Point Of Care Diagnostics Research Articles

Rapid reagent-free anaemia screening using plant-derived “HemoQR” paper-strips and smartphone: A study on 200 human subjects

Detecting anaemia in resource-limited settings is challenging due to limited access to diagnostic tools, healthcare infrastructure, and trained personnel, often resulting in underdiagnosis and delayed treatment. This study aims to evaluate HemoQR, a novel point-of-care (POC) diagnostic method for haemoglobin (Hb) level detection using a smartphone-based system integrated with a paper strip without needing any reagent or additional auxiliary device. The goal is to assess the efficacy of this portable, user-friendly, rapid and accurate alternative for Hb measurement in large-scale intervention programmes such as Anaemia-mukt Bharat (AMB), which is crucial for diagnosing anaemia and monitoring various health conditions. The method involves the transfer of one drop of the blood sample to a paper strip that is made indigenously using plant based cellulosic sources for optimal functionality. The colour signal around the test spot is quantitatively analysed using a smartphone camera equipped with a custom application. The application processes the images through a machine learning algorithm to determine the Hb concentration. The system was evaluated in a clinical trial involving 200 participants, comparing its performance with the gold-standard laboratory Hb assay using automated haematology analyser. The smartphone-based method demonstrated high sensitivity (87.09 %) and specificity (96.11 %) for anaemia screening, as compared with laboratory results. The average detection time was significantly reduced to under q minute, and the system showed excellent user-friendliness and portability with no auxiliary device needed beyond the smartphone. Clinical trial results indicated that the device was accurate and reliable across different Hb levels. Summarily, HemoQR evidenced to be a promising tool for POC diagnostics. Its high accuracy, rapid results, and ease of use make it suitable for both clinical settings and remote areas with limited access to laboratory facilities. Future research will prioritize further validation and integration of the anaemia screening outcome with electronic health records from a large population cohort. This will help identify early indicators and signals of numerous emerging disease burdens that could escalate into secondary and tertiary health crises if not addressed in time.

B-237 Engineered Reverse Transcriptases and Taq DNA Polymerases deliver robust yield in the presence of clinically relevant inhibitors

Abstract Background The past five years has seen explosive growth in molecular diagnostic assays that rely on sensitive and accurate detection of nucleic acids across a wide range of sample types. As the assays are required to become increasingly sensitive, the need for highly functional and purpose-built enzymatic tools has grown. Reverse transcriptase (RT) and Taq DNA Polymerase (Taq DNAP) are commonly used to detect RNA and/or DNA viruses in patient samples for point of care (POC) diagnostics. However, crude patient samples introduce inhibitors that decrease enzyme activity and increase false negative results in POC testing. In addition, there is a need for faster turn-around time for patient diagnosis which can be enabled by engineering more active and sensitive Taq variants. Herein, we present on our engineered RT and Taq DNAP enzymes that were developed to confer greater activity, thermostability, inhibitor tolerance and/or sensitivity. Methods We take a 3-pronged approach to protein engineering: rational design, computational design and directed evolution. Using these techniques, we identified and characterized many RT and Taq DNAP variants. We deeply characterized each RT variant by measuring processivity, inhibitor tolerance and thermostability. First strand synthesis was done at temperatures from 42-65°C in the presence and absence of various clinically relevant inhibitors followed by qPCR to assess cDNA yield. Next, we characterized the inhibitor tolerance of the Taq variants by performing qPCR reactions in the presence of inhibitors. To determine if the Taq variant would be an appropriate fit for fast PCR, the polymerization activity of the Taq DNAP variants were characterized using a primer extension assay. Results Reverse transcriptases termed; StellarScript, StellarScript HT and StellarScript HT+ exhibit high processivity at 42°C. StellarScript HT+ exhibited the highest inhibitor tolerance, processivity and thermostability at temperatures up to 65°C thus making StellarScript HT+ well suited to produce high yields with clinical samples. Taq Pol variants were identified that efficiently amplify DNA in the presence of inhibitors. Conclusions Based on the increased polymerase activity and inhibitor tolerance, WMG Taq variants are well suited to give robust DNA yields with clinical samples. We were successful in engineering RT and Taq DNAP variants that are highly thermostable, active and inhibitor tolerant and thus will enable breakthroughs in existing POC diagnostics.

