Single-Molecule Evaluation of the SARS-CoV-2 Nucleocapsid Protein Using Gold Particle-in-a-Frame Nanostructures Enhanced Fluorescent Assay

Abstract

Ultrasensitive evaluation of low-abundance analytes, particularly with limits approaching a single molecule, is a key challenge in the design of an assay for profiling severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antigen. Herein, we report an aptamer claw strategy for directly evaluating the SARS-CoV-2 antigen based on gold particle-in-a-frame nanostructures (Au PIAFs). Au PIAF was used as a metal-enhanced fluorescence material. The assay integrated with a microplate reader achieved a sensitivity of 44 fg·mL–1 in under 3 min and accurately detected the SARS-CoV-2 nucleocapsid protein (N protein) in human saliva samples. When our assay is combined with a single-molecule counting platform, the limit of detection can be as low as 0.84 ag·mL–1. This rapid and ultrasensitive assay holds promise as a tool for screening SARS-CoV-2 and other contagious viruses.

Optical biosensors for diagnosis of COVID-19: nanomaterial-enabled particle strategies for post pandemic era

“…Zhu et al developed an aptamer-based biosensor using in-frame gold particle nanostructures (Au PIAFs) as metal-enhanced fluorescent material to detect SARS-CoV-2 N protein [ 105 ]. Au PIAFs were synthesized with the use of Ag prism seeds in the presence of a mixture consisting of AgNO 3 , sodium citrate, PVP, H 2 O 2 and NaBH 4 , potassium iodide (KI), and hydrogen tetrachloroaurate(III) trihydrate (HAuCl 4 -3H 2 O).…”