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Click chemistry driven aggregate reaction dramatically enhances the sensitivity of dynamic light scattering
Weipeng Tong1, Mingjian Yao1, Hao Fang1
1State Key Laboratory of Food Science and Resource, Nanchang University, Nanchang, 330047, China; School of Food Science and Technology, Nanchang University, Nanchang, 330047, China.
Traditional dynamic light scattering (DLS) immunosensors rely on antigen-antibody interactions to directly trigger DLS probe aggregation, in which immune recognition and signal transduction are tightly coupled. However, the limited aggregation efficiency of DLS probes caused by suboptimal binding affinity of antigen-antibody interactions and inefficient target valency restrict the sensitivity of DLS immunosensors. Herein, we introduce a Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC)-amplified DLS immunosensor (CAD-immunosensor) that decouples immune recognition from click chemistry-driven aggregation. In this strategy, antigen-antibody binding enables specific target recognition, while CuAAC independently drives covalent aggregation of immunocomplexes through a polyvalent protein crosslinker. The valency and spatial configuration of the protein crosslinker were examined to improve the kinetics and efficiency of immunocomplex aggregation. Owing to the irreversibility of the click chemistry reaction, the aggregation efficiency of DLS probes is significantly increased, leading to a remarkable enhancement in the sensitivity of the CAD-immunosensor for detecting staphylococcal enterotoxin A with a limit of detection of 0.70 pg mL-1, representing 560-fold and 2986-fold sensitivity improvements over conventional DLS immunosensor and enzyme-linked immunosorbent assay (ELISA). Furthermore, the sensitivity of the developed CAD-immunosensor for detecting malachite green is also increased by 3675-fold compared to the ELISA method. The test time is reduced from 90 min with ELISA to 25 min with the CAD-immunosensor, and sample preparation is significantly simplified, especially for malachite green detection in fish and shrimp samples. Collectively, the sensing performance of this DLS immunosensor highlights its strong potential for ultrasensitive detection of trace analytes, characterized by enhanced sensitivity, reduced detection time, and robust operational performance.
Traditional dynamic light scattering (DLS) immunosensors rely on antigen-antibody interactions to directly trigger DLS probe aggregation, in which immune recognition and signal transduction are tightly coupled. However, the limited aggregation efficiency of DLS probes caused by suboptimal binding affinity of antigen-antibody interactions and inefficient target valency restrict the sensitivity of DLS immunosensors. Herein, we introduce a Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC)-amplified DLS immunosensor (CAD-immunosensor) that decouples immune recognition from click chemistry-driven aggregation. In this strategy, antigen-antibody binding enables specific target recognition, while CuAAC independently drives covalent aggregation of immunocomplexes through a polyvalent protein crosslinker. The valency and spatial configuration of the protein crosslinker were examined to improve the kinetics and efficiency of immunocomplex aggregation. Owing to the irreversibility of the click chemistry reaction, the aggregation efficiency of DLS probes is significantly increased, leading to a remarkable enhancement in the sensitivity of the CAD-immunosensor for detecting staphylococcal enterotoxin A with a limit of detection of 0.70 pg mL-1, representing 560-fold and 2986-fold sensitivity improvements over conventional DLS immunosensor and enzyme-linked immunosorbent assay (ELISA). Furthermore, the sensitivity of the developed CAD-immunosensor for detecting malachite green is also increased by 3675-fold compared to the ELISA method. The test time is reduced from 90 min with ELISA to 25 min with the CAD-immunosensor, and sample preparation is significantly simplified, especially for malachite green detection in fish and shrimp samples. Collectively, the sensing performance of this DLS immunosensor highlights its strong potential for ultrasensitive detection of trace analytes, characterized by enhanced sensitivity, reduced detection time, and robust operational performance.
