Improving the Performance of Selective Solid-State Nanopore Sensing Using a Polyhistidine-Tagged Monovalent Streptavidin.
ACS Sens
; 9(3): 1602-1610, 2024 03 22.
Article
en En
| MEDLINE
| ID: mdl-38451864
ABSTRACT
Solid-state (SS-) nanopore sensing has gained tremendous attention in recent years, but it has been constrained by its intrinsic lack of selectivity. To address this, we previously established a novel SS-nanopore assay that produces translocation signals only when a target biotinylated nucleic acid fragment binds to monovalent streptavidin (MS), a protein variant with a single high-affinity biotin-binding domain. While this approach has enabled selective quantification of diverse nucleic acid biomarkers, sensitivity enhancements are needed to improve the detection of low-abundance translational targets. Because the translocation dynamics that determine assay efficacy are largely governed by constituent charge characteristics, we here incorporate a polyhistidine-tagged MS (hMS) to alter the component detectability. We investigate the effects of buffer pH, salt concentration, and SS-nanopore diameter on the performance with the alternate reagent, achieve significant improvements in measurement sensitivity and selectivity, and expand the range of device dimensions viable for the assay. We used this improvement to detect as little as 1 nM miRNA spiked into human plasma. Overall, our findings improve the potential for broader applications of SS-nanopores in the quantitative analyses of molecular biomarkers.
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1
Base de datos:
MEDLINE
Asunto principal:
Ácidos Nucleicos
/
Nanoporos
/
Histidina
Idioma:
En
Revista:
ACS Sens
Año:
2024
Tipo del documento:
Article