Solid Contact Microfabricated Electrode for Point-of-Care Electrochemical Determination of a Parasitic Infection Managing Medication Based on a Transducer Layer Prussian Blue Analogue Decorated with Multi-Walled Carbon Nanotubes

Point-of-care diagnostics (POC), often known as on-site testing has evolved as a quick and accurate methodology for analyzing and therapeutic monitoring of drugs. The aim of this study is to provide cost-effective, simple, portable, reliable, and ecofriendly methodology for determination of Tinidazole (TD); potentiometric sensors are found to be an ideal fit for achieving these goals. An advanced microfabricated ion selective electrode is provided employing two-steps procedure. The first is selecting the most selective ionophore for TD determination while the second is to improve the stability and detection limit upon doping the nanoparticles as ion to electron transducer layer between the microfabricated copper electrode and the ion selective media as it can decrease potential drift from 8.0 to ∼1.0 mV h−1. Nernstian potentiometric results were obtained for TD in range of concentration 1.0 × 10−3 to 1.0 × 10−8 M, its slope was −57.43 mV/decade with lower detection limits 2.55 × 10−9 M. Essential values of the adopted sensor are fast and stable response time (9 ± 2 s). The provided potentiometric approach succeeds to asses TD in its pure and pharmaceutical forms, furthermore in different biological fluids with satisfactory results.

Portable, smartphone-linked, and miniaturized photonic resonator absorption microscope (PRAM Mini) for point-of-care diagnostics.

We report the design, development, and characterization of a miniaturized version of the photonic resonator absorption microscope (PRAM Mini), whose cost, size, and functionality are compatible with point-of-care (POC) diagnostic assay applications. Compared to previously reported versions of the PRAM instrument, the PRAM Mini components are integrated within an optical framework comprised of an acrylic breadboard and plastic alignment fixtures. The instrument incorporates a Raspberry Pi microprocessor and Bluetooth communication circuit board for wireless control and data connection to a linked smartphone. PRAM takes advantage of enhanced optical absorption of ∼80 nm diameter gold nanoparticles (AuNP) whose localized surface plasmon resonance overlaps with the ∼625 nm resonant reflection wavelength of a photonic crystal (PC) surface. When illuminated with wide-field low-intensity collimated light from a ∼617 nm wavelength red LED, each AuNP linked to the PC surface results in locally reduced reflection intensity, which is visualized by observing dark spots in the PC-reflected image with an inexpensive CMOS image sensor. Each AuNP in the image field of view can be easily counted with digital resolution. We report upon the selection of optical/electronic components, image processing algorithm, and contrast achieved for single AuNP detection. The instrument is operated via a wireless connection to a linked mobile device using a custom-developed software application that runs on an Android smartphone. As a representative POC application, we used the PRAM Mini as the detection instrument for an assay that measures the presence of antibodies against SARS-CoV-2 infection in cat serum samples, where each dark spot in the image represents a complex between one immobilized viral antigen, one antibody molecule, and one AuNP tag. With dimensions of 23 × 21 × 10 cm3, the PRAM Mini offers a compact detection instrument for POC diagnostics.

The Development of a Versatile Activation Protocol for Carbon-Infused Thermoplastic-Based 3D Printed Electrochemical Sensors

