Single-molecule protein profiling using nanopores