Multiplexed Detection of Respiratory Viruses Using Surface Plasmon Resonance Spectroscopy With Ultrathin Carbon Nanomembranes

ABSTRACT The simultaneous circulation of multiple respiratory viruses demands diagnostic platforms capable of rapid, sensitive, and multiplexed detection in complex clinical samples. Here, we present a label‐free, multi‐channel surface plasmon resonance (MC‐SPR) biosensor for simultaneous, real‐time detection of three major respiratory viruses. A molecular 2D material, ∼1 nm thick azide‐terminated carbon nanomembrane (N 3 ‐CNM), is used to functionalize the MC‐SPR sensor surface, enabling covalent antibody immobilization via click chemistry. This approach provides controlled functionalization of multiple channels with virus‐specific antibodies, resulting in robust and reproducible multiplexed detection of the respective antigens. We demonstrate specific detection of SARS‐CoV‐2, Influenza A, and respiratory syncytial virus (RSV) antigens from their mixtures, both in physiological buffer and clinically relevant nasopharyngeal swab matrices, with sensitivity in the pM range, negligible cross‐reactivity, and high reproducibility. Unknown analyte concentrations can be directly determined from the SPR data with high accuracy using the respective calibration curves. This work establishes a robust and versatile platform based on molecular 2D materials for the development of multiplexed biosensors and medical diagnostic applications.