Point-of-care (POC) diagnostics is rapidly expanding as researchers continually develop advanced sensors and integrated devices with the goal of streamlining on-site detection and analysis. Recently, the utilization of 3D printing for rapid fabrication of housings, electrodes, and sensors for use at the POC has been realized and exploited towards diverse applications. Particularly interesting is the use of 3D printed carbon electrodes (3DpCEs) in POC diagnostics owing to its simplicity, affordability, and mass production capabilities for developing selective and sensitive sensors either for direct use or through post-printing surface modifications. Herein, a comprehensive investigation of an alkaline-assisted electrochemical activation (AAEA) protocol for the fabrication of sensitive and selective 3DpCEs was completed. In this study we explored the influence and effect of basicity, activation cycles, activation time, and scan rate. Following the optimization of the activated 3DpCEs, a comparative study was performed using nonactivated and alkaline immersed 3DpCEs, where AAEA 3DpCEs demonstrated higher current responses, improved reversibility, and significantly increased reproducibility. The AAEA protocol for 3DpCEs also provides a versatile modification protocol for the design of sensors capable of selectively detecting biomolecules in the presence of potential interferents. Using the developed protocol, the AAEA 3DpCEs were applied toward i) the selective separation and sensing of catechol and hydroquinone and ii) the detection of glucose using copper deposited 3DpCEs.

Temperature-programmed microfluidic CRISPR diagnostics enable rapid and automatous point-of-care testing for syphilis

The rising incidence of syphilis highlights the urgent need to develop molecular tests of Treponema pallidum at the point-of-care (POC). Traditional molecular approaches based on polymerase chain reaction (PCR) often involve expensive equipment, expert technicians, and lengthy turnaround time. Recently emerged approaches coupling CRISPR detection with isothermal amplification techniques hold great promise to advance the POC diagnostics, but commonly require multiple manual operations for postamplification reaction transfer or suffer from compromised performance in terms of sensitivity or assay time. To address these issues, we develop a thermometer-inspired microfluidic biosensing platform that seamlessly integrates CRISPR detection and isothermal amplification through temperature-programmed air expansion for intended fluidic manipulation. This platform not only allows for automated running of the canonical two-step recombinase polymerase amplification-CRISPR/Cas12a assay via a simple two-step temperature profile, but also provides a smartphone-enabled fluorescence readout with a limit of detection of a few DNA copies in 30 min. We detail the rational design of a thermally-responsive microfluidic cartridge and showcase how it harnesses the temperature difference in preamplification and CRISPR detection to implement the assay workflow without the need for an external driving mechanism or human intervention. Furthermore, we validate its applicability for rapid and sensitive detection of Treponema pallidum DNA in clinical samples (n = 30), exhibiting 100 % sensitivity and 100 % specificity compared to the quantitative PCR technique. The simplicity and effectiveness may lead this platform to be a powerful upgrade kit compatible with the existing CRISPR approaches, thus facilitating their widespread point-of-care diagnostic applications.

Biosensors–The Point of Care Diagnostics (POCD): A Review

Biosensors–The Point of Care Diagnostics (POCD): A Review

Antibody-Driven Assembly of Plasmonic Core-Satellites to Increase the Sensitivity of a SERS Vertical Flow Immunoassay.

Here, we describe a SERS-based vertical flow assay as a platform technology suitable for point-of-care (POC) diagnostic testing. A capture substrate is constructed from filter paper embedded with spherical gold nanoparticles (AuNPs) and functionalized with an appropriate capture antibody. The capture substrate is loaded into a filtration device and connected to a syringe to rapidly and repeatedly pass the sample through the sensor for efficient antigen binding. The antigen is then labeled with a SERS-active detection probe. We show that only a few Raman reporter molecules, exclusively located adjacent to the plasmonic capture substrate, generate detectible signals. To maximize the signal from underutilized Raman reporter molecules, we employ a secondary signal enhancing probe that undergoes antibody-directed assembly to form plasmonic core-satellites. This facile enhancement step provides a 3.5-fold increase in the signal and a detection limit of 0.23 ng/mL (1.6 pM) for human IgG. This work highlights the potential to rationally design plasmonic architectures using widely available and reproducible spherical AuNPs to achieve large SERS enhancements for highly sensitive POC diagnostics.

Filtered Saliva for Rapid and Accurate Analyte Detection for POC Diagnostics.

