Thermally imprinted microcone structure-assisted lateral-flow immunoassay platforms for detecting disease marker proteins.

Staphylococcal enterotoxins (SEs) are among the leading causes of foodborne illnesses in the Philippines. Rapid and sensitive detection of these toxins in food, particularly staphylococcal enterotoxin B (SEB) – the most potent type of SE – is imperative. Rapid toxin detection can be achieved using a lateral flow immunoassay (LFIA) device. However, low sensitivity has been a persistent issue with LFIA devices. To develop an LFIA device with improved sensitivity, this study investigated the exclusive use of antibodies as biorecognition molecules in a dual-detection probe LFIA device format and the configuration of the LFIA device to release the probes nonsequentially and sequentially. The LFIA device was fitted with two layers of glass fiber sample pads, loaded separately with the two detection probes, and partially separated by a Scotch tape film. For the nonsequential LFIA device, the two sample pads were pretreated with blocking buffers containing the same sugar concentration, whereas for the sequential LFIA device, the two sample pads were pretreated with blocking buffers containing different sugar concentrations. When tested against a filtered, homogenized durian candy sample, the nonsequential LFIA device generated higher signal intensity than the sequential LFIA device. This result suggests that aggregation of the detection probes was superior to sequential binding in enhancing the signal of the LFIA device for SEB detection. The sensitivity of the nonsequential LFIA device at 5 ng/mL was two-fold higher compared to previous POCT LFIA devices for SEB detection in food samples.

Real-world evaluation of COVID-19 lateral flow device (LFD) mass-testing in healthcare workers at a London hospital; a prospective cohort analysis

Lateral flow immunoassays (LFIAs) have been employed extensively for the rapid, accurate, and portable detection of protein biomarkers in healthcare. However, the cross-reactivity, especially in the multiplexed detection, leads to false-positive errors that would further limit their practical applications. In this work, we report a highly sensitive and accurate chemiluminescent LFIA based on the synthesis of the Au nanoparticle-antibody-horseradish peroxidase-polyethylene glycol conjugate for detecting cardiac troponin I (cTnI), a major biomarker of acute myocardial infarction. Due to the presence of polyethylene glycol, the accuracy of the LFIA was improved significantly from a clear false positive signal to the absence of a false positive signal. In addition, the device exhibited a highly sensitive detection of cTnI in the concentration range of 1-90 ng mL-1, and the detection limit can be as low as 10 pg mL-1. The method further enabled the multiplex detection of cTnI and myoglobin successfully. It is anticipated that this work may open new paradigms for the development of a variety of lateral flow devices with high sensitivity and accuracy and further lead to widespread practical applications in clinical diagnosis.

Aflatoxin B1 (AFB1) is a highly toxic and carcinogenic compound produced by certain fungi (e.g., Aspergillus flavus and Aspergillus parasiticus). Rapid and ultra-sensitive detection methods for AFB1 in various commodities are in high demand. This study aimed to enhance the sensitivity of a competitive lateral flow immunoassay (LFIA) for AFB1 detection by leveraging a previously developed experimental design strategy, named 4S. This approach comprises four phases—START, SHIFT, SHARPEN, and STOP—and involves the analysis of two reference conditions: NEG (0 ng/mL AFB1) and POS (1 ng/mL AFB1). By generating and overlaying response surfaces, regions of optimal NEG signal and POS/NEG signal ratio (IC%) were identified. Four variables were optimized: two related to the labeled antibody (its concentration and antibody-to-label ratio) and two to the competitor antigen (its concentration and hapten-to-protein ratio). An initial design defined the parameter space, while three subsequent designs did not yield further improvements in sensitivity. A strong anti-correlation was observed between the IC% and competitor parameters. The optimized LFIA-1 exhibited enhanced sensitivity, achieving a limit of detection of 0.027 ng/mL compared to 0.1 ng/mL for the original device. Additionally, the amount of expensive antibody required for device fabrication was reduced by around a factor of four.

