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Helicobacter pylori (Hp) prevail globally as the primary cause of gastritis, gastric ulcer, and potential gastric cancer, highlighting the need for rapid and precise point-of-care (POC) detection of Hp nucleic acid. Upconversion nanoparticle-based lateral flow assay (UCNPs-LFA) exhibit great potential in POC detection, due to their high optical stability and absence of background fluorescence. However, insufficient sensitivity for nucleic acid detection remains a key challenge. This study systematically optimizes UCNPs-LFA by focusing on target capture, signal transduction, signal separation, and signal analysis, to enhance its detection capabilities for Hp nucleic acid. The optimized UCNPs-LFA platform features a significantly decreased detection limit, a broadened detection range, and high reliability. Results demonstrate that the limit of detection (LOD) is 25 fM, a 105-fold improvement over the initial platform. This systematic optimization strategy is versatile and can be applied to optimize other nanoparticle-based LFAs. [ABSTRACT FROM AUTHOR]

Lateral Flow Assays (LFAs) are cost‐effective and widely utilized for rapid diagnostics, yet they often suffer from limited sensitivity. This study introduces a straightforward yet highly effective method to enhance LFAs performance by integrating a poly(ethylene glycol) diacrylate (PEGDA) hydrogel to create a "reaction trap." This hydrogel reaction trap optimizes the flow rate and reaction time at the sensing components, substantially improving assay performance. By applying various hydrogel concentrations (6%, 9%, 12%, 15%, and 18% w/v), significant enhancements across a range of detection systems are achieved. An optimized 18% hydrogel concentration shows a 1.5 times increase in sensitivity in the tested commercial pregnancy test. Additionally, 12% hydrogel concentration is tested in pregnancy, ovulation, and Coronavirus disease 2019 (COVID‐19) commercial kits, and the improvement reached up to a sevenfold increase in signal intensity. The enhancement in detection illustrates the profound impact of this simple modification and shows the importance of hydrogel concentration optimization to maximize detection improvement. These results demonstrate that hydrogel‐coated LFAs offer a scalable and highly effective solution for boosting the reliability and sensitivity of rapid diagnostics across different healthcare settings, with broad potential for global health diagnostics applications. [ABSTRACT FROM AUTHOR]

Nipah and Hendra viruses belong to the Paramyxoviridae family, which pose a significant threat to human health, with sporadic outbreaks causing severe morbidity and mortality. Early symptoms include fever, cough, sore throat, and headache, which offer little in terms of differential diagnosis. There are no specific therapeutics and vaccines for these viruses. This review comprehensively covers a spectrum of diagnostic techniques for Nipah and Hendra virus infections, discussed in conjunction with appropriate type of samples during the progression of infection. Serological assays, reverse transcriptase Real-Time PCR assays, and isothermal amplification assays are discussed in detail, along with a listing of few commercially available detection kits. Patents protecting inventions in Nipah and Hendra virus detection are also covered. Despite several outbreaks of Nipah and Hendra infections in the past decade, in-depth research into their pathogenesis, Point-of-Care diagnostics, specific therapies, and human vaccines is lacking. A prompt and accurate diagnosis is pivotal for efficient outbreak management, patient treatment, and the adoption of preventative measures. The emergence of rapid point-of-care tests holds promise in enhancing diagnostic capabilities in real-world settings. The patent landscape emphasizes the importance of innovation and collaboration within the legal and business realms. [ABSTRACT FROM AUTHOR]

Abstract Lateral Flow Assays (LFAs) are cost‐effective and widely utilized for rapid diagnostics, yet they often suffer from limited sensitivity. This study introduces a straightforward yet highly effective method to enhance LFAs performance by integrating a poly(ethylene glycol) diacrylate (PEGDA) hydrogel to create a “reaction trap.” This hydrogel reaction trap optimizes the flow rate and reaction time at the sensing components, substantially improving assay performance. By applying various hydrogel concentrations (6%, 9%, 12%, 15%, and 18% w/v), significant enhancements across a range of detection systems are achieved. An optimized 18% hydrogel concentration shows a 1.5 times increase in sensitivity in the tested commercial pregnancy test. Additionally, 12% hydrogel concentration is tested in pregnancy, ovulation, and Coronavirus disease 2019 (COVID‐19) commercial kits, and the improvement reached up to a sevenfold increase in signal intensity. The enhancement in detection illustrates the profound impact of this simple modification and shows the importance of hydrogel concentration optimization to maximize detection improvement. These results demonstrate that hydrogel‐coated LFAs offer a scalable and highly effective solution for boosting the reliability and sensitivity of rapid diagnostics across different healthcare settings, with broad potential for global health diagnostics applications.

In the quest for more effective colorimetric reporters compared with traditional gold nanoparticles (AuNPs), a family of Au@Ag core–shell nanoparticles was designed and synthesized using a seed growth-mediated approach starting from commercial 37 nm AuNPs. This method enabled precise control over the thickness of the silver shell by employing hydroquinone for the reduction of silver and citrate for stabilization of the resulting core–shell particles. Core–shell NPs with an Ag shell of 7 nm (Au@Ag5NPs) and 18 nm (Au@Ag10NPs) were synthesized, resulting in orange and milky yellow suspension, respectively. Additionally, the impact of an external gold layer on Au@Ag10NPs (Au@Ag10@AuNPs), which significantly altered their optical properties from milky yellow to gray, was investigated. The core–shell Au@AgNPs exhibited substantially higher molar extinction coefficients than their parent AuNPs: from 3.5-fold for Au@Ag5NPs and 9-fold for Au@Ag10NPs and Au@Ag10@AuNPs. Subsequently, all core–shell NPs were functionalized with a calix[4]-arene layer, imparting superior stability against external stresses, such as dispersion in PBS, when compared to NPs functionalized with traditional ligands. This calixarene coating enabled the covalent bioconjugation of antibodies on all NP types without inducing noticeable aggregation. Their performance as colorimetric reporters was evaluated in a lateral flow assay for troponin I detection, demonstrating positive signals down to 1 ng/mL, surpassing the detection limit of the parent gold NPs (2.5 ng/mL). Notably, the gray color of the core–shell Au@Ag10@AuNPs provided strong contrast against the white NC membrane, facilitating T-line visualization even at low signal intensity. Despite the lack of optimization of our LFA, it competes with the limit of quantification of commercial LFAs for troponin I detection, offering the potential for the development of a highly sensitive assay. The diverse core–shell NPs employed in this study form a library of colorimetric reporters with distinct optical properties, paving the way for multiplexed detection systems targeting multiple proteins simultaneously and enhancing diagnostic reliability. Furthermore, the choice of colorimetric reporters allows tailoring the detection range based on the pertinent limit of quantification desired for the analyte, dictated by the reporter's light extinction properties. [ABSTRACT FROM AUTHOR]