Saliva has shown considerable promise as a diagnostic medium for point-of-care (POC) and over-the-counter (OTC) diagnostic devices due to the non-invasive nature of its collection. However, a significant limitation of saliva-based detection is undesirable interference in a sensor's readout caused by interfering components in saliva. In this study, we develop standardized sample treatment procedures to eliminate bubbles and interfering molecules while preserving the sample's target molecules such as spike (S) protein and glucose. We then test the compatibility of the pretreatment system with our previously designed SARS-CoV-2 and glucose diagnostic biosensing systems for detecting S protein and glucose in subject saliva. Ultimately, the effectiveness of each filter in enhancing biomarker sensitivity is assessed. The results show that a 20 mg nylon wool (NW) filter shows an 80% change in viscosity reduction with only a 6% reduction in protein content, making it an appropriate filter for the salivary S protein diagnostic system. Meanwhile, a 30 mg cotton wool (CW) filter is identified as the optimal choice for salivary glucose detection, achieving a 90% change in viscosity reduction and a 60.7% reduction in protein content with a minimal 4.3% reduction in glucose content. The NW pretreatment filtration significantly improves the limit of detection (LOD) for salivary S protein detection by five times (from 0.5 nM to 0.1 nM) and it reduces the relative standard deviation (RSD) two times compared to unfiltered saliva. Conversely, the CW filter used for salivary glucose detection demonstrated improved linearity with an R2 of 0.99 and a sensitivity of 36.6 μA/mM·cm2, over twice as high as unfiltered saliva. This unique filtration process can be extended to any POC diagnostic system and optimized for any biomarker detection, making electrochemical POC diagnostics more viable in the current market.

Diagnostic and Management Challenges of Acute Fatty Liver of Pregnancy: A Case Series

Introduction: Acute fatty liver of pregnancy remains an elusive and potentially fatal complication of a rare liver disease in late pregnancy. Due to sparse research, diagnostic dilemmas, and multi-organ involvement of the disease, it is imperative to elaborate the evaluation and management principles to confine the outcomes. Case Reports: Here, we aim to discuss in detail the peripartum manifestations and goal-directed management protocols in a case series of four patients managed at a tertiary care setup in India. Conclusion: It was found that all four cases had varied presentations ranging from asymptomatic hypoglycemia to liver failure and multiorgan dysfunction requiring a multidisciplinary perioperative care approach to prevent complications by incorporating supportive care along with latest available point of care diagnostics, interventional care, Extracorporeal Membrane Oxygenation (ECMO) and Continuous renal replacement therapy (CRRT).

Point-of-Care Diagnostic System for Viable Salmonella Species via Improved Propidium Monoazide and Recombinase Polymerase Amplification Based Nucleic Acid Lateral Flow.

Salmonella species are prominent foodborne microbial pathogens transmitted through contaminated food or water and pose a significant threat to human health. Accurate and rapid point-of-care (POC) diagnosis is gaining attention in effectively preventing outbreaks of foodborne disease. However, the presence of dead bacteria can interfere with an accurate diagnosis, necessitating the development of methods for the rapid, simple, and efficient detection of viable bacteria only. Herein, we used an improved propidium monoazide (PMAxx) to develop a nucleic acid lateral flow (NALF) assay based on recombinase polymerase amplification (RPA) to differentiate viable Salmonella Typhimurium. We selected an RPA primer set targeting the invA gene and designed a probe for NALF. RPA-based NALF was optimized for temperature (30-43 °C), time (1-25 min), and endonuclease IV concentration (0.025-0.15 unit/µL). PMAxx successfully eliminated false-positive results from dead S. Typhimurium, enabling the accurate detection of viable S. Typhimurium with a detection limit of 1.11 × 102 CFU/mL in pure culture. The developed method was evaluated with spiked raw chicken breast and milk with analysis completed within 25 min at 39 °C. This study has potential as a tool for the POC diagnostics of viable foodborne pathogens with high specificity, sensitivity, rapidity, and cost-effectiveness.

