Ultra-sensitive detection of SARS-CoV-2 S1 protein by coupling rolling circle amplification with poly(N-isopropylacrylamide)-based sandwich-type assay

Abstract

In the past few years, the COVID-19 pandemic, caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) seriously threatens global public health security due to its high contagiousness. It remains of vital importance to develop a rapid and sensitive assay for SARS-CoV-2. In this work, we proposed a sandwich-type assay based on poly(N-isopropylacrylamide) (PNIPAM), allowing efficient detection of the SARS-CoV-2 S1 protein in the homogeneous solution. Firstly, a direct sandwich-type assay was established with a linear range of 0.2 − 2 μg/mL and a limit of detection (LOD) of 0.11 μg/mL, which could realize rapid detection in about one hour. Furthermore, the sandwich-type assay coupled with rolling circle amplification (RCA) obtained an increase in sensitivity of 5.9×104 folds with a wide linear range of 0.01 − 100 ng/mL and a LOD of 1.88 pg/mL. The average recoveries in unpretreated saliva were 90% − 113.0%, indicating the potential of the developed method for application in practical samples. Given the high selectivity and sensitivity of the developed method, it has a significant potential for rapid and early detection of SARS-CoV-2.