The Monkeypox virus (MPXV), a DNA virus classified under the Orthpoxvirus genus alongside variola virus, has recently garnered significant global health attention due to its increasing transmission and emerging genomic mutations. Point-of-care testing is essential for effective clinical response and outbreak mitigation. In this article, we developed a novel class of colored microspheres designed for application in a lateral flow immunoassay (LFIA) platform targeting MPXV-specific biomarkers. Polystyrene-maleic anhydride (SMA-MAA) microspheres were synthesized with a high-temperature soap-free emulsion polymerization optimized in our lab. Subsequent alkali and acid treatments were employed to introduce porosity into the microsphere matrix. Solvent Red 27 and Disperse Red 60 were incorporated via solvent-swelling and thermal-swelling methods, respectively, to generate high brightness (HB) carriers. A surface coating composed of a tannic acid–iron (TA–Fe3⁺) coordination complex was applied to form a stable metal–polyphenol film (MPF). This coating not only minimized dye leaching by establishing a robust shell but also improved dye distribution, thereby enhancing overall color intensity. The final HB-LFIA system, configured in a sandwich immunoassay format, demonstrated favorable sensitivity and linear detection range for Monkeypox antigen, indicating strong potential for clinical diagnostic use.

Ultrabright magneto-fluorescent nanoparticles-enhanced lateral flow immunoassay for ultrasensitive detection of Escherichia coli O157:H7.

Natural toxin (for example mycotoxin and phycotoxin) contamination of food is of safety and economic concern, so much effort is devoted to the development of screening methods which enable the toxins to be continuously and widely monitored in food and feed. More generally speaking, rapid and non-instrumental assays for detection of a variety of food contaminants are generating ever-increasing scientific and technological interest because they enable high-throughput, economical, on-site monitoring of such contaminants. Among rapid methods for first-level screening of food contaminants, lateral-flow immunoassay (LFIA), also named immunochromatographic assay or immune-gold colloid immunoassay, has recently attracted scientific and industrial interest because of its attractive property of enabling very rapid, one-step, in-situ analysis. This review focuses on new aspects of the development and optimization of lateral-flow devices for mycotoxin and phycotoxin detection, including strategies for management of matrix interference and, particularly, for investigation of the improvements achieved by signal-enhancing strategies or by application of non-gold nanoparticle signal reporters.

Integrating lateral flow device with controllable gold in situ growth for sensitive detection of staphylococcal enterotoxin A in milk

Limitations in the sensitivity, linear detection range, and cross-reaction of lateral flow immunoassays mainly hamper their application in rapid screening for multiple targets. In this work, we designed a new time-resolved fluorescence immunoassay (TRFIA) platform to overcome these limitations. This platform uses europium chelate polystyrene (PS@Eu) nanoparticles conjugated with monoclonal antibodies to sense multiple mycotoxins. We employed a competitive TRFIA protocol in which the conjugated PS@Eu was used on the surfaces of photonic microbead arrays (PMAs). The TRFIA signal of PMAs on the pad was recorded with the digital time-resolved fluorescence reader. The developed TRFIA shows wide detection linear ranges (0.01-1000 ng/mL for DON, 0.1-100 ng/mL for OTA, and 0.01-100 ng/mL for AFB1), low limits of detection (LODs) (7.9 pg/mL for DON, 18 pg/mL for OTA, and 7.7 pg/mL for AFB1), good specificity, good recovery ratios (76.68-117.26%), and good reproducibility in grain samples. The simulated fluorescence enhancement effect of PMA indicated that the electric field distribution on the surface of PS@Eu on PMA is twice higher than that on the surface of PS@Eu. The new TRFIA for three kinds of mycotoxins was 1000-fold more sensitive than the classical TRFIA, and it has great potential application in rapid screening for multiple targets.