Nonenzymatic Detection of Glucose Using 3D Printed Carbon Electrodes in Human Saliva

One of the most prevalent diseases where point-of-care (POC) diagnostics has focused is diabetes, which impacts hundreds of millions of people globally. Due to the severe negative outcomes including renal failure, nerve damage, and stroke, many POC sensors have been designed to streamline low-cost testing. Recently, the utility of 3D printing for rapidly fabricating housings, electrodes, and sensors for use at the POC has been exploited toward diverse applications. Particularly interesting are 3D printed carbon electrodes (3DpCEs) in POC diagnostics owing to their simplicity, affordability, and mass production capabilities for developing sensors either for direct use or through post-printing surface modifications. Herein, we report a copper modified 3DpCE as a sensitive and selective nonenzymatic biosensor for glucose. Copper deposition, paired with an optimized activation protocol, produced a sensitive and selective sensor for glucose with a larger detection range, enhanced sensitivity, and better reproducibility compared to nonactivated and alkaline immersed 3DpCEs. The sensor displayed excellent linearity between 10–1800 μM and proved to be highly selective over common biologically relevant interferants. The 3D printed sensor successfully determined biologically relevant concentrations of glucose in human saliva which resulted in percent recoveries of 101 ± 8%, 106 ± 6%, and 98 ± 6% for 74, 402, and 652 μM glucose, respectively.

Gold Nanoparticle-based Biosensors for Point-of-Care Diagnostics: A Review of Sensing Nanoparticle Applications and Future Prospects.

Point of Care Diagnostics (POCD) is quintessential in hospitals and the healthcare sector as the secants uplift the quality of medical care and the life of a patient by facilitating quick identification of the underlying pathological condition. Nanotechnology can provide opportunities and has potential in the development of new-age sensing/diagnostic tools. Owing to extraordinary features (e.g., higher density, effective catalysis, good conduction, biocompatibility, inertness, and greater surface-to-volume ratio), gold nanoparticles (AuNPs) are frequently employed in POCT (Point-of-Care-Testing). Gold nanoparticles-based colorimetric methods are widely used in the rapid, sensitive, and selective detection of analytes/target molecules. AuNPs description is critical for their possible utility in prophylaxis, diagnostics, and treatment of an ailment. AuNPs interact with organic/inorganic target molecules to generate colorimetric shift that enables the accurate, precise, and subtle recognition of biologicals (e.g., microorganisms, cellular components, and proteins) and metal ions. This review focused on the need for AuNPs-based colorimetric application in prophylaxis, diagnostics, and treatment in healthcare and reviewed the future outlook of these AuNPs for biological applications. Different synthesis methods of AuNPs, their morphology, and characterization, including their surface functionalization, will be discussed in detail. AuNPs are very much preferable nanomaterials owing to exclusive optical, electrical, and photothermal features. AuNPs-based colorimetric biosensors are simple and possess great utility, yet these offer a robust technique to enable visual, quantitative analysis.

Performance evaluation of a Test&Treat rapid detection kit for the diagnosis of septic effusions in dogs and cats.

To evaluate the sensitivity and specificity of a veterinary point-of-care (POC) luminometer-based kit for the diagnosis of septic peritoneal or pleural effusion in dogs and cats. Prospective study performed between January 2020 and July 2021. University teaching hospital. Forty-eight animals with naturally occurring peritoneal or pleural effusion collected by aseptic abdominocentesis or thoracocentesis. Effusion samples were split into filtered (using a 10-micron filter) and unfiltered aliquots and analyzed by the POC instrument according to the manufacturer's instructions and following variable incubation periods. Samples were also plated aerobically on standard and blood agar plates. Proprietary reagents were added to samples, causing bacterial ATP to generate bioluminescence that is detected by the luminometer. Bioluminescence values (relative light units [RLUs]) were recorded and compared with the presence of bacterial growth on the culture plates. Nucleated cell counts in native and filtered effusion samples were recorded. Twenty-one samples were septic based on positive culture. RLUs were higher in septic effusions for filtered and native effusions compared with sterile effusions. The use of a filter reduced cell counts. In filtered samples incubated for 30minutes before testing, the sensitivity and specificity of the luminometer for diagnosis of infection in cavitary effusions were 81% and 82%, respectively, using a cutoff of 12,202 RLUs. The luminometer kit evaluated in this study represents a viable screening tool for diagnosis of septic cavitary effusions and could be used in conjunction with other POC diagnostics to support the rapid diagnosis of infection.

