Polydopamine nanoparticle-based colorimetric–photothermal dual-mode lateral flow immunoassay for thromboembolism diagnosis

Linker-optimized bivalent nanobodies combined with photothermal gold liposome nanocomposites for enhanced dual-mode lateral flow immunoassay.

Lateral flow immunoassay (LFIA) has been widely used for rapid on-site analysis because of its simplicity, portability, and low cost. However, its performance in competitive assays, particularly for small molecule toxins, remains limited, as the properties of antibodies and the reliance on single-mode signal often lead to insufficient sensitivity and reliability. To address this challenge, we developed an enhanced colorimetric-photothermal LFIA by integrating linker-optimized bivalent nanobodies (BvNbs) and gold-liposome nanocomposites (Au-LNCs). Using tetrodotoxin (TTX) as a representative analyte, a series of BvNbs with different linker lengths were constructed. Molecular docking and molecular dynamics simulations further demonstrated that linker length modulates BvNbs detection performance by regulating conformational stability, local flexibility, and persistent interactions with free target molecules. In parallel, photothermal Au-LNCs were prepared, which exhibited broadband near-infrared absorption and a photothermal conversion efficiency as high as 78.06%. The resulting LFIA achieved a photothermal LOD of 1.89 ng/mL, corresponding to an approximately 10.7-fold improvement in sensitivity compared with conventional colloidal gold-based LFIA, while maintaining good reliability in real samples. Overall, this study provides mechanistic insight into the rational design of BvNbs and a promising strategy for improving LFIA performance.

Dual-Mode Lateral Flow Strip with Förster Resonance Energy Transfer for Rapid and Accurate Thrombin Detection

A dual-signal lateral flow biosensing platform for visual detection of miRNA-21 in serum.

Herein, we report a rationally designed dual-mode lateral flow assay (LFA) biosensor for the visual detection of microRNA-21 (miRNA-21), a critical breast cancer biomarker. The platform utilizes a synergistic readout of colorimetric signals from gold nanoparticles (AuNPs) and fluorescent signals from quantum dot nanobeads (QDNBs). Upon the specific introduction of target miRNA-21, the conformation of optimized hairpin nucleic acid probes conjugated on the nanomaterials is disrupted. This structural transition exposes capture sites, generating an AuNP-mediated colorimetric signal and a restored QDNBs fluorescence signal on the test lines by mitigating the inner filter effect (IFE) between them. Compared with conventional single-signal or amplification-dependent miRNA sensors, the proposed LFA integrates amplification-free target recognition, complementary AuNP-based colorimetric and QDNB-based fluorescent readouts, and IFE-regulated fluorescence recovery on a single strip, enabling visual screening and smartphone-assisted semi-quantitative analysis without complex instrumentation. Under optimal conditions, both the colorimetric and fluorescent readouts display broad linear correlations with miRNA-21 concentrations (5-1000 nM and 2.5-1000 nM, respectively), the limits of detection (LOD) of 1.599 nM and 1.354 nM. Furthermore, the platform demonstrates remarkable specificity against homologous miRNAs and robust stability. In spiked human serum, the assay achieves satisfactory recovery rates (91.8%-113.2%) with relative standard deviations (RSDs) below 6.0%. Furthermore, a preliminary evaluation using serum samples from five breast cancer patients and five healthy individuals showed clearly distinguishable colorimetric and fluorescent responses between the two groups, supporting the effectiveness of the proposed probe for miRNA-21 detection in real serum samples. These results indicate the potential of the dual-signal LFA platform for further clinical application.

MOF-assisted colorimetric-photothermal dual-readout lateral flow assay for rapid detection of Salmonella typhimurium.

