Protein Stacking on the APTES-Functionalized Pyrochlore Bi₂Ru₂O₇ Clusters for Ultrasensitive and Selective Immunosensing

With their unique physicochemical properties, such as metallic-like conductivity, favorable (electro)catalytic properties, electrochemical stability, and ease of functionalization, pyrochlores have found applications in various fields such as solid oxide fuel cells, batteries, thick film resistors, and temperature sensors; however, there are no reports on their application in electrochemical immunosensing. In this study, we exploited the (electro)catalytic nature and stability of the pyrochlore Bi₂Ru₂O₇ clusters silanized with (3-aminopropyl)triethoxysilane (APTES) to demonstrate their potential for the effective stacking of functional proteins. Characterization of the clusters by XPS disclosed a dual environment of Bi, also indicating the presence of Bi₂O₃ alongside APTES-Bi₂Ru₂O₇ clusters and, importantly, the predominant involvement of pyrochlore moieties in subsequent protein stacking. After stacking protein A and antibodies, the immunosensor revealed a nearly interference-free operation, high sensitivity, a detection limit of 118 fM SARS-CoV-2 spike protein, and operation in a wide examined concentration range of 10^-5-10^-1 μg mL^-1 with an <i>r</i>² of 0.98. In combination with a short incubation time of 30 min, the pyrochlore-based immunosensor provides a solid platform for future point-of-need applications.