approaches, presenting their advantages and disadvantages, to assist in finding suitable microfluidic device bonding methods. In addition, we highlight current applications of thermoplastic microfluidics to analyses and diagnostics and introduce future perspectives on thermoplastic bonding strategies

Recent Advances in Molecular and Immunological Diagnostic Platform for Virus Detection: A Review

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused an ongoing coronavirus disease (COVID-19) outbreak and a rising demand for the development of accurate, timely, and cost-effective diagnostic tests for SARS-CoV-2 as well as other viral infections in general. Currently, traditional virus screening methods such as plate culturing and real-time PCR are considered the gold standard with accurate and sensitive results. However, these methods still require sophisticated equipment, trained personnel, and a long analysis time. Alternatively, with the integration of microfluidic and biosensor technologies, microfluidic-based biosensors offer the ability to perform sample preparation and simultaneous detection of many analyses in one platform. High sensitivity, accuracy, portability, low cost, high throughput, and real-time detection can be achieved using a single platform. This review presents recent advances in microfluidic-based biosensors from many works to demonstrate the advantages of merging the two technologies for sensing viruses. Different platforms for virus detection are classified into two main sections: immunoassays and molecular assays. Moreover, available commercial sensing tests are analyzed

Recent Progress in Nanotechnology-Based Approaches for Food Monitoring

Throughout the food supply chain, including production, storage, and distribution, food can be contaminated by harmful chemicals and microorganisms, resulting in a severe threat to human health. In recent years, the rapid advancement and development of nanotechnology proposed revolutionary solutions to solve several problems in scientific and industrial areas, including food monitoring. Nanotechnology can be incorporated into chemical and biological sensors to improve analytical performance, such as response time, sensitivity, selectivity, reliability, and accuracy. Based on the characteristics of the contaminants and the detection methods, nanotechnology can be applied in different ways in order to improve conventional techniques. Nanomaterials such as nanoparticles, nanorods, nanosheets, nanocomposites, nanotubes, and nanowires provide various functions for the immobilization and labeling of contaminants in electrochemical and optical detection. This review summarizes the recent advances in nanotechnology for detecting chemical and biological contaminations in the food supply chain

Design Strategy and Application of Deep Eutectic Solvents for Green Synthesis of Nanomaterials

The first report of deep eutectic solvents (DESs) was released in 2003 and was identified as a new member of ionic liquid (IL), involving innovative chemical and physical characteristics. Using green solvent technology concerning economical, practical, and environmental aspects, DESs open the window for sustainable development of nanomaterial fabrication. The DESs assist in different fabrication processes and design nanostructures with specific morphology and properties by tunable reaction conditions. Using DESs in synthesis reactions can reduce the required high temperature and pressure conditions for decreasing energy consumption and the risk of environmental contamination. This review paper provides the recent applications and advances in the design strategy of DESs for the green synthesis of nanomaterials. The strategy and application of DESs in wet-chemical processes, nanosize reticular material fabrication, electrodeposition/electrochemical synthesis of nanostructures, electroless deposition, DESs based nano-catalytic and nanofluidic systems are discussed and highlighted in this review

Fluorescence Enhancement Using Bimetal Surface Plasmon-Coupled Emission from 5-Carboxyfluorescein (FAM)

We demonstrate the enhancement of fluorescence emission from a dye, 5-carboxyfluorescein (FAM), which couples with surface plasmons at the spectral channels of excitation and emission. Experiments and calculations revealed that bimetallic (gold-silver) plasmon, as compared to the monometallic ones, allowed such coupling to be enhanced, at both the spectral channels. We achieved the maximum fluorescence enhancement level of 46.5-fold, with markedly high reproducibility (coefficient of variation ~ 0.5%) at a FAM concentration of 10 nM. We also found that higher fluorescence enhancement was more likely to be reproducible. This encourages the use of this technology for practical applications in fluorescence-based biochemical assays. Moreover, we investigated a FAM concentration-dependent enhancement of fluorescence. It was found that fluorescence enhancement decreased and saturated at above 10 nM concentration possibly due to partial photo-bleaching of FAM molecules

Fabrication of Polymerase Chain Reaction Plastic Lab-on-a-Chip Device for Rapid Molecular Diagnoses

Purpose: We aim to fabricate a thermoplastic poly(methylmethacrylate) (PMMA) Lab-on-a-Chip device to perform continuous- flow polymerase chain reactions (PCRs) for rapid molecular detection of foodborne pathogen bacteria. Methods: A miniaturized plastic device was fabricated by utilizing PMMA substrates mediated by poly(dimethylsiloxane) interfacial coating, enabling bonding under mild conditions, and thus avoiding the deformation or collapse of microchannels. Surface characterizations were carried out and bond strength was measured. The feasibility of the Lab-on-a-Chip device for performing on-chip PCR utilizing a lab-made, portable dual heater was evaluated. The results were compared with those obtained using a commercially available thermal cycler. Results: A PMMA Lab-on-a-Chip device was designed and fabricated for conducting PCR using foodborne pathogens as sample targets. A robust bond was established between the PMMA substrates, which is essential for performing miniaturized PCR on plastic. The feasibility of on-chip PCR was evaluated using Escherichia coli O157:H7 and Cronobacter condimenti, two worldwide foodborne pathogens, and the target amplicons were successfully amplified within 25 minutes. Conclusions: In this study, we present a novel design of a low-cost and high-throughput thermoplastic PMMA Lab-on-a-Chip device for conducting microscale PCR, and we enable rapid molecular diagnoses of two important foodborne pathogens in minute resolution using this device. In this regard, the introduced highly portable system design has the potential to enable PCR investigations of many diseases quickly and accurately