Solid-state nanopore analysis of human genomic DNA shows unaltered global 5-hydroxymethylcytosine content associated with early-stage breast cancer

Improving the Performance of Selective Solid-State Nanopore Sensing Using a Polyhistidine-Tagged Monovalent Streptavidin

“…The reduction was well-described by a semilog fit (linear on a semilog scale, Figure 4b, black line), similar to past results. 34 For the smallest SS-nanopore studied here (7.3 nm), we observed significant transient clogging of the device compared to larger-diameter pores (Supplementary Figure S6), demonstrating that steric hindrance was too strong under that condition to enable smooth translocations. From this, we concluded that 7.5 nm was the minimum diameter possible for assay implementation, correlating with the dimensions of the nucleoprotein complex and with our prior findings.…”

“…Indeed, small differences in diameter can partially account for the variability that has been observed between measurements and can, in principle, be corrected for through a quantitative understanding of its dependence. While pore diameter dependence has been measured for our assay previously, 34 its impact on the improved performance realized with hMS must be determined explicitly. To achieve this, we finally probed DNA, hMS, and DNA−hMS complex using a series of 14 independent SS-nanopores across the diameter range of 7.3 to 23.1 nm.…”

“…The sensitivity and selectivity of our assay depend critically on many experimental parameters that have been explored systematically in prior studies to identify optimal conditions for assay performance. ,, One critical factor determining the dynamics of translocation is the charge associated with both the molecular components and the SS-nanopore itself. We recently investigated the impact of modified salt conditions on our assay, thereby varying charge screening and influencing electrokinetics.…”

The Emerging Role of Circulating Tumor DNA in the Management of Breast Cancer

“…Similarly, Beaver et al reported a sensitivity of 93.3% and specificity of 100% among a small cohort (29 patients) with PIK3CA mutations [17]. Other groups have investigated additional mutations among ctDNA as biomarker targets for the detection of early breast cancer, including 5-Hydroxymethylcytosine, MDM2 and MDM4, and AKT1 [18][19][20].…”

Nanopore-based technologies beyond DNA sequencing

“…These mutations were sensed optically by tagging the probe oligonucleotides with fluorescent dyes and supplementing the electrical sensing of the nanopore with a single-molecule optical detection approach. Solid-state nanopores were also used to quantify global 5-hydroxymethylcytosine epigenetic DNA modifications in human tissue derived from both healthy breast tissue and stage 1 breast tumour tissue 125 . In another recent study, the high selectivity of DNA aptamers was used to fabricate specific DNA 'carriers' with high affinities for specific protein biomarkers in a plasma sample, producing characteristic electric current traces when translocated through a nanopore formed at the end of a glass-pulled pipette 126 .…”