Lateral Flow Sensors Research Articles

Resistivity detection of perfluoroalkyl substances with fluorous polyaniline in an electrical lateral flow sensor

Perfluoroalkyl substances (PFAS), known as "forever chemicals," are a growing concern in the sphere of human and environmental health. In response, rapid, reproducible, and inexpensive methods for PFAS detection in the environment and home water supplies are needed. We have developed a simple and inexpensive perfluoroalkyl acid detection method based on an electrically read lateral flow assay (e-LFA). Our method employs a fluorous surfactant formulation with undoped polyaniline (F-PANI) fabricated to create test lines for the lateral flow assay. In perfluoroalkyl acid sensing studies, an increase in conductivity of the F-PANI film is caused by acidification and doping of PANI. A conductivity enhancement by 104-fold can be produced by this method, and we demonstrate a limit of detection for perfluorooctanoic acid (PFOA) of 400 ppt and perfluorobutanoic acid of 200 ppt. This method for PFOA detection can be expanded for wide-scale environmental and at-home water testing.

Graphene Nanobeacons with High‐Affinity Pockets for Combined, Selective, and Effective Decontamination and Reagentless Detection of Heavy Metals (Small 33/2022)

Heavy Metal Detection In article number 2201003 by Aristides Bakandritsos, Michal Otyepka, and co-workers, recyclable fluorescent nanographenes with high-affinity pockets for toxic metals enable selective and highly effective decontamination of natural water and quality monitoring with sub-nanomolar sensitivity for lead and cadmium via photoluminescence quenching or optical lateral-flow sensors.

Prototype Digital Lateral Flow Sensor Using Impact Electrochemistry in a Competitive Binding Assay.

This work demonstrates a lateral flow assay concept on the basis of stochastic-impact electrochemistry. To this end, we first elucidate requirements to employ silver nanoparticles as redox-active labels. Then, we present a prototype that utilizes nanoimpacts from biotinylated silver nanoparticles as readouts to detect free biotin in solution based on competitive binding. The detection is performed in a membrane-based microfluidic system, where free biotin and biotinylated particles compete for streptavidin immobilized on embedded latex beads. Excess nanoparticles are then registered downstream at an array of detection electrodes. In this way, we establish a proof of concept that serves as a blueprint for future "digital" lateral flow sensors.

(Digital Presentation) Stochastic Impact Electrochemistry in a Lateral-Flow Sensor Architecture

During the recent SARS-CoV-2 pandemic, lateral flow assays (LFAs) have written an indescribable story of success, as they provide means for decentralized, low-cost, and easy-to-use testing. However, LFAs in their most common form support only qualitative results and their sensitivity and limit of detection is significantly limited by the colorimetric readout method. In contrast, single-impact electrochemistry offers the possibility to quantify species beyond picomolar concentrations by recording individual species collisions with a biased microelectrode. Within this work, we investigate the integration of stochastic sensing into a LFA-architecture by combining a wax-patterned microchannel with a microelectrode array in order to detect silver nanoparticles by their oxidative dissolution. Here, we demonstrate the possibility to resolve individual nanoparticle collisions in a paper-based microchannel using a simplified reference-on-chip setup. Furthermore, we simulated a lateral-flow sensor, by flushing previously dried silver nanoparticles towards the electrode array, where the particles are subsequently detected. This proof-of-principle illustrates that single-impact electrochemistry might be a promising technique to extend the capability of LFAs. Especially, the integration of functionalized nanoparticle labels could enable the rapid and on-site detection of very dilute species with exceptional sensitivity.

A paper-based lateral flow sensor for the detection of thrombin and its inhibitors

Thrombin is a biomarker of blood-related diseases. Its detection and inhibitor screening are of great significance in the fields of medical and biological research. Herein, a novel paper-based lateral flow sensor for thrombin detection and inhibitors screening is developed. The formation of cross-links of fibrin from fibrinogen during the blood clotting process can efficiently capture water molecules. Based on the principle, in the presence of thrombin, the water of the plasma solution cannot flow along with the test paper as the water molecules are trapped in the fibrin. Upon the inactivation of thrombin by the inhibitors, water can flow along the test paper. The detection limit of thrombin reaches about 16.1 mU/mL and the sensing platform also exhibits excellent selectivity, reproducibility, and stability. Furthermore, the inhibition efficiency of argatroban, a clinical drug used as a thrombin inhibitor, is also successfully investigated using the assay. Overall, this strategy exhibits the advantages of the specific enzymatic reaction, low-cost, and label-free detection of thrombin with high sensitivity and remarkable specificity. This method is also competent to facilitate the screening of thrombin inhibitors, which is promising in the discovery of therapeutic drugs for thrombus-related diseases.

