From Upconversion Nanoparticles to Proteins: Probing Hydration-Water Density Fluctuations by Luminescence Thermometry.

Water appears simple, yet its anomalous behavior reveals an unexpected structural complexity. A growing body of evidence indicates that many of water's anomalies arise from fluctuations between low-density (LD) and high-density (HD) local structural motifs, a form of polymorphism that is well established in the supercooled regime and increasingly supported at ambient conditions. Yet, how these structural motifs manifest within hydration layers, where water interacts with nanoparticles, proteins, and charged interfaces, remains far less understood. This interfacial water governs colloidal stability, biomolecular function, and chemical reactivity, but its microscopic organization is difficult to probe directly with conventional bulk techniques. In this Account, we describe how luminescence nanothermometry provides a powerful and versatile approach to accessing density fluctuations in the hydration layer. By monitoring temperature-dependent optical and Brownian observables of luminescent probes, structural reorganizations of the surrounding hydration layer can be inferred with nanoscale sensitivity. Over the past several years, our group has shown that lanthanide-doped upconversion nanoparticles (UCNPs) and fluorescent proteins, such as enhanced green fluorescent protein (EGFP), act as local reporters of hydration-water density fluctuations. A central observation emerging from these studies is the existence of a crossover temperature,