Hybridization chain reaction circuit-based electrochemiluminescent biosensor for SARS-cov-2 RdRp gene assay

In this work, we designed an ECL ratiometric biosensor with a three-stranded Y-type DNA (Y-DNA) probe and induced a hybridization chain reaction (HCR) for the highly sensitive detection of SARS-CoV-2 nucleic acid. The important component of this system is the self-assembled Y-Shaped probe based on three nucleic acids. Y1, Y2, and Y3 can be linked by complementary base pairing to Hairpin1 (H1), Hairpin2 (H2), and Ru modified DNA (Ru1), respectively. H1 and H2 can trigger the HCR reaction when activated by the SARS-CoV-2 RdRp gene and the 5′ end of Ru1. The 5′ end of Ru1 is modified with the Ru complex, which can produce a strong electrochemiluminescence luminescence signal at 620 nm under an applied voltage. Through the amplification of Y-DNA-induced HCR reaction, Ru1 on the electrode surface gradually increased, the ECL signal at 460 nm was gradually quenched, and the signal at 620 nm was steadily generated. The SARS-CoV-2 RdRp gene can be quantified according to the degree of decrease of ECL signal at 460 nm and the increase of ECL signal at 620 nm. Combining the two signal amplification strategies, this ratiometric ECL biosensor can accurately and efficiently detect the target gene with a detection limit of 59 aM. • An HCR based ratiometric ECL biosensor was fabricated based on a Y- DNA. • Y-DNA provides a nucleic acid scaffold for the amplification reaction on the electrode surface. • The biosensor was employed for the detection of SARS-CoV-2 RdRp gene with a LOD of 59 aM. • The method avoids the problem of enzyme inactivation in PCR. • The biosensor can be used for the assay target DNA in human Pharyngeal swabs.