Detection of parvovirus B19 genomic fragments using an electrochemical biosensor based on argonaute-assisted silver metallization

Abstract

The human pathogen Parvovirus B19 (B19V) is known to cause a spectrum of diseases. Presently, no vaccines or antiviral medications are accessible for its prevention or cure. This highlights the pressing need for the advancement of efficient and affordable diagnostic technologies in vitro, which are essential for managing and curbing the spread of infectious diseases. In our research, we have introduced an innovative nanocomposite material, g-C₃N₄/DWCNT-COOH/PtNPs, synthesized via a one-step hydrothermal process. This material is applied on the electrode surface to enhance the primary signal. The capture probe cDNA is affixed to the electrode via a Pt-S bond, a method that ensures a stable immobilization for detection purposes, and in conjunction with nucleic acid detection mediated by Clostridium butyricum Argonaute (CbAgo) from the mesophilic bacterium, an electrochemical Argonaute biosensor utilizing silver metallization was constructed, targeting the non-structure protein 1 (NS1) region of the virus genome. Benefiting from the lack of sequence constraints similar to the Protospacer Adjacent Motif (PAM), CbAgo can utilize an easily designed and cost-effective guide DNA (gDNA) to recognize the target sequence. By incorporating a polymerase chain reaction (PCR) pre-amplification step, along with silver metallization for secondary signal amplification and electrochemical analysis techniques, a highly sensitive and specific detection is obtained. The biosensing platform exhibits a detection range from 10^-17 M to 10^-12 M, with a detection limit as low as 2.62 aM. Compared to current detection methods for this virus, it demonstrates superior analytical performance and validates its effectiveness for use with real serum samples.