FSH, estrogen and progesterone testing are widely utilized in clinical practice. Lateral flow assays (LFAs) are cost-effective tools used for diagnosing infectious diseases, pregnancy, and substance testing. The focus of this narrative review is the potential for the wider utilization of listed hormone LFAs. A search was conducted with PubMed, Google Scholar and Wiley online libraries using keywords without any limitation on the publication date; animal studies were excluded. Clinical guidelines for the related conditions were included. According to published data, E3G and PdG are used to determine ovulatory cycles and can be utilized for research purposes to establish the normal range of menstrual cycles, as there is currently disagreement among guidelines. FSH measurement in blood samples is utilized to predict oocyte yield in assisted cycles and to differentiate women with premature ovarian insufficiency from hypothalamic amenorrhea, and can be replaced with more convenient urine testing. PdG was tested to assess the risk of pregnancy complications, specifically miscarriage and ectopic pregnancy, and might become a screening tool for miscarriage in the future. PMS, PMDD and ovarian carcinogenesis could be extensively studied using LFAs to gain a better understanding of the biology behind these conditions. Before implementing these LFAs into clinical practice, the reproducibility of progesterone assays should be evaluated. The results are critical for treatment decisions, and universally recognized standards for estradiol measurement should be developed. [ABSTRACT FROM AUTHOR]

The combination of upconverting nanoparticles (UCNPs) and immunochromatography has become a widely used and promising new detection technique for point-of-care testing (POCT). However, their low luminescence efficiency, non-specific adsorption, and image noise have always limited their progress toward practical applications. Recently, artificial intelligence (AI) has demonstrated powerful representational learning and generalization capabilities in computer vision. We report for the first time a combination of AI and upconversion nanoparticle-based lateral flow assays (UCNP-LFAs) for the quantitative detection of commercial internet of things (IoT) devices. This universal UCNPs quantitative detection strategy combines high accuracy, sensitivity, and applicability in the field detection environment. By using transfer learning to train AI models in a small self-built database, we not only significantly improved the accuracy and robustness of quantitative detection, but also efficiently solved the actual problems of data scarcity and low computing power of POCT equipment. Then, the trained AI model was deployed in IoT devices, whereby the detection process does not require detailed data preprocessing to achieve real-time inference of quantitative results. We validated the quantitative detection of two detectors using eight transfer learning models on a small dataset. The AI quickly provided ultra-high accuracy prediction results (some models could reach 100% accuracy) even when strong noise was added. Simultaneously, the high flexibility of this strategy promises to be a general quantitative detection method for optical biosensors. We believe that this strategy and device have a scientific significance in revolutionizing the existing POCT technology landscape and providing excellent commercial value in the in vitro diagnostics (IVD) industry.

Here we develop Lateral Flow Assays (LFAs) that employ as functional elements DNA‐based structures decorated with reporter tags and recognition elements. We have rationally re‐engineered tile‐based DNA tubular structures that can act as scaffolds and can be decorated with recognition elements of different nature (i.e. antigens, aptamers or proteins) and with orthogonal fluorescent dyes. As a proof‐of‐principle we have developed sandwich and competitive multiplex lateral flow platforms for the detection of several targets, ranging from small molecules (digoxigenin, Dig and dinitrophenol, DNP), to antibodies (Anti‐Dig, Anti‐DNP and Anti‐MUC1/EGFR bispecific antibodies) and proteins (thrombin). Coupling the advantages of functional DNA‐based scaffolds together with the simplicity of LFAs, our approach offers the opportunity to detect a wide range of targets with nanomolar sensitivity and high specificity. [ABSTRACT FROM AUTHOR]

Mycobacterium avium subsp. paratuberculosis (MAP) causes bovine paratuberculosis (PTB). PTB is responsible for significant economic losses in dairy herds around the word. PTB control programs that rely on testing and culling of test-positive cows have been developed. Current diagnostics, such as ELISA for detecting MAP antibodies in serum samples and PCR detecting MAP DNA in feces, have inadequate sensitivity for detecting subclinical animals. Innovative “omics” technologies such as next-generation sequencing (NGS) technology-based RNA-sequencing (RNA-Seq), proteomics and metabolomics can be used to find host biomarkers. The discovered biomarkers (RNA, microRNAs, proteins, metabolites) can then be used to develop new and more sensitive approaches for PTB diagnosis. Traditional approaches for measuring host antibodies and biomarkers, such as ELISAs, northern blotting, quantitative reverse-transcriptase polymerase chain reaction (RT-qPCR), cDNA microarrays, and mass spectrometry are time-consuming, expensive, and sometimes exhibit poor sensitivity. With the rapid development of nanotechnology, low-cost monitoring devices for measuring antibodies against MAP proteins in point-of-care (POC) settings have been developed. Lateral flow assays (LFAs), in particular, are thought to be appropriate for the on-site detection of antibodies to MAP antigens and/or host biomarkers. This review aims to summarize LFAs that have recently been developed to accurately detect antibodies against MAP antigens, as well as the benefits that host biomarkers linked with MAP infection give to PTB diagnosis. The identification of these novel biomarkers could be the basis for the development of new LFAs. The dairy industry and producers are likely to benefit from reliable and rapid technologies capable of detecting MAP infection in situ to establish a quick and sensitive PTB diagnosis.

