Highly sensitive and label-free detection of SARS-CoV-2 proteins via surface plasmon resonance using biofunctionalization with 1 nm thick carbon nanomembranes

Here we report a novel platform for the detection of nucleocapsid (N) and receptor-binding domain (RBD) of spike (S) proteins of SARS-CoV-2 viruses using the surface plasmon resonance (SPR) technique. We demonstrate that the functionalization of SPR sensors with molecular 2D materials - 1 nm thick carbon nanomembranes (CNMs) significantly enhances sensitivity. CNMs terminated with azide linker (N₃-CNM) enable covalent bonding of SARS-CoV-2 antibodies for specific immobilization of the N- and S-proteins to the sensor surface. The successful and stable hierarchical functionalization is confirmed by multiparametric SPR measurements complemented with X-ray photoelectron spectroscopy and polarization modulation infrared reflection absorption spectroscopy. The obtained equilibrium dissociation constants (K_D) for the N-protein and the S-protein in the physiological buffer are 570 ± 50 pM and 22 ± 2 pM and the low detection limits (LODs) are ~ 190 pM and ~ 10 pM, respectively. The high specificity of the developed sensors is shown via their negligible cross-reactivity with SARS-CoV-1 and MERS-CoV proteins. Finally, detection of SARS-CoV-2 proteins in nasopharyngeal swab samples with the LOD of ~ 40 pM is demonstrated. The proposed methodology enables the development of biosensors that cover clinically relevant range for the direct and immediate detection of SARS-CoV-2 without any amplification or labeling.