A Folding-Based Electrochemical Aptasensor for the Single-Step Detection of the SARS-CoV-2 Spike Protein

Efficient and timely testing has taken center stage in the management, control and monitoring of the current COVID-19 pandemic. Simple, rapid, cost-effective diagnostics are needed that can complement current polymerase chain reaction (PCR)-based methods and lateral flow immunoassays. Here, we report the development of an electrochemical sensing platform based on single-walled carbon nanotube screen-printed electrodes (SWCNT-SPEs) functionalized with a redox-tagged DNA aptamer that specifically binds to the receptor binding domain (RBD) of the SARS-CoV-2 spike protein S1 subunit. Single-step, reagentless detection of the S1 protein is achieved through a binding-induced, concentration-dependent folding of the DNA aptamer that reduces the efficiency of the electron transfer process at the electrode surface and causes a suppression of the resulting amperometric signal. This aptasensor is specific for the target S1 protein with a dissociation constant (KD) value of 43 ± 4 nM and a limit of detection (LOD) of 7 nM. We demonstrate that the target S1 protein can be detected both in a buffer solution and in an artificial viral transport medium widely used for the collection of nasopharyngeal swabs, and that no cross-reactivity is observed in the presence of different, non-target viral proteins. This SWCNT-SPE-based format of electrochemical aptasensor may be extended to the detection of other protein targets for which nucleic acid aptamer ligands are made available.