Enhancing the Peroxidase-Mimicking Activity of Gold Nanoparticles for Lateral Flow Assays: Quantitative Evaluation in a Kinetic View.

Highly sensitive lateral flow immunoassays (LFIAs) are essential for various point-of-care applications, and gold nanoparticles (Au NPs) are by far the most commonly used labels. However, conventional LFIAs often suffer from high detection limits (LOD) or low sensitivity. In this study, we investigated three strategies to enhance the sensitivity of LFIAs by improving the peroxidase-mimicking (POD) activity of Au NPs. The POD activity of unmodified Au NPs was negligible (<0.01 units/mg, U/mg). The first strategy involved coupling Au NPs with horseradish peroxidase (HRP), which increased the POD activity to 65 U/mg. The second approach involved forming a thin palladium or iridium shell on Au NPs, which elevated the POD activity to 0.69-0.71 U/mg. The third strategy involved binding mercury ions (Hg²⁺) to Au NPs, resulting in a POD activity of up to 3 U/mg. Finally, we developed a simple quantitative model to estimate the LOD of LFIAs based on the POD kinetic parameters. Using Au-HRP conjugates, we demonstrated that the experimentally measured LOD was consistent with the calculated values. The developed model provides a framework for evaluating LFIAs with catalytic signal amplification and can be used to guide the development of highly sensitive assays.