The enzyme-linked immunosorbent assay (ELISA) is a widely used diagnostic technique due to its high sensitivity and specificity. However, its reliance on natural enzymes like horseradish peroxidase (HRP) poses limitations, including poor stability under harsh conditions and high production costs. This study explores metallic nanoparticles, including gold (Au), palladium (Pd), and platinum (Pt) nanoparticles, as peroxidase mimics in ELISA systems. Their peroxidase-like activity was evaluated through catalytic oxidation of 3,3', 5,5' tetramethylbenzidine (TMB) in the presence of hydrogen peroxide. Then these NPs were integrated into the ELISA platform through conjugating detection antibodies to the NP surface using established bioconjugation techniques. The developed nanozyme-based ELISA exhibited enhanced sensitivity, lower detection limits, and robust performance after prolonged storage, compared to HRP-based ELISA. This approach promises more stable, cost-effective, and accessible diagnostic assays, potentially transforming ELISA applications into clinical diagnostics, point-of-care testing, and resource-limited settings.