Microneedle Sensors for Point-of-Care Diagnostics.

Point-of-care (POC) has the capacity to support low-cost, accurate and real-time actionable diagnostic data. Microneedle sensors have received considerable attention as an emerging technique to evolve blood-based diagnostics owing to their direct and painless access to a rich source of biomarkers from interstitial fluid. This review systematically summarizes the recent innovations in microneedle sensors with a particular focus on their utility in POC diagnostics and personalized medicine. The integration of various sensing techniques, mostly electrochemical and optical sensing, has been established in diverse architectures of "lab-on-a-microneedle" platforms. Microneedle sensors with tailored geometries, mechanical flexibility, and biocompatibility are constructed with a variety of materials and fabrication methods. Microneedles categorized into four types: metals, inorganics, polymers, and hydrogels, have been elaborated with state-of-the-art bioengineering strategies for minimally invasive, continuous, and multiplexed sensing. Microneedle sensors have been employed to detect a wide range of biomarkers from electrolytes, metabolites, polysaccharides, nucleic acids, proteins to drugs. Insightful perspectives are outlined from biofluid, microneedles, biosensors, POC devices, and theragnostic instruments, which depict a bright future of the upcoming personalized and intelligent health management.

Distance-based paper analytical device for multiplexed quantification of cytokine biomarkers using carbon dots integrated with molecularly imprinted polymer.

This article introduces distance-based paper analytical devices (dPADs) integrated with molecularly imprinted polymers (MIPs) and carbon dots (CDs) for simultaneous quantification of cytokine biomarkers, namely C-reactive protein (CRP), tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6) in human biological samples for diagnosis of cytokine syndrome. Using fluorescent CDs and MIP technology, the dPAD exhibits high selectivity and sensitivity. Detection is based on fluorescence quenching of CDs achieved through the interaction of the target analytes with the MIP layer on the paper substrate. Quantitative analysis is easily accomplished by measuring the distance length of quenched fluorescence with a traditional ruler and naked eye readout enabling rapid diagnosis of cytokine syndrome and the underlying infection. Our sensor demonstrated linear ranges of 2.50-24.0 pg mL-1 (R2 = 0.9974), 0.25-3.20 pg mL-1 (R2 = 0.9985), and 1.50-16.0 pg mL-1 (R2 = 0.9966) with detection limits (LODs) of 2.50, 0.25, and 1.50 pg mL-1 for CRP, TNF-α, and IL-6, respectively. This sensor also demonstrated remarkable selectivity compared to a sensor employing a non-imprinted polymer (NIP), and precision with the highest relative standard deviation (RSD) of 5.14%. The sensor is more accessible compared to prior methods relying on expensive reagents and instruments and complex fabrication methods. Furthermore, the assay provided notable accuracy for monitoring these biomarkers in various human samples with recovery percentages ranging between 99.22% and 103.58%. By integrating microfluidic systems, nanosensing, and MIPs technology, our developed dPADs hold significant potential as a cost-effective and user-friendly analytical method for point-of-care diagnostics (POC) of cytokine-related disorders. This concept can be further extended to developing diagnostic devices for other biomarkers.

Sample preparation and detection methods in point-of-care devices towards future at-home testing.

Timely and accurate diagnosis is critical for effective healthcare, yet nearly half the global population lacks access to basic diagnostics. Point-of-care (POC) testing offers partial solutions by enabling low-cost, rapid diagnosis at the patient's location. At-home POC devices have the potential to advance preventive care and early disease detection. Nevertheless, effective sample preparation and detection methods are essential for accurate results. This review surveys recent advances in sample preparation and detection methods at POC. The goal is to provide an in-depth understanding of how these technologies can enhance at-home POC devices. Lateral flow assays, nucleic acid tests, and virus detection methods are at the forefront of POC diagnostic technology, offering rapid and sensitive tools for identifying and measuring pathogens, biomarkers, and viral infections. By illuminating cutting-edge research on assay development for POC diagnostics, this review aims to accelerate progress towards widely available, user-friendly, at-home health monitoring tools that empower individuals in personalized healthcare in the future.

