Robust SERS Platform Enabled by Silver-Nanoparticle-Loaded Hydrogel Millibeads for Trace Detection in High-Salt and Protein-Rich Complex Matrixes

Reliable trace detection in complex matrixes (e.g., seawater, biofluids) is challenging for conventional surface-enhanced Raman scattering (SERS) platforms due to salt-induced aggregation, protein adsorption, and coffee-ring effects. We present a robust SERS platform using evaporative shrinkage of a silver-nanoparticle (AgNP)-loaded hydrogel millibead (Ag@Gel-MB). The hydrogel millibead selectively enriches analytes while excluding proteins, and its subsequent shrinkage coconcentrates analytes and AgNPs into ultrasmall volumes to form dense hotspots, while the 3D network prevents salt-induced aggregation. Using Nile Blue, we achieved high sensitivity (LOD < 10⁻¹⁰ M), excellent uniformity (RSD ∼ 4.36%), and batch reproducibility (RSD ∼ 4.39%). Unlike droplet methods needing superhydrophobic surfaces, our platform avoids the coffee-ring effect on any support and achieves 15-fold and 100-fold improvements in salt and protein tolerance (150 mM NaCl, 100 μg/mL BSA) over conventional methods. It successfully detected malachite green in seawater and fish serum, demonstrating its practical reliability for the analysis of complex samples.