Single-molecule detection methods are becoming increasingly important for diagnostic applications. Practical early detection of disease requires sensitivity down to the level of single copies of the targeted biomarkers. Of the candidate technologies that can address this need, solid-state nanopores show great promise as digital sensors for single-molecule detection. Here, we present work detailing the use of solid-state nanopores as downstream sensors for a polymerase chain reaction (PCR)-based assay targeting group A streptococcus (strep A), which can be readily extended to detect any pathogen that can be identified with a short nucleic acid sequence. We demonstrate that with some simple modifications to the standard PCR reaction mixture, nanopores can be used to reliably identify strep A in clinical samples. We also discuss methodological best practices for both adapting PCR-based assays to solid-state nanopore readout and analytical approaches by which to decide on sample status.
Keywords: Group A Streptococcus; PCR Amplicons; digital sensors; nanopore; single-molecule detection.