Nanodiamond conjugated SARS-CoV-2 spike protein: electrochemical impedance immunosensing on a gold microelectrode

A promising immunosensing strategy in diagnosing SARS-CoV-2 is proposed using a 10-µm gap-sized gold interdigitated electrode (AuIDE) to target the surface spike protein (SP). The microelectrode surface was modified by (3-glycidyloxypropyl) trimethoxysilane to enforce the epoxy matrix, which facilitates the immobilization of the anti-SP antibody. The immunosensing performance was evaluated by integrating a nanosized (~ 10 nm) diamond-complexed SP as a target. The proposed immunoassay was quantitatively evaluated through electrochemical impedance spectroscopy (EIS) with the swept frequency from 0.1 to 1 MHz using a 100 mV_RMS AC voltage supply. The immunoassay performed without diamond integration showed low sensitivity, with the lowest SP concentration measured at 1 pM at a determination coefficient of R² = 0.9681. In contrast, the nanodiamond-conjugated SP on the immunosensor showed excellent sensitivity with a determination coefficient of R² = 0.986. SP detection with a nanodiamond-conjugated target on AuIDE reached the low limit of detection at 189 fM in a linear detection range from 250 to 8000 fM. The specificity of the developed immunosensor was evaluated by interacting influenza-hemagglutinin and SARS-CoV-2-nucleocapsid protein with anti-SP. In addition, the authentic interaction of SP and anti-SP was validated by enzyme-linked immunosorbent assay.