Screening for Group A Streptococcal disease via Solid-State Nanopore Detection of PCR Amplicons



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 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 both for adapting PCR-based assays to solid-state nanopore readout as well as analytical approaches by which to decide on sample status.


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Supplementary material

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Supporting Information document for main manuscript
Document containing the following sections: Section S1: Nanopore signals from PowerUp Sybr Green PCR Master Mix; Section S2: Homebrew PCR optimization process; Section S3: Homebrew PCR mixture event signatures; Section S4: Method of classifying events; Section S5: Signal classifications; Section S6: Classified Event Detection