Bimetallic gold/silver nanoclusters-gold nanoparticles based fluorescent sensing platform via the inner filter effect for hyaluronidase activity detection

Bimetallic nanoclusters, compared to monometallic nanoclusters, possess better chemical performance. Herein, with the assistance of chondroitin sulfate, a novel fluorescent gold/silver nanoclusters (Au/Ag NCs) have been developed and applied to establish a sensing platform for sensitive detection of hyaluronidase (HAase). The Au/Ag NCs fluorescence intensity can be distinctly quenched by gold nanoparticles (Au NPs) due to the inner-filter effect (IFE). With the introduction of protamine (PRO), the positively charged PRO can bind to negatively charged Au NPs via electrostatic attraction resulting in the aggregation of gold nanoparticles. When PRO was preferential interaction with the negatively charged hyaluronic acid (HA), it would lead to the disaggregation of Au NPs and quench the fluorescence again. However, HAase can hydrolyze HA into small fragments which prevented HA interacting with protamine, and leading to the aggregation of Au NPs and weaken the IFE of Au NPs on Au/Ag NCs. Thus, the fluorescence of bimetallic nanoclusters was “turn-on”. This “turn-on” sensing system can be utilized to detect HAase level in the range of 0.5–37.5 U mL⁻¹. The developed strategy not only expands the application scope of bimetallic nanoclusters, but also provides a new approach for the determination of HAase.