BACKGROUND Foodborne pathogens remain a major threat to public health and food safety, among which Salmonella typhimurium is a leading cause of bacterial foodborne illness. Conventional culture-, immunoassay-, and nucleic-acid-based methods are reliable but often require laboratory facilities, trained personnel, and relatively long processing times, limiting their use for rapid on-site screening. Lateral flow immunoassays are attractive for field detection but commonly suffer from limited sensitivity and subjective visual interpretation. This study addresses the need for a rapid, specific, and more sensitive on-site assay for S. typhimurium. RESULTS We developed a phage-assisted colorimetric-photothermal dual-readout lateral flow immunoassay using zeolitic imidazolate framework-67-aggregated gold nanoparticles (ZIF-67@AuNPs) as signal probes. The ZIF-67 scaffold promoted dense AuNP assembly and aggregation-induced localized surface plasmon resonance coupling, resulting in broadened visible absorption around 650 nm and enhanced photothermal conversion. ZIF-67@AuNPs achieved a photothermal conversion efficiency of 45.7%, markedly higher than that of AuNPs alone (22.7%). Phage-based capture and antibody-functionalized ZIF-67@AuNPs enabled selective recognition of S. typhimurium on the test strip. In the colorimetric mode, the assay showed a linear range of 3 × 104-1 × 107 CFU/mL with a visual limit of detection (LOD) of 3.0 × 104 CFU/mL. In the photothermal mode, the linear range was 103-106 CFU/mL and the LOD improved to 0.997 × 103 CFU/mL, giving approximately 30-fold higher sensitivity. SIGNIFICANCE AND NOVELTY This work introduces a MOF-assisted LFIA that integrates phage-based biorecognition with colorimetric screening and photothermal quantification in a single test strip. The strategy reduces reliance on subjective visual judgment while retaining operational simplicity. The platform provides a practical approach for rapid on-site detection of S. typhimurium and can be extended to other foodborne pathogens by changing the recognition elements.

Pre-colored Prussian blue: A substrate-free dual-mode immunosensor for visible-to-thermal Staphylococcus aureus quantification.

Rapid identification of postoperative Staphylococcus aureus (S. aureus) infections is essential for timely treatment and improved clinical outcomes. Herein, we developed a substrate-free dual-mode lateral flow immunoassay (LFIA) using intrinsically colored Prussian Blue nanoparticles (PB NPs) as multifunctional signal reporters. The intrinsic blue color of PB NPs enables direct colorimetric visualization, while their excellent near-infrared photothermal conversion provides quantitative temperature readout. This self-signaling strategy eliminates enzymatic amplification and substrate incubation, simplifying the assay into a one-step sandwich immunoreaction. Under optimized conditions, the proposed LFIA achieved a wide linear detection range of 10-106 CFU/mL, with detection limits of 10 CFU/mL for the colorimetric mode and 1 CFU/mL for the photothermal mode. Analysis of real samples demonstrated good agreement with a commercial ELISA kit, with relative deviations below 10% and coefficients of variation below 15.5%, confirming the accuracy and practicality of the proposed platform. The dual-readout strategy improves analytical reliability through mutual signal verification, providing a simple, rapid, and reliable approach for S. aureus detection with strong potential for point-of-care testing.

Electrochemiluminescence Imaging Digital Immunoassay of Cytokines Secreted by Activated T Cells.

High-sensitivity cytokine detection is essential for predicting immunotherapy efficacy and monitoring treatment. Conventional methods such as ELISA suffer from limited sensitivity, while single-molecule immunoassays, although highly sensitive, often require physical compartmentalization, resulting in high cost and limited clinical applicability. Herein, we report a compartmentalization-free electrochemiluminescence (ECL) digital immunoassay based on silica-coated gold nanorod (AuNR@SiO2) for detecting tumor necrosis factor-α (TNF-α) secreted by activated T cells. The AuNR@SiO2 nanoparticles served as efficient nanoaccelerators, increasing the ECL photon emission rate of the tris(2,2'-bipyridine)ruthenium(II) (Ru(bpy)32+)/tri-n-propylamine (TPrA) system by 17-30-fold. Mechanistic investigations suggest that the enhancement originates from two complementary effects: plasmonic modulation associated with the localized surface plasmon resonance (LSPR) of the AuNR core, and nanoconfinement provided by the mesoporous SiO2 shell, which promotes local enrichment of ECL reactants and increases the effective reaction frequency around individual nanoparticles. Using this platform, TNF-α was detected with approximately one-order-of-magnitude higher sensitivity than conventional ELISA and a wide linear range of 10-50,000 pg/mL. The method also allowed direct analysis of TNF-α in cell culture supernatants without pretreatment, revealing activation-dependent secretion kinetics and confirming a positive feedback circuit in T cell cytokine production. Compared with existing single-molecule immunoassays, this ECL platform eliminates the need for precisely fabricated microchambers or time-consuming signal amplification, enabling straightforward digital readout while remaining compatible with standard immunoassay workflows. This work provides a simple and practical strategy for isolation-free ECL digital immunoassays and demonstrates promising potential for clinical translation.