Already for some decades lateral flow assays (LFAs) are 'common use' devices in our daily life. Also, for forensic use LFAs are developed, such as for the analysis of illicit drugs and DNA, but also for the detection of explosives and body fluid identification. Despite their advantages, including ease-of-use, LFAs are not yet frequently applied at a crime scene. This review describes (academic) developments of LFAs for forensic applications, focusing on biological and chemical applications, whereby the main advantages and disadvantages of LFAs for the different forensic applications are summarized. Additionally, a critical review is provided, discussing why LFAs are not frequently applied within the forensic field and highlighting the steps that are needed to bring LFAs to the forensic market. [ABSTRACT FROM AUTHOR]

Nitrocellulose (NC) membrane was fabricated and tested for its potential use in various paper-based biosensors for use in point-of-care testing. However, contemporary technologies are complex, expensive, non-scalable, limited by conditions, and beset with potentially adverse effects on the environment. Herein, we proposed a simple, cost-effective, scalable technology to prepare nitrocellulose/cotton fiber (NC/CF) composite membranes. The NC/CF composite membranes with a diameter of 20 cm were fabricated in 15 min using papermaking technology, which contributes to scalability in the large-scale production of these composites. Compared with existing commercial NC membranes, the NC/CF composite membrane is characterized by small pore size (3.59 ± 0.19 μm), low flow rate (156 ± 55 s/40 mm), high dry strength (up to 4.04 MPa), and wet strength (up to 0.13 MPa), adjustable hydrophilic-hydrophobic (contact angles ranged from 29 ± 4.6 to 82.8 ± 2.4°), the good adsorption capacity of protein (up to 91.92 ± 0.07 μg). After lateral flow assays (LFAs) detection, the limit of detection is 1 nM, which is similar to commercial NC membrane (Sartorius CN 140). We envision the NC/CF composite membrane as a promising material for paper-based biosensors of point-of-care testing applications. [ABSTRACT FROM AUTHOR]

Purpose of Review: The true incidence of fungal disease is hampered by conventionally poor diagnostic tests, limited access to advanced diagnostics, and limited surveillance. The availability of serological testing has been available for over two decades and generally underpins the modern diagnosis of the most common forms of fungal disease. This review will focus on technical developments of serological tests for the diagnosis of fungal disease, describing advances in clinical performance when available. Recent Findings: Despite their longevity, technical, clinical, and performance limitations remain, and tests specific for fungal pathogens outside the main pathogens are lacking. The availability of LFA and automated systems, capable of running multiple different tests, represents significant developments, but clinical performance data is variable and limited. Summary: Fungal serology has significantly advanced the diagnosis of the main fungal infections, with LFA availability increasing accessibility to testing. Combination testing has the potential to overcome performance limitations. [ABSTRACT FROM AUTHOR]

Lateral Flow Assays (LFA) have been one of the most widely adopted technologies in clinical diagnosis over recent years, especially during the COVID-19 pandemic, due to their feasibility, compactness, and rapid readout. However, the precise dispensing of antibodies–a key part of the fabrication process–requires costly line dispenser equipment, which poses a challenge to researchers with limited budgets. This study aims to resolve this key issue by introducing a Syringe-based Arduino-operated Low-cost Antibody Dispenser (SALAD). By utilizing a microneedle, stepper motor-driven syringe pump, and conveyor belt, SALAD can form micro-droplets to create an even band of antibodies. Our evaluation results showed comparable performance between SALAD and a commercialized model – Claremont ALFRD, with SALAD exceeding in affordability and feasibility. SALAD yielded an even signal, uniform bandwidth, and low background noise, yet optimization in the conveyor belt should be considered to enhance stability. With a low manufacturing cost ($200.61) compared to the commercialized models, our model is expected to provide an affordable approach for LFA researchers.

Background: Dengue, ranked as one of the most critical viral diseases, requires rapid, accurate, and early diagnosis for better patient care. Objectives: This study pertains to the development and clinical validation of an in house easy to do, affordable kit for the detection of dengue NS1 antigen based on lateral flow immunoassay in serum samples of patients. Study Design: Clinically uncharacterized serum specimens were obtained and analyzed using the developed NS1 detection kit and compared against the ELISA results. Results: The performance of the kit was evaluated with both positive and negative patients’ serum samples for NS1 antigen and found to be highly specific and sensitive. An overall sensitivity of 92.16 % and specificity of 97.25 % were recorded. Kit stability tests were also carried out and the performance of the kit was found to be similar to real time tests. Conclusion: The results indicate that the developed NS1 detection kit has good reliability with comparable performance to ELISA results.

Recently, CRISPR/Cas13a-based biosensors have been combined with non-traditional lateral flow assays (N-TLFAs) successfully used to clinical detection of target RNA due to their simplicity, portability, and cost-effectiveness. However, N-TLFAs used in such biosensors have several disadvantages, such as inversion of test and control zones, in which color intensity depends on reporter probe cleavage. Herein, we propose a novel biosensor in which CRISPR/Cas13a is integrated with traditional lateral flow assays (TLFAs) to facilitate the sequential direct location of test/control zones by dual hairpin probes, namely DNA hairpin reporter probe B-HPs (5′ Biotin-hairpins, B-HPs, loop containing 5 rU) and F-HPs (5′ FAM-hairpins, F-HPs). Cas13a activated by crRNA-targeted RNA duplexes can indiscriminately cleave B-HPs reporter probes to release short sequences to open hairpin F-HPs reporter probes to form Biotin-dsDNA-FAM reporter probes. The advantage of this platform is that detection results only rely on the formed Biotin-dsDNA-FAM reporter probes, which can be used for SARS-CoV-2 RNA detection after preliminary detection. Under optimized conditions, the SARS-CoV-2 RNA detection range was 10 aM to 1 µM and the limit of detection was as low as 5.34 aM. This proof-of-concept study demonstrates that dual-hairpin reporter-probe binding to TLFAs opens up new opportunities for CRISPR/Cas13a activity detection and bioanalytical applications. • The dual hairpin combined with the TLFAs improves the disadvantage of the reverse positioning of the N-TLFAs. • Rapid detection of SARS-CoV-2 RNA without amplification for 35 min by CRISPR/Cas13a combined with TLFAs. • The Cas13a-TLFAs biosensors platform is capable of achieving nanomolar-level detection ranges. [ABSTRACT FROM AUTHOR]