Rapid and multiplexed detection of biomarkers plays an indispensable role in disease diagnosis. Although paper-based lateral flow immunoassays have been widely used in this field, the speediness and throughput are still challenging issues. Herein, an electrochemical vertical flow immunoassay device (eVFID) is fabricated for rapid, ultrasensitive, and multiplexed detection of inflammatory biomarkers. Working electrodes with excellent electrochemical performance and permeability properties were directly fabricated on the nitrocellulose membrane to enable both the vertical flow of the sample solution and electrochemical detection. This vertical configuration can remarkably improve the speediness of the immunoassay and effectively inhibit the cross-talk reactions among immunomolecules, thus allowing rapid and simultaneous detection of multiplexed biomarkers. Furthermore, a signal amplification strategy based on horseradish peroxidase and tetramethylbenzidine was integrated into the eVFID to substantially increase the sensitivity of the electrochemical detection. A low limit of detection of 0.1 and 0.22 pg mL-1 was obtained for two low-abundance inflammatory biomarkers, interleukin-6 (IL-6) and procalcitonin (PCT), respectively. Finally, using a two-channel eVFID, simultaneous detection of IL-6 and PCT in human plasma samples was successfully realized within 5 min. We believe that the eVFID holds great promise for speedy and high-throughput biomarker detection at the point of care.

Biological fluid collection to identify and analyze different disease markers is a routine and normal procedure in health care settings. Body fluids are as varied as urine, blood, mucus, cerebrospinal fluid (CSF), tears, semen, etc. The volumes of the collected fluids range from micro liters (e.g., tears, CSF) to tens and hundreds of milliliters (blood, urine, etc.). In some manifestations, a disease marker (particularly protein markers) can occur in trace amounts, yet the fluids collected are in large volumes. To identify these trace markers, cumbersome methods, expensive instruments, and trained personnel are required. We developed an easy method to rapidly capture, concentrate, and identify protein markers in large volumes of test fluids. This method involves the utilization of two antibodies recognizing two different epitopes of the protein biomarker. Antibody-1 helps to capture and concentrate the biomarker and Antibody-2 adsorbed or conjugated to nanogold beads will detect the biomarker. This method was validated in capturing and detecting lipocalin type prostaglandin-D2 synthase, a marker in urine that implicates diabetic nephropathy. A one-step collection, concentration, and detection device was designed based on this method. This device can replace many of the normal body fluid collection devices such as tubes and containers. A one-step fluid collection and biomarker capture and concentration device for rapid diagnosis of diseases has tremendous advantage in terms of cost and providing timely results.

Biotin interferences: Have we neglected the impact on serological markers?

Afterglow time-resolved lateral flow immunoassay platform with a low-cost handheld reader for on-site detection of methamphetamine

Novel electrical detection of label-free disease marker proteins using piezoresistive self-sensing micro-cantilevers

Background: Chronic ischemic heart disease (CIHD) is characterized by persistent myocardial ischemic due to long-term reduced coronary blood flow. In the past, we mainly relied on surgical intervention or drug therapy to alleviate symptoms, but effective targeted treatments were scarce. Proteomics serves as a key tool to identify novel therapeutic targets. Methods: This study performed a bidirectional Mendelian randomization (MR) analysis by integrating genome-wide association study (GWAS) data on CIHD (10,894,596 single-nucleotide polymorphisms) with plasma proteomic data encompassing 4907 proteins. We conducted Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis to identify pathways linked to candidate protein biomarkers, searched the National Genomics Data Center (NGDC) database for existing evidence of their association with CIHD, and evaluated druggability through multi-dimensional analysis integrating the DSIGDB, ChEMBL, and clinical trial databases. Results: After eliminating the reverse effect, ultimately identifying 28 protein markers, including 16 risk-associated and 12 protective proteins. We also investigated their roles in the pathways related to CIHD. Meanwhile, the search confirmed that five of them were newly discovered protein markers. Ultimately, through evaluation, three priority therapeutic targets (CXCL12, PLAU, CD14) were identified for development. Conclusions: This study identified some biomarkers related to CIHD and analyzed the possible pathways involved. It also provided some new insights into the identification of the target and druggability.