RESUMEN
The exploration of nanomaterials with mimic enzyme activity (named nanozyme) has gained extensive attention in the fields of advanced analytical chemistry and materials science. Herein, the gold nanoparticles doped covalent organic frameworks (COFs) were prepared, which exhibited not only excellent mimic nitroreductase activity but also robust stability. By replacing the traditional natural enzyme tag in an enzyme-linked immunosorbent assay (ELISA), we employed the proposed nanozyme to label the detecting antibody. According to the catalytic properties of the nanozyme, 4-nitrothiophenol (4-NTP) was introduced as the substrate, which can be transformed to 4-aminothiophenol (4-ATP) in the presence of NaBH4. In a surface enhanced Raman scattering (SERS) assay, 4-ATP was capable of functioning as a powerful bridge to connect the gold nanostars (with excellent SERS performance) by both the Au-S bond and electrostatic force to further produce a Raman "hot spot". Meanwhile, the Raman signal of 4-nitrothiophenol at 1573 cm-1 was weakened, and a new signal at 1591 cm-1 generated by 4-ATP was turned on, leading to the generation of a ratiometric SERS signal. Based on this performance, a ratiometric nanozyme-linked immunosorbent assay (NELISA) strategy was developed delicately, which was applied to detect ß-lactoglobulin (allergenic protein) by monitoring the ratiometric signal of I1591/I1573 with a limit of detection (LOD) of 0.01 ng/mL. The linear range is 25.65-6.2 × 104 ng/mL, covering more than 3 orders of magnitude. The developed method showed many advantages such as low-cost, higher recovery, and lower cross-reactivity, providing new insight into the application of SERS technology for trace target analysis.