The global coronavirus disease 2019 (COVID-19) pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spawned an ongoing demand for new research reagents and interventions. Herein we describe a panel of monoclonal antibodies raised against SARS-CoV-2. One antibody showed excellent utility for immunohistochemistry, clearly staining infected cells in formalin-fixed and paraffin embedded lungs and brains of mice infected with the original and the omicron variants of SARS-CoV-2. We demonstrate the reactivity to multiple variants of concern using ELISAs and describe the use of the antibodies in indirect immunofluorescence assays, Western blots, and rapid antigen tests. Finally, we illustrate the ability of two antibodies to reduce significantly viral tissue titers in K18-hACE2 transgenic mice infected with the original and an omicron isolate of SARS-CoV-2. [ABSTRACT FROM AUTHOR]

Objective: Respiratory disease is a rapidly growing global health issue that impacts the quality of living of tens of millions of people around the world. Neutrophil elastase (NE) represents a key inflammatory biomarker and has previously been demonstrated to have the capability of predicting exacerbation risk related to respiratory diseases. This paper utilises a low-cost Point of Care (PoC) approach using Lateral Flow Assays (LFAs) to provide quantitative measurement of active NE in a patient’s sputum. Methods and procedures: The main aim of this study is to develop a quantitative platform using a Complementary Metal-Oxide-Semiconductor (CMOS) to image the LFAs and with an adaptable image analysis algorithm to measure a target biomarker concentration. This result could be used to monitor a patient’s health and quality of living. In the paper, NE is used as the target biomarker to determine if the patient is suffering from a high risk of exacerbations. Results: The results presented in the paper indicate the CMOS reader approach is promising for rapid and low-cost PoC devices, with the current system able to provide quantitative trends of NE concentrations as low as 100 ng/ml and is comparable to a research-based laboratory lateral flow reader. Conclusion: The image analysis algorithm used in the CMOS reader can estimate the minimum NE concentration of 250 ng/ml to indicate the high-risk category for exacerbations from respiratory illnesses with the same accuracy as expensive a research-based laboratory reader but by using low-cost components and onboard image analysis. Clinical impact: The image analysis algorithm is evaluated to analyse LFAs with NE biomarker to determine the patient in a high-risk category for exacerbations. The device communicates the analysis result to medical professionals for daily historical logging for daily health monitoring without regular hospital appointments. The low-cost approach of the proposed system and image analysis approach can be adapted to analyse different biomarkers for other health concerns including multiplex LFAs without additional hardware in the reader design.

Globodera rostochiensis is an important quarantine pest, it causes serious potato yield losses annually. Reliable and rapid molecular detection of G. rostochiensis is pivotal to effective early disease diagnosis and managements. Herein, recombinase polymerase amplification integrated with lateral flow assays method (RPA-LFA) was developed to target the internal transcribed spacer of nuclear ribosomal DNA (ITS rDNA) of the golden cyst nematode (G. rostochiensis), which allowed for the rapid diagnosis and detection of this nematode from crude extracts of cysts and juveniles within 30 min. Sensitivity test results showed that 10−1 single juvenile and 10−3 single cyst can be reliably detected. Moreover, the RPA-LFA method can directly diagnose and detect G. rostochiensis from infested field soil. This is the first RPA-LFA method for diagnosis G. rostochiensis, it is a fast, accurate, and sensitive detection method and can be developed for detection of G. rostochiensis in fields and laboratories lacking large instrument and equipment. [ABSTRACT FROM AUTHOR]

Abstract Background COVID-19 has resulted in significant morbidity and mortality worldwide. Lateral flow assays can detect anti-Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) antibodies to monitor transmission. However, standardized evaluation of their accuracy and tools to aid in interpreting results are needed. Methods We evaluated 20 IgG and IgM assays selected from available tests in April 2020. We evaluated the assays’ performance using 56 pre-pandemic negative and 56 SARS-CoV-2-positive plasma samples, collected 10–40 days after symptom onset, confirmed by a molecular test and analyzed by an ultra-sensitive immunoassay. Finally, we developed a user-friendly web app to extrapolate the positive predictive values based on their accuracy and local prevalence. Results Combined IgG + IgM sensitivities ranged from 33.9 to 94.6%, while combined specificities ranged from 92.6 to 100%. The highest sensitivities were detected in Lumiquick for IgG (98.2%), BioHit for both IgM (96.4%), and combined IgG + IgM sensitivity (94.6%). Furthermore, 11 LFAs and 8 LFAs showed perfect specificity for IgG and IgM, respectively, with 15 LFAs showing perfect combined IgG + IgM specificity. Lumiquick had the lowest estimated limit-of-detection (LOD) (0.1 μg/mL), followed by a similar LOD of 1.5 μg/mL for CareHealth, Cellex, KHB, and Vivachek. Conclusion We provide a public resource of the accuracy of select lateral flow assays with potential for home testing. The cost-effectiveness, scalable manufacturing process, and suitability for self-testing makes LFAs an attractive option for monitoring disease prevalence and assessing vaccine responsiveness. Our web tool provides an easy-to-use interface to demonstrate the impact of prevalence and test accuracy on the positive predictive values.

