Molecular Crowding Modulates SARS‐CoV‐2 Aptamer Affinity

SARS‐CoV‐2 aptamer is a favorable candidate for the recognition and detection of SARS‐CoV‐2, owing to its small size and easy synthesis. However, the issue of compromised binding affinities in real samples and targeting mutant SARS‐CoV‐2 hinder wide applications of the aptamer. In this study, it is discovered that molecular crowding could increase binding affinity of CoV2‐6C3 aptamer against RBD (Receptor Binding Domain) of SARS‐CoV‐2 via increasing the absolute value of the enthalpy change. The values of the equilibrium dissociation constant in molecular crowding decrease by 70% and 150%, respectively, against wild‐type and mutant RBD compared with those in buffer without crowding. Moreover, the detection limit of SARS‐CoV‐2 pseudovirus is up to 5 times lower under molecular crowding compared to dilute conditions. The discovery deepens the understanding of aptamer‐target interaction mechanisms in crowding conditions and provides an effective way to apply SARS‐CoV‐2 aptamer for virus recognition and detection.