Integration of high refractive index sulfonated polystyrene opals with aptamers for rapid testing of neuron-specific enolase

We introduce a microplate assay for Neuron-Specific Enolase (NSE), a pivotal biomarker in neurodegenerative disorders, neuroendocrine tumors, and lung cancers, including small-cell lung cancer (SCLC) and non-small-cell lung cancer (NSCLC). The assay involves the self-assembly of polystyrene microspheres into a photonic crystal array (PCA) via pre-adsorption within microplate wells. Sulfonation of the PCA with sulfuric acid yields sulfonated-PCA (SPCA) with a high refractive index. NSE-specific aptamers are crosslinked to SPCA using EDC crosslinking, resulting in an aptamer-linked sulfonated photonic crystals assay (APSA). This APSA platform is employed to detect NSE in human serum samples. Aptamer-NSE binding induces shifts in wavelength values (∆PWV), generating discernible color changes in SPCA. This binding is further analyzed by molecular dynamics simulations. As NSE concentrations increase, the refractive index decreases, causing reflection peak shifts across the entire visible wavelength range. The assay demonstrates remarkable sensitivity with a limit of detection (LOD) of 3.21 ± 0.45 pg mL⁻¹ and a rapid response time of 30 s. This sensitivity outperforms existing biosensing methods accompanied by better selectivity. The presented APSA platform serves as a robust NSE detection tool in human serum and holds promise for customization to target other molecules, offering significant potential for clinical applications.