Rethinking the Role of AuNPs in Colorimetric Aptasensors: Dominant Nonspecific Interactions with Antibiotics over...

Gold nanoparticles (AuNPs) have been widely used in aptasensors due to the distinct color differences between their dispersed and aggregated states. However, recent studies have revealed that such colorimetric signals may originate from direct interactions between small molecule targets and AuNPs, rather than from specific aptamer-target binding. Here, we investigated the interactions between 26 antibiotics and AuNPs, and found that all antibiotics except NFZ exhibited strong binding to AuNPs, leading to decreased salt stability. The aggregation induced by these antibiotics could be reversed by mPEG-SH, indicating that it occurs in the secondary minimum. Using density functional theory, we analyzed the molecular structures and charge distribution characteristics of these antibiotics and proposed mechanisms for their interactions with AuNPs. Finally, we proposed the underlying mechanism of AuNPs aggregation induced by antibiotics. Additionally, we found that the aptamers adsorbed on AuNPs could not be desorbed via specific target-aptamer binding, as the aptamer's affinity was significantly reduced due to the adsorption. Our study demonstrates that AuNPs are unsuitable as carriers for developing colorimetric aptasensors or validating aptamer affinity, and researchers should exercise caution when using AuNP-based systems.