SERS-based microdevices for use as in vitro diagnostic biosensors.

Advances in surface-enhanced Raman scattering (SERS) detection have helped to overcome the limitations of traditional in vitro diagnostic methods, such as fluorescence and chemiluminescence, owing to its high sensitivity and multiplex detection capability. However, for the implementation of SERS detection technology in disease diagnosis, a SERS-based assay platform capable of analyzing clinical samples is essential. Moreover, infectious diseases like COVID-19 require the development of point-of-care (POC) diagnostic technologies that can rapidly and accurately determine infection status. As an effective assay platform, SERS-based bioassays utilize SERS nanotags labeled with protein or DNA receptors on Au or Ag nanoparticles, serving as highly sensitive optical probes. Additionally, a microdevice is necessary as an interface between the target biomolecules and SERS nanotags. This review aims to introduce various microdevices developed for SERS detection, available for POC diagnostics, including LFA strips, microfluidic chips, and microarray chips. Furthermore, the article presents research findings reported in the last 20 years for the SERS-based bioassay of various diseases, such as cancer, cardiovascular diseases, and infectious diseases. Finally, the prospects of SERS bioassays are discussed concerning the integration of SERS-based microdevices and portable Raman readers into POC systems, along with the utilization of artificial intelligence technology.

Vibration mixing for enhanced paper-based recombinase polymerase amplification.

Isothermal nucleic acid amplification tests (NAATs) are a vital tool for point-of-care (POC) diagnostics. These assays are well-suited for rapid, low-cost POC diagnostics for infectious diseases compared to traditional PCR tests conducted in central laboratories. There has been significant development of POC NAATs using paper-based diagnostic devices because they provide an affordable, user-friendly, and easy to store format; however, the difficulties in integrating separate liquid components, resuspending dried reagents, and achieving a low limit of detection hinder their use in commercial applications. Several studies report low assay efficiencies, poor detection output, and poorer limits of detection in porous membranes compared to traditional tube-based protocols. Recombinase polymerase amplification is a rapid, isothermal NAAT that is highly suited for POC applications, but requires viscous reaction conditions that has poor performance when amplifying in a porous paper membrane. In this work, we show that we can dramatically improve the performance of membrane-based recombinase polymerase amplification (RPA) of HIV-1 DNA and viral RNA by employing a coin cell-based vibration mixing platform. We achieve a limit of detection of 12 copies of DNA per reaction, nearly 50% reduction in time to threshold (from ∼10 minutes to ∼5 minutes), and an overall fluorescence output increase up to 16-fold when compared to unmixed experiments. This active mixing strategy enables reactions where the target and reaction cofactors are isolated from each other prior to the reaction. We also demonstrate amplification using a low-cost vibration motor for both temperature control and mixing, without the requirement of any additional heating components.

Point-of-care devices engaging green graphene: an eco-conscious and sustainable paradigm.

The healthcare landscape has experienced a profound and irreversible transformation, primarily driven by the emergence of nanomaterial-assisted point-of-care (POC) devices. The inclusion of nanomaterials in POC devices has revolutionized healthcare by enabling rapid, on-site diagnostics with minimal infrastructure requirements. Among the materials poised to lead this technological revolution, green graphene emerges as a compelling contender. It possesses a unique combination of exceptional material properties and environmentally conscious attributes. These attributes include its substantial surface area, unparalleled electrical conductivity, and inherent biocompatibility. This article embarks on an exploration of POC devices incorporating green graphene. It meticulously dissects the intricacies of their design, performance characteristics, and diverse applications. Throughout the exposition, the transformative impact of green graphene on the advancement of POC diagnostics takes centre stage. It underscores the material's potential to drive sustainable and effective healthcare solutions, marking a significant milestone in the evolution of healthcare technology.