[Display omitted] • Discuss the problems of long PCR times and the non-portability of PCR instruments. • Discuss the pros and cons of antigen or antibody detection and nucleic acid testing. • Introduce the performance of integrated detection platforms and devices. In recent years, the widespread spread of many viruses among populations, e.g., severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), monkeypox, and Middle East respiratory syndrome, has posed a huge threat to human life and health and the global economy. Especially, the coronavirus disease 2019 (COVID-19) outbreak caused by SARS-CoV-2 has been a wake-up call and stimulated the development of rapid detection and point-of-care testing (POCT) field. How to quickly establish POCT methods and rapidly detect pathogens to control the virus epidemic in the population has become a hot research topic. In this review, we first analyzed the pros and cons of different detection methods based on target types, including nucleic acids, antigens, and antibodies. Then we summarized the performances of the integrated detection platforms and devices in combination with these detection methods, such as isothermal amplification, lateral flow assays (LFAs), clustered regularly interspaced short palindromic repeats (CRISPR), electrochemistry, surface-enhanced Raman scattering (SERS) biosensor. It also discusses how to solve the problems of long polymerase chain reaction (PCR) times and the non-portability of PCR instruments. This summary can enhance the comprehension of researchers in rapidly establishing POCT methods, thereby facilitating their industrial application to scientifically respond to the next outbreak of novel or historical pathogens. This endeavor aims to effectively reduce the societal and economic ramifications of new emerging pathogens in epidemics, particularly in countries with large populations or limited medical infrastructure. [ABSTRACT FROM AUTHOR]

Zoonoses and animal diseases threaten human health and livestock biosecurity and productivity. Currently, laboratory confirmation of animal disease outbreaks requires centralized laboratories and trained personnel; it is expensive and time-consuming, and it often does not coincide with the onset or progress of diseases. Point-of-care (POC) diagnostics are rapid, simple, and cost-effective devices and tests, that can be directly applied on field for the detection of animal pathogens. The development of POC diagnostics for use in human medicine has displayed remarkable progress. Nevertheless, animal POC testing has not yet unfolded its full potential. POC devices and tests for animal diseases face many challenges, such as insufficient validation, simplicity, and portability. Emerging technologies and advanced materials are expected to overcome some of these challenges and could popularize animal POC testing. This review aims to: (i) present the main concepts and formats of POC devices and tests, such as lateral flow assays and lab-on-chip devices; (ii) summarize the mode of operation and recent advances in biosensor and POC devices for the detection of farm animal diseases; (iii) present some of the regulatory aspects of POC commercialization in the EU, USA, and Japan; and (iv) summarize the challenges and future perspectives of animal POC testing. [ABSTRACT FROM AUTHOR]

Objective Numerous manufacturers market lateral flow assays for the detection of SARS-CoV-2 antibodies, but there are many questions about the reliability and efficacy of these tests. Materials and Methods Serum specimens from 60 individuals were analyzed using 2 lateral flow antibody assays, an in-house enzyme-linked immunosorbent assay (ELISA), and the Abbott SARS-CoV-2 IgG chemiluminescent immunoassay. Results The BioMedomics and Premier Biotech lateral flow assays were positive for IgM in 73.3% and 70% and for IgG in 80% and 73.3% of specimens, respectively. The ELISA assay was positive for IgM and IgG in 73.3% and 86.7% of specimens from infected individuals, whereas the Abbott assay was positive in 80%. The specificities of the 4 assays ranged from 96.7% to 100% for IgM and from 93.3% to 100% for IgG. Conclusion Results of the 2 lateral flow assays were comparable to those of the ELISA and Abbott assays. Assay efficacy depended on length of time after SARS-CoV-2 infection. [ABSTRACT FROM AUTHOR]

Point-of-care (POC) diagnostics, in particular lateral flow assays (LFA), represent a great opportunity for rapid, precise, low-cost and accessible diagnosis of disease. Especially with the ongoing coronavirus disease 2019 (COVID-19) pandemic, rapid point-of-care tests are becoming everyday tools for identification and prevention. Using smartphones as biosensors can enhance POC devices as portable, low-cost platforms for healthcare and medicine, food and environmental monitoring, improving diagnosis and documentation in remote, low-resource locations. We present an open-source, all-in-one smartphone-based system for quantitative analysis of LFAs. It consists of a 3D-printed photo box, a smartphone for image acquisition, and an R Shiny software package with modular, customizable analysis workflow for image editing, analysis, data extraction, calibration and quantification of the assays. This system is less expensive than commonly used hardware and software, so it could prove very beneficial for diagnostic testing in the context of pandemics, as well as in low-resource countries. [ABSTRACT FROM AUTHOR]

The development of the lateral flow assay (LFA) has received much attention in both academia and industry because of their broad applications to food safety, environmental monitoring, clinical diagnosis, and so forth. The user friendliness, low cost, and easy operation are the most attractive advantages of the LFA. In recent years, quantitative detection has become another focus of LFA development. Here, the most recent studies of quantitative LFAs are reviewed. First, the principles and corresponding formats of quantitative LFAs are introduced. In the biomaterial and nanomaterial sections, the detection, capture, and signal amplification biomolecules and the optical, fluorescent, luminescent, and magnetic labels used in LFAs are described. The invention of dedicated strip readers has drawn further interest in exploiting the better performance of LFAs. Therefore, next, the development of dedicated reader devices is described and the usefulness and specifications of these devices for LFAs are discussed. Finally, the applications of LFAs in the detection of metal ions, biotoxins, pathogenic microorganisms, veterinary drugs, and pesticides in the fields of food safety and environmental health and the detection of nucleic acids, biomarkers, and viruses in clinical analyses are summarized. [ABSTRACT FROM AUTHOR]

ABSTRACT During the last year, mass screening campaigns have been carried out to identify immunological response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and establish a possible seroprevalence. The obtained results gained new importance with the beginning of the SARS-CoV-2 vaccination campaign, as the lack of doses has persuaded several countries to introduce different policies for individuals who had a history of COVID-19. Lateral flow assays (LFAs) may represent an affordable tool to support population screening in low-middle-income countries, where diagnostic tests are lacking and epidemiology is still widely unknown. However, LFAs have demonstrated a wide range of performance, and the question of which one could be more valuable in these settings still remains. We evaluated the performance of 11 LFAs in detecting SARS-CoV-2 infection, analyzing samples collected from 350 subjects. In addition, samples from 57 health care workers collected at 21 to 24 days after the first dose of the Pfizer-BioNTech vaccine were also evaluated. LFAs demonstrated a wide range of specificity (92.31% to 100%) and sensitivity (50% to 100%). The analysis of postvaccination samples was used to describe the most suitable tests to detect IgG response against S protein receptor binding domain (RBD). Tuberculosis (TB) therapy was identified as a potential factor affecting the specificity of LFAs. This analysis identified which LFAs represent a valuable tool not only for the detection of prior SARS-CoV-2 infection but also for the detection of IgG elicited in response to vaccination. These results demonstrated that different LFAs may have different applications and the possible risks of their use in high-TB-burden settings. IMPORTANCE Our study provides a fresh perspective on the possible employment of SARS-CoV-2 LFA antibody tests. We developed an in-depth, large-scale analysis comparing LFA performance to enzyme-linked immunosorbent assay (ELISA) and electrochemiluminescence immunoassay (ECLIA) and evaluating their sensitivity and specificity in identifying COVID-19 patients at different time points from symptom onset. Moreover, for the first time, we analyzed samples of patients undergoing treatment for endemic poverty-related diseases, especially tuberculosis, and we evaluated the impact of this therapy on test specificity in order to assess possible performance in TB high-burden countries.

Research in electrochemical detection in lateral flow assays (LFAs) has gained significant momentum in recent years. The primary impetus for this surge in interest is the pursuit of achieving lower limits of detection, especially given that LFAs are the most widely employed point-of-care biosensors. Conventionally, the strategy for merging electrochemistry and LFAs has centered on the superposition of screen-printed electrodes onto nitrocellulose substrates during LFA fabrication. Nevertheless, this approach poses substantial limitations regarding scalability. In response, we have developed a novel method for the complete integration of reduced graphene oxide (rGO) electrodes into LFA strips. We employed a CO 2 laser to concurrently reduce graphene oxide and pattern nitrocellulose, exposing its backing to create connection sites impervious to sample leakage. Subsequently, rGO and nitrocellulose were juxtaposed and introduced into a roll-to-roll system using a wax printer. The exerted pressure facilitated the transfer of rGO onto the nitrocellulose. We systematically evaluated several electrochemical strategies to harness the synergy between rGO and LFAs. While certain challenges persist, our rGO transfer technology presents compelling potential for setting a new standard in electrochemical LFA fabrication. • Reduced graphene oxide electrodes and lateral flow assay strips fully integrated with a novel methodology. • Streamlined dual-purpose laser solution to reduce graphene oxide, ensure connectivity and impede sample leaking. • Pressure exerted uniformly using a wax printer mechanism to transfer rGO onto nitrocellulose without compromising fluidics. • Exploration of several electrochemical strategies to understand the synergy between rGO and nitrocellulose, a porous matrix. • Proof-of-concept application in an electrochemical LFA for cortisol that still allows optical readouts. [ABSTRACT FROM AUTHOR]

[Display omitted] • Introduction of the use of electrochemical impedance spectroscopy to explore ion transport dynamics in LFA. • Fluid velocity and Péclet number analysis performed for electro-fluidic insights in LFA. • Diffusive and convective ion transport in LFA discovered to be crucial for eLFA designs. • LFA dead volume modification to ensure continuous sample flow during EIS measurement. • Reusable LFA electrodes cartridges fabricated, promoting waste reduction in biosensors. Lateral Flow Assays (LFAs) are among the most widely used biosensors. Conventional colorimetric LFAs yield binary, non-quantifiable results. Electrochemical LFAs (eLFAs) aim to overcome this limitation. However, the reported disadvantage of extreme sensitivity to experimental conditions poses a significant challenge in the development of new eLFAs. In this work, impedimetric measurements were performed to probe the impact of experimental factors such as electrolyte conductivity/ionic strength on the eLFA outcomes. The time-dependent evolution of non-Faradaic impedance measurements coupled with flow velocity computations were employed to investigate ion transport across different LFA configurations and buffer compositions. The analysis of the Péclet number, a characteristic dimensionless parameter of flow dynamics within the LFA, revealed the coexistence of diffusive and convective ionic transport regimes over the LFA operational time. In addition, our findings underscore the critical role of absorbent pad dead volume adjustments in governing the capacity to maintain sample flow within the membrane over extended durations. These electro-fluidics phenomena are essential considerations for conducting electrochemical measurements within LFAs. Overall, this study offers insights into key design parameters for the integration of electrochemistry into LFAs. [ABSTRACT FROM AUTHOR]

Mycotoxins are responsible for several diseases in human and animals which impose huge medical and economical burdens worldwide, especially in low-income countries. Therefore, many countries set regulations for maximum permissible amounts of mycotoxins in food that need detection methods to monitor the mycotoxin concentration. Conventional methods like Enzyme‐linked immunosorbent assay (ELISA), high-performance liquid chromatography (HPLC) with fluorescence detection (HPLC-FLD), liquid chromatography-tandem mass spectrometry (LC-MS/MS) and gas chromatography (GC) are widely used to quantitatively detection of mycotoxins. However, these methods have limitations that make them suitable for laboratories and factories, but cannot be used regularly in the daily markets. Thus, the pursuit for sensitive, specific, handheld, and low-cost methods as effective elective strategies for point-of-care testing (POCT) has gotten much consideration in recent years. POCT is one of the ultimate objectives of mycotoxin determination, so the test must be quick, particular, delicate, open, and simple to be used. In this review, after a brief introduction to mycotoxins and their importance, the common tools of mycotoxin detection and their advantages and disadvantages are discussed, then lateral flow assay (LFA) systems working principles, classifications, and their advantages as a POCT method are explained. Furthermore, some studies regarding the development of LFA in detecting several types of mycotoxins and their innovations in LFAs' design and techniques for overcoming traditional LFAs' limitations are comprehensively reviewed. Various strategies are utilized to extend sensitivity and give quantitative sensing. Utilizing numerous visualization strategies, utilizing distinctive labeled reporters, coordination LFA with other detection strategies to take advantage of both LFA and detection gadgets, and utilize restrictive membranes to upgrade reagent reactivity, are a few of the strategies that are examined in this literature update regarding the use of LFA in the determination of mycotoxins, which covers. The period of time from 2015 till 2023 and based on the use of data bases. • LFAs offer rapid, cost-effective mycotoxin detection in food and feed samples. • Development of LFAs now covers a wide range of mycotoxins for comprehensive monitoring. • Improved LFA technology enhances sensitivity and specificity for reliable screening. • Integration with smartphone-based systems enhances portability and accessibility. • Ongoing research promises further improvements in LFA performance and applicability. [ABSTRACT FROM AUTHOR]

[Display omitted] • Nanozymes-based LFAs with respective to design, principle and application were firstly reviewed. • The activity of nanozymes associated with different compositions was discussed. • Trends in developing higher-performance nanozymes-based LFAs were prospected. • Advance in such LFAs significantly guaranteed the human health and public safety. Development of low-cost, portable and disposable sensors for rapid, on-site and reliable analysis of various analytes is of high urgency and importance for ensuring people's livelihood and public safety. With the emergence and popularity of digital color imaging technology, lateral flow assays (LFAs) have gradually become one of the most important sensors for point-of-care application. Compared with colloidal gold nanoparticles, nanozymes not only provide color strength via target-induced specific enrichment of nanozymes at test line, but also deepen color signal via nanozymes-assisted catalytic oxidation of colorless substrates into colored products, hence supplying a great potential for constructing higher-performance LFAs. Driven by this, considerable efforts have been devoted to nanozymes-based LFAs. In this review, we summarize the recent progress on the design and synthesis of nanozymes with different compositions showing different activity, and construction and application of nanozymes-based LFAs for pesticides, foodborne pathogens, biotoxins, food additives, antioxidants, disease biomarkers, and viruses. Further, the current challenges and future development directions are briefed for readers to develop higher-performance nanozymes-based LFAs. We expect this review will not only drive the significant advance of LFAs, but also promote their practical application in real samples, guaranteeing human health and public safety. [ABSTRACT FROM AUTHOR]

The increasing food allergic cases have posed a great challenge to the human health worldwide. Avoiding the exposure to food allergens is still the only efficient method to prevent food allergic reactions for food allergic sufferers because there are no effective treatments for food allergy up to now. Therefore, it is crucial to develop analytical technologies for rapid and on-site detection of food allergens, helping allergy-prone groups make proper option when buying or eating foods. Recently, various detection technologies and portable analytical devices have been widely developed for rapid, accurate, and on-site screening of food allergens. In this review, first of all, the traditional detection methods were simply introduced and discussed. Then, the latest advances on emerging biosensing technologies (i.e., colorimetric biosensing, fluorescent biosensing, electrochemical biosensing, surface enhanced Raman spectroscopy (SERS) biosensing, surface plasmon resonance (SPR) biosensing, and dual-mode biosensing) and portable analytical devices (such as lateral flow assays (LFAs), microfluidic chips, and paper-based microfluidic devices) for the detection of food allergens were comprehensively summarized. Furthermore, the advantages and weaknesses of these analytical methods for food allergen detection were also compared and discussed. Most importantly, the remaining challenges and chances in this important field were also proposed. Emerging biosensing technologies have shown a great potential in the on-site detection of food allergens. Impressively, the integration of biosensing technologies and portable detection devices (e.g., LFAs, microfluidic chips, and paper-based microfluidic devices) is one of the most promising methods for reliable and on-site detection of food allergens. [Display omitted] • Emerging biosensors and portable analytical devices are promising for food allergen detection. • Food sample preparation methods before food allergen detection were discussed. • Advantages and disadvantages of existing food allergen detection technologies were discussed. • Remaining challenges and chances in this promising field were proposed. [ABSTRACT FROM AUTHOR]

The prevalence and significant health risks associated with mycotoxin contamination in traditional Chinese medicines underscore the need for on-site detection methods. The complexity of matrices necessitates sample pretreatment before analysis, leading to prolonged analysis times and intricate procedures. In this study, we employed aptamer-modified magnetic nanoparticles (aptamer-MNPs) as labels and devised a three-in-one lateral flow aptasensor for on-site detection of ochratoxin A (OTA) in Astragalus membranaceus. By integrating aptamer-MNPs that serve in recognition, separation, and signal amplification roles, we streamlined purification, enrichment, and detection within a single aptasensor in 30 minutes. The developed aptasensor demonstrated high sensitivity (0.053 ng/mL), excellent specificity, and satisfactory accuracy. Analyzing twelve actual samples revealed three as OTA-positive, aligning well with HPLC analysis results. These results showcase the potential of the lateral flow aptasensor as a cost-effective, reliable, and robust on-site screening technique for trace-level mycotoxins in complex matrices. • Aptamer-modified MNPs were employed as labels in lateral flow assays. • A 3-in-1 aptasensor for purification, enrichment, and detection was developed. • The developed aptasensor exhibits high sensitivity, specificity, and accuracy. • Outcomes of real samples analysis by the aptasensor aligned well with HPLC. [ABSTRACT FROM AUTHOR]

Background: COVID-19 has resulted in significant morbidity and mortality worldwide. Lateral flow assays can detect anti-Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) antibodies to monitor transmission. However, standardized evaluation of their accuracy and tools to aid in interpreting results are needed. Methods: We evaluated 20 IgG and IgM assays selected from available tests in April 2020. We evaluated the assays' performance using 56 pre-pandemic negative and 56 SARS-CoV-2-positive plasma samples, collected 10–40 days after symptom onset, confirmed by a molecular test and analyzed by an ultra-sensitive immunoassay. Finally, we developed a user-friendly web app to extrapolate the positive predictive values based on their accuracy and local prevalence. Results: Combined IgG + IgM sensitivities ranged from 33.9 to 94.6%, while combined specificities ranged from 92.6 to 100%. The highest sensitivities were detected in Lumiquick for IgG (98.2%), BioHit for both IgM (96.4%), and combined IgG + IgM sensitivity (94.6%). Furthermore, 11 LFAs and 8 LFAs showed perfect specificity for IgG and IgM, respectively, with 15 LFAs showing perfect combined IgG + IgM specificity. Lumiquick had the lowest estimated limit-of-detection (LOD) (0.1 μg/mL), followed by a similar LOD of 1.5 μg/mL for CareHealth, Cellex, KHB, and Vivachek. Conclusion: We provide a public resource of the accuracy of select lateral flow assays with potential for home testing. The cost-effectiveness, scalable manufacturing process, and suitability for self-testing makes LFAs an attractive option for monitoring disease prevalence and assessing vaccine responsiveness. Our web tool provides an easy-to-use interface to demonstrate the impact of prevalence and test accuracy on the positive predictive values. [ABSTRACT FROM AUTHOR]

Zoonoses and animal diseases threaten human health and livestock biosecurity and productivity. Currently, laboratory confirmation of animal disease outbreaks requires centralized laboratories and trained personnel; it is expensive and time-consuming, and it often does not coincide with the onset or progress of diseases. Point-of-care (POC) diagnostics are rapid, simple, and cost-effective devices and tests, that can be directly applied on field for the detection of animal pathogens. The development of POC diagnostics for use in human medicine has displayed remarkable progress. Nevertheless, animal POC testing has not yet unfolded its full potential. POC devices and tests for animal diseases face many challenges, such as insufficient validation, simplicity, and portability. Emerging technologies and advanced materials are expected to overcome some of these challenges and could popularize animal POC testing. This review aims to: (i) present the main concepts and formats of POC devices and tests, such as lateral flow assays and lab-on-chip devices; (ii) summarize the mode of operation and recent advances in biosensor and POC devices for the detection of farm animal diseases; (iii) present some of the regulatory aspects of POC commercialization in the EU, USA, and Japan; and (iv) summarize the challenges and future perspectives of animal POC testing.

The working principle of lateral flow assays, such as the widely used COVID-19 rapid tests, is based on the capillary-driven liquid transport of a sample fluid to a test line using porous polymeric membranes as the conductive medium. In order to predict this wicking process by simplified analytical models, it is essential to determine an effective capillary radius for the highly porous and open-pored membranes. In this work, a parametric study is performed with selected simplified structures, representing the complex microstructure of the membrane. For this, a phase-field approach with a special wetting boundary condition to describe the meniscus formation and the corresponding mean surface curvature for each structure setup is used. As a main result, an analytical correlation between geometric structure parameters and an effective capillary radius, based on a correction factor, are obtained. The resulting correlation is verified by applying image analysis methods on reconstructed computer tomography scans of two different porous polymeric membranes and thus determining the geometric structure parameters. Subsequently, a macroscale flow model that includes the correlated effective pore size and geometrical capillary radius is applied, and the results are compared with wicking experiments. Based on the derived correction function, it is shown that the analytical prediction of the wicking process in highly porous polymeric membranes is possible without the fitting of experimental wicking data. Furthermore, it can be seen that the estimated effective pore radius of the two membranes is 8 to 10 times higher than their geometric mean pore radii.

Green/sustainable cellulose paper-based platforms at point-of-care testing have received much interest in biomedical fields at resource-limited settings. In such platforms, biomolecules would have to be immobilized onto paper substrate to achieve detection. Therefore, effective molecule immobilization is critical for the performance of paper-based testing, especially the detection sensitivity. Although various methods have been developed to improve the detection sensitivity, the immobilization capacity of biomolecules has not been considered yet. In this study, we developed a facile method for modifying paper materials by incorporating chitosan into NC membrane, which has positive effect on the pore size, porosity, and surface groups of NC membrane, hence, enhancing the immobilization capacity of biomolecules on paper substrate. We demonstrated the effectiveness of the above mentioned approach by using nucleic acid lateral flow assay (NALFAs) as a model paper-based platform; in comparison with the unmodified NALFAs, the results showed ten-fold increase in the detection sensitivity for HBV. We envision that this facile modification method will have a great potential for developing other highly sensitive paper-based detecting platforms. [ABSTRACT FROM AUTHOR]

The transmission of Tuberculosis (TB) is very rapid and the burden it places on health care systems is felt globally. The effective management and prevention of this disease requires that it is detected early. Current TB diagnostic approaches, such as the culture, sputum smear, skin tuberculin, and molecular tests are time-consuming, and some are unaffordable for low-income countries. Rapid tests for disease biomarker detection are mostly based on immunological assays that use antibodies which are costly to produce, have low sensitivity and stability. Aptamers can replace antibodies in these diagnostic tests for the development of new rapid tests that are more cost effective; more stable at high temperatures and therefore have a better shelf life; do not have batch-to-batch variations, and thus more consistently bind to a specific target with similar or higher specificity and selectivity and are therefore more reliable. Advancements in TB research, in particular the application of proteomics to identify TB specific biomarkers, led to the identification of a number of biomarker proteins, that can be used to develop aptamer-based diagnostic assays able to screen individuals at the point-of-care (POC) more efficiently in resource-limited settings.

Lateral flow assays (LFAs) have been introduced and developed over the last half century. This technology is widely used as a tool for diagnosis in several fields such as environment, food quality and healthcare. Point-of-care (POC) diagnosis using LFAs has been attracting attention of the research community, particularly aiming for the development of a platform that can evaluate of biological markers in bodily fluids such as saliva and urine. The existence of a disease or the pregnancy can be determined by a test device, before further investigation and medical treatment. LFAs make use of a disposable test strip, which can provide diagnosis result on the spot within minutes. Thus, LFAs is a promising alternative of preliminary diagnosis for laboratory instruments that are costly, time consuming and require trained personnel. This paper includes a brief overview of the conventional LFAs: material selection based on its roles and characteristics, working principles, fundamentals, applications, and design criteria. We mainly discuss the technical challenges in both engineering and biochemical aspects and recommends possible solutions. We identify current research trends and provide perspectives of advanced technologies for enhancing assay performance. [ABSTRACT FROM AUTHOR]