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.
A dual-mode colorimetric–photothermal LFIA platform was successfully constructed for the detection of thrombosis-related biomarkers, including thrombomodulin and α2-plasmin inhibitor–plasmin complex, in clinical serum samples, enhancing analytical sensitivity and reliability.
Jun Yin, Yu Qiu, Yao Nie et al.· RSC Advances· 0 citations
The LIGMA assay is established as a viable electrokinetic strategy for improving paper-based biosensors, providing a foundation for expanded applications in clinical diagnostics, multiplexed detection, and portable field-deployable biosensing platforms.
D. McCornack, C. Ivory, Zhihong Zhang et al.· Analytical Chemistry· 0 citations
Non-invasive imaging of inflammation is critical for disease diagnosis, monitoring, and therapeutic evaluation. Nanobodies, due to their many unique properties, are emerging as powerful imaging agents; however, conventional non-specific conjugation strategies often yield heterogeneous products that limit in vivo performance. Here, we report a chemoenzymatic strategy that enables precise and modular assembly of nanobody conjugates using farnesyltransferase (FTase)-mediated site-specific modification. We developed a trifunctional FTase substrate that introduces two orthogonal bioorthogonal handles, an aldehyde and an azide, onto nanobodies in a single enzymatic step. These handles enable independent and controlled conjugation of imaging payloads (fluorophores or radioisotopes) and pharmacokinetic modifiers (i.e. PEG), which enhance in vivo signal-to-noise ratios. Applying this strategy to nanobodies targeting CD45 and CD11b, markers broadly expressed on all immune cells and myeloid subsets, respectively, we generated well-defined, dually modified constructs that enabled high-contrast in vivo visualization of immune organs, including the spleen and lymph nodes. In a mouse model of localized inflammation, these probes robustly delineated inflamed tissue with high sensitivity and specificity. Together, this FTase-enabled dual-labeling approach provides a versatile and generalizable platform for the rapid generation of precisely defined nanobody-based imaging agents, advancing non-invasive detection and monitoring of inflammation across diverse disease contexts.
Sneha Venkatachalapathy, Y. Lee, A. Farid et al.· Angewandte Chemie· 0 citations
We have developed a one-pot, two-stage method for the synthesis of highly monodispersed fluorescent polystyrene nanoparticles with surface carboxyl functional groups. The fluorescent nanoparticles have a tunable size range of 150−300 nm and excellent photo/chemical stability. Through conjugating antibodies to the surface carboxyl groups, various lateral flow immunoassays (LFIAs) using the fluorescent nanoparticles as the reporters have been developed, achieving a visual limit of detection (LOD) of 39 mIU mL−1 using human chorionic gonadotropin as the model sample. This LOD is 8-fold and 64-fold better as compared to LFIAs using AuNPs and free dye as reporters, respectively. The LODs of our optimized LFIAs are 3 pg mL−1 for HIV-1 p24, 4 ng mL−1 for Zika NS1, and 1 and 0.5 ng mL−1 for Staphylococcus aureus enterotoxin types A and B, respectively. These LOD values are below the clinical cutoffs and are comparable to ELISA results reported in the literature.
K. Chan, Xiaojun Chen, J. Soh et al.· ACS Materials Letters· 0 citations
This programmable dual-aptamer amplified luminescent proximity homogeneous assay (AlphaLISA) provides a rapid, homogeneous, and highly adaptable strategy for tumor-derived sEVs analysis, offering promising potential for noninvasive cancer diagnostics.
D-Proline has been reported as a potential biomarker for gastric (stomach) cancer, creating a demand for practical and rapid analytical approaches for early diagnosis using simple and non-invasive clinical samples. In this work, a microfluidic paper-based analytical device (µPAD) integrating molecularly imprinted polymer-functionalized upconversion nanoparticles (UCNPs@MIPs) was developed for the selective fluorescence-based detection of D-proline. Owing to near-infrared excitation (980 nm), UCNPs provide background-free fluorescence with high photostability, while MIPs serve as artificial receptors to impart molecular selectivity. The sensing mechanism relies on fluorescence quenching of UCNPs@MIPs induced by D-proline through multiphonon relaxation, resulting in an inverse relationship between fluorescence intensity and analyte concentration. Quantitative analysis was achieved using a homemade IR-laser-controlled light box and smartphone-based image capture, with signal evaluation based on blue-channel intensity. The proposed platform exhibited a linear response over the concentration range of 0.01-0.30 mM, with a limit of detection of 0.016 mM. The method was successfully applied to the determination of spiked D-proline in human saliva and serum samples. Owing to its simplicity, low sample consumption, portability, and adaptability, the proposed sensing strategy shows strong potential for on-site analysis and can be extended to broader applications in chemical analysis, biosensing, and environmental monitoring.
Akarapong Prakobkij, Jiajia Zhou, Mahnaz Maddahfar et al.· In Analysis· 0 citations