Electrostatic Control of Silica Microbeads Improves Performance of Microscale Surface Ion Conduction Biosensors

In microscale surface ion conduction (μSIC) sensing, binding of charged analytes to functionalized bead surfaces alters interfacial ion transport, making sensor performance highly dependent on surface electrostatics. Previously, probe immobilization relied on commercially available streptavidin-coated beads, which exhibit large intrinsic zeta potentials and fixed biotin-based coupling chemistry. Here, we report an in-house surface functionalization strategy for silica microbeads that enables controlled probe immobilization and substantially reduced baseline surface charge. Sequential APTES-glutaraldehyde-ACE2 modification yielded beads with near-neutral ζ (-2.54 ± 0.73 mV), compared to -48.83 ± 0.73 mV for streptavidin-coated polystyrene beads. The lower intrinsic surface charge reduces electrostatic repulsion between the sensing interface and negatively charged viral targets, improving adsorption efficiency and surface coverage. Using ACE2-functionalized silica beads, the μSIC sensor achieved a limit of detection (LOD) of 560 pM for SARS-CoV-2 spike S1 protein and 9.4 fM (5.7 × 10⁶ copies/mL) for the inactivated SARS-CoV-2 whole virus, with excellent selectivity against the Influenza A virus. Notably, viral detection performance improved relative to previous streptavidin-based implementations despite the use of ACE2 rather than polyclonal antibodies. These results demonstrate that engineering baseline surface electrostatics is a powerful design parameter in surface ion conduction sensing and establish a tunable bead-functionalization strategy for adaptable, label-free pathogen detection.