Hydrogel-assisted paper-based lateral flow sensor for the detection of trypsin in human serum

The detection of trypsin and its inhibitor is significantly important for both clinical diagnosis and disease treatment. Herein, we demonstrate a hydrogel-assisted paper-based lateral flow sensor for the detection of trypsin and its inhibitor for the first time. The gelatin hydrogel is hydrolyzed based on the gel-to-sol transition in the presence of trypsin, which results in the release of the trapped water molecules in the gelatin hydrogel. By placing one end of a pH indicator strip onto the hydrolyzed gelatin hydrogel, water is flowing along the pH indicator strip. However, in the absence of trypsin, water cannot flow along the pH indicator strip as the water molecules are trapped in the gelatin hydrogel. The detection limit of the system reaches as low as 1.0 × 10−6 mg/mL, and it is also applied to the quantitative detection of trypsin in human serum. In addition, the detection of a clinical drug aprotinin that is an inhibitor of trypsin is also successfully achieved. Noteworthy, only the gelatin hydrogel, pH indicator strip, and PS substrate are needed to fulfill the detection of trypsin without the need of other chemicals or reagents. Overall, we develop a particularly simple, elegant, robust, competitive, high-throughput, and low-cost approach for the rapid and label-free detection of trypsin and its inhibitor, which is very promising in the development of commercial products for sensing, diagnostic, and pharmaceutical applications. Besides, the hydrogel-assisted paper-based lateral flow sensor can also be employed to detect other analytes of interest by use of different stimuli-responsive hydrogel systems.

16S rRNA Monitoring Point-of-Care Magnetic Focus Lateral Flow Sensor

The detection and profiling of pathogenic bacteria is critical for human health, environmental, and food safety monitoring. Herein, we propose a highly sensitive colorimetric strategy for naked eye screening of 16S ribosomal RNA (16S rRNA) from pathogenic agents relevant to infections, human health, and food safety monitoring with a magnetic focus lateral flow sensor (mLFS) platform. The method developed was demonstrated in model 16S rRNA sequences of the pathogen Escherichia coli O157:H7 to detect as low as 1 fM of targets, exhibiting a sensitivity improved by ∼5 × 105 times compared to the conventional GNP-based colorimetric lateral flow assay used for oligonucleotide testing. Based on the grayscale values, semi-quantitation of up to 1 pM of target sequences was possible in ∼45 min. The methodology could detect the target 16S rRNA from as low as 32 pg/mL of total RNA extracted from pathogens. Specificity was demonstrated with total RNA extracted from E. coli K-12 MG1655, Bacillus subtilis (B. subtilis), and Pseudomonas aeruginosa (P. aeruginosa). No signal was observed from as high as 320 pg/mL of total RNA from the nontarget bacteria. The recognition of target 16S rRNA from 32 pg/mL of total RNA in complex matrices was also demonstrated. The proposed mLFS method was then extended to monitoring B. subtilis and P. aeruginosa. Our approach highlights the possibility of extending this concept to screen specific nucleic acid sequences for the monitoring of infectious pathogens or microbiome implicated in a range of diseases including cancer.

Polymerase chain reaction with lateral flow sensor assay for the identification of horse meat in raw and processed meat products

A simple, rapid and accurate detection method for the authentication of animal species is urgently required in the food detection field. The present study established a horse-specific polymerase chain reaction integrated with a lateral flow sensor assay (Horse-PCR-LFS) for the rapid detection of horse meat. In this test, a cytb gene sequence of horse was amplified using PCR, the PCR amplicon was checked with the lateral flow sensor assay, and the result of the sensor can be read within 2–3 min by the naked eye. The detection limit of the test was up to 0.01% horse meat in artificially adulterated meat mixtures, the assay also successfully detected horse DNA in various commercial food samples. As a rapid and user-friendly molecular detection tool, this test provides an accurate detection format for the identification of horse and offers solutions to problems related to animal meat adulteration and animal-origin food safety and traceability.

Controlled copper in situ growth-amplified lateral flow sensors for sensitive, reliable, and field-deployable infectious disease diagnostics

A polyethyleneimine (PEI)-assisted copper in-situ growth (CISG) strategy was proposed as a controlled signal amplification strategy to enhance the sensitivity of gold nanoparticle-based lateral flow sensors (AuNP-LFS). The controlled signal amplification is achieved by introducing PEI as a structure-directing agent to regulate the thermodynamics of anisotropic Cu nanoshell growth on the AuNP surface, thus controlling shape and size of the resultant AuNP@Cu core–shell nanostructures and confining free reduction and self-nucleation of Cu2+ for improved reproducibility and decreased false positives. The PEI-CISG-enhanced AuNP-LFS showed ultrahigh sensitivities with the detection limits of 50 fg mL−1 for HIV-1 capsid p24 antigen and 6 CFU mL−1 for Escherichia coli O157:H7. We further demonstrated its clinical diagnostic efficacy by configuring PEI-CISG into a commercial AuNP-LFS detection kit for SARS-CoV-2 antibody detection. Altogether, this work provides a reliable signal amplification platform to dramatically enhance the sensitivity of AuNP-LFS for rapid and accurate diagnostics of various infectious diseases.

Gold nanoparticle-streptavidin conjugates for rapid and efficient screening of aptamer function in lateral flow sensors using novel CD4-binding aptamers identified through Crossover-SELEX.

To decrease the burden of laborious and reagent-intensive screening of modified aptamers, their binding function requires assessment in assay formats compatible with the end diagnostic application. Here, we report on the use of an alternative and cost-effective approach: a rapid lateral flow assay (LFA) utilising streptavidin-conjugated gold nanoparticles (AuNP) as reporter molecules to screen novel ssDNA aptamers for their ability to detect CD4. Crossover-SELEX was employed to identify CD4-targeting aptamers from a ssDNA library enriched against a recombinant human CD4 protein (hCD4) conjugated to magnetic-beads and to endogenous CD4 expressed by U937 cells. Counter-selection with IgG-conjugated beads and CD4-negative Ramos RA-1 cells was employed. Following SELEX, four sequences (U4, U14, U20 and U26) were selected for candidate screening. Fluorescence confocal microscopy showed comparable localization of the Cy5-labeled aptamer U26, compared to antibodies binding CD4's cytoplasmic domain. Aptamer-hCD4 binding kinetics were evaluated by a qPCR-based magnetic-bead binding assay to unmodified aptamers. U26 exhibited the highest binding affinity (Kd = 2.93 ± 1.03 nM) to hCD4-conjugated beads. Citrate-stabilized gold nanoparticles (mean particle diameter, 10.59 ± 1.81 nm) were functionalized with streptavidin to allow immobilization of biotin-labeled aptamers. Except for U4, the aptamer-gold nanoparticle conjugates (Apt-AuNP) remained stable under physiological conditions with their size (approx. 15 nm) appropriate for use in the LFAs. Lateral-flow based screening was used to evaluate the suitability of the Apt-AuNPs as CD4-detecting reporter molecules by immobilizing hCD4 and flowing the nanoparticle conjugates across the LFA. Using this approach, two novel sequences were identified as being suitable for the detection of hCD4: visual detection at 9 min was obtained using U20 or U26. After 20 min, equivalent colorimetric hCD4 responses were observed between anti-CD4 monoclonal antibody (ΔI = 94.19 ± 3.71), an existing CD4 aptamer F1-62 (ΔI = 90.31 ± 19.31) and U26 (ΔI = 100.14 ± 14.61) LFA's, each demonstrating high specificity to hCD4 compared to IgG. From the above, Crossover-SELEX allowed for the successful identification of ssDNA aptamers able to detect hCD4. Streptavidin-conjugated AuNPs, when bound to candidate aptamers, show potential application here as screening tools for the rapid evaluation of aptamer performance in low-cost lateral flow diagnostics.

Photothermal enhancement in sensitivity of lateral flow assays for detection of E-coli O157:H7

Lateral flow immunoassay (LFA) is a well-known point-of-care technology for the detection of various analytes. However, low sensitivity and lack of quantitative results are some of its critical drawbacks. Here we report a photothermal enhanced lateral flow sensor on the basis of the photothermal properties of reduced graphene oxide (rGO) for the detection of E-coli O157:H7 as a model pathogen. The calibration curve of the photothermal method exhibited a linear range from 5 × 10⁵ to 5 × 10⁷ CFU/ml with a correlation coefficient of R2 = 0.96 and a regression equation of y = 8.1x-43 for standard bacteria solutions in phosphate buffer. The limit of detection was ∼5 × 10⁵ CFU/ml for standard bacteria solutions, which was a 10-fold enhancement in sensitivity compared to the qualitative results. Specificity experiments showed that the photothermal method can only detect the target bacteria among 6 types of bacteria strains. It was confirmed that the developed technique could be a highly potential method for the rapid detection field because it can provide fast quantitative results with improved sensitivity.

Magnetic Focus Lateral Flow Sensor for Detection of Cervical Cancer Biomarkers.

We report on a magnetic focus lateral flow biosensor (mLFS) for ultrasensitive detection of protein biomarkers in a practical format. With valosin-containing protein as a target protein, we show that the developed mLFS concept could detect as low as 25 fg/mL with magnetic focus to enhance target capture efficiency to deliver a 106-fold improvement in sensitivity compared to that of conventional lateral flow (LF) systems. The conceptualized strategy utilizes a simple magnet placed beneath the three-dimensional printed LF device to concentrate the targets at the signal zone without any additional instrumentation. In addition, protein mixtures extracted from the tissue of cervical cancer patients was also utilized to validate the sensor. To investigate the effect of magnetic focus on sensitivity, surface-enhanced Raman spectroscopy and dark-field imaging was utilized to characterize the distribution and movement of Fe3O4 core-Au shell nanoprobes in a model LF strip. Our experiments show that the magnetic focus results in an increased interaction time between the magnetic probe-labeled targets and the capture antibody, yielding a higher capture efficiency, allowing for ultrasensitive detection of the target not possible before with LF. The proposed mLFS can be utilized to detect a range of trace protein biomarkers for early diagnosis and can be combined with diverse pretreatment and signal amplification steps to query complex samples.

Plasmonic enhancement in lateral flow sensors for improved sensing of E. coli O157:H7

We propose a plasmonic enhanced lateral flow sensor (pLFS) concept with an enhanced colorimetric signal by utilizing liposome encapsulating reagent to trigger the aggregation of gold nanoparticles (GNPs). Our signal enhancement strategy incorporates the simplicity of lateral flow immunoassays (LFIA) utilizing plasmonic enhancement. The conceptualized hybrid pLFS for onsite rapid detection of pathogens in low numbers in a user friendly format requiring simple steps is the first step in the translation of plasmonic enhancement sensing to a practical regime. The pLFS was carried out with a biotinylated liposome label ruptured to release branched polyethylenimine (BPEI) to trigger the aggregation of GNPs for colorimetric signal generation. BPEI has multiple amino groups and more positive charges in PBS buffer, therefore few of the BPEI groups could induce the aggregation of GNPs, resulting in an enhanced colorimetric signal to detect E. coli O157:H7. Compared with the reported conventional LFIA, the proposed pLFS demonstrated more than 1000-fold improvement in sensitivity. The pLFS could detect as low as 100 CFU/ml of E. coli O157:H7 in buffer and 600 CFU/ml E. coli O157:H7 in liquid food systems.

Disposable Printed Lateral Flow Electrochemical Immunosensors for Human Cardiac Troponin T

Here we report an electrochemical ELISA approach for cardiac Troponin T (cTnT) determination based on a lateral flow membrane with underlying screen printed electrodes ( $ /cm2). The thick film transducer was modified with the anti-cardiac Troponin T antibody via physisorption and the electrochemical performance of the immunosensor was evaluated using cyclic voltammetry. The capture antibody coated immunoelectrode employed for electrochemical determination of Troponin T antigen used a sandwich assay format with horseradish peroxidase conjugated signaling antibodies held in the track of a lateral flow sensor strip. A simple two step procedure realized signal acquisition within <20 min (total assay time). The lateral flow electrochemical immunosensor response resulted in a calibration curve with linear response (0–700 ng/ml cTnT) with limit of detection of 0.15 ng/ml.

Electrically-Actuated Valves for Woven Fabric Lateral Flow Devices.

The integration of flow control elements into low-cost biosensors presents a significant engineering challenge. This Article describes the development and integration of active, chemical valves into lateral flow devices, using a scalable, single-step, weaving-based manufacturing approach. The valve was constructed from an electrically conductive polymer, polypyrrole. The polymer switches between wetting and nonwetting states when it is reduced and oxidized via the application of an electrochemical potential. In this work, yarns were first coated with polypyrrole and integrated into fabric lateral flow sensors. The coated yarns were stimulated in situ via integrated electrodes. Coated textiles were characterized for their response to variations in the applied electrical potential, the duration for which the potential is applied, and the chemical composition of the polymer. Among these tuning parameters, the concentration of iron (iii) chloride utilized to catalyze the synthesis of the polymer, was found to be a significant determinant in the wetting range of the polymer. Complete ON/OFF flow control was achieved at applied potentials of 20 V.cm-1, within 120 s of stimulation, using 0.1 M iron (iii) chloride, making the valve fairly easy to incorporate into point-of-care format. The practical utility of the valve was demonstrated by performing a Lowry protein assay in the device, wherein fluid flow was deactivated to allow individual reaction steps to go to completion prior to reactivation. Significant improvements in the sensitivity and linear range of the devices are reported in a simple straight-channel, lateral flow device, with the potential to develop more complex channel geometries via the weaving-based approach.

Ultrasensitive detection of microbial cells using magnetic focus enhanced lateral flow sensors.

We report on an improved lateral flow immunoassay (LFIA) sensor with a magnetic focus for ultrasensitive naked-eye detection of pathogenic microorganisms at a near single cell limit without any pre-enrichment steps, by allowing the magnetic probes to focus the labelled pathogens to the target zone of the LF strip.

Development of a novel dual-functional lateral-flow sensor for on-site detection of small molecule analytes

Recently, more sensitive methods for analyzing small molecule analytes were required. Lateral-flow sensors (LFSs) have been used as reliable, rapid and cost-effective on-site screening technique to detect small molecule analytes. However, available LFSs for small molecule analytes were competitive and performed in “turn-off” mode, allowing low-sensitivity detection by the naked eye. In this study, we used dual-functional LFSs for monitoring small molecule analytes (clenbuterol CL), which based a fluorescence-quenching effect of gold nanoparticles (AuNPs). The most important feature of this sensor is its dual function as “turn-on” LFSs under excitation light and turn-off LFSs under natural light. The sensitivity of “turn-on” LFSs reached 0.04ng/m under excitation light; however, under natural light they performed as common “turn-off” AuNPs LFSs with 5.0ng/mL sensitivity. This novel LFS can be used in either traditional or highly sensitive tools to detect a variety of small analytes.

Nanoparticle embedded enzymes for improved lateral flow sensors

In this study, combining the nanoparticle embedded sensors with lateral flow assays, a novel strategy for ensuring the quality of signalling in lateral flow assays (LFAs) was developed. A LFA for reactive oxygen species (ROS) is reported that is based on horse radish peroxidase (HRP) which is co-entrapped with Texas Red dextran inside porous polyacrylamide nanoparticles. In this system, enzymes are protected in the porous matrix of polyacrylamide which freely allows the diffusion of the analyte. The sensor is rapid and sensitive for quantification of hydrogen peroxide concentrations. A test solution of hydrogen peroxides was quantified with this novel LFA-ROS sensor to obtain a linear range between 1 and 25 μM. Nanoparticle embedding of enzymes is proposed here as a general strategy for developing enzyme-based lateral flow assays, eliminating adverse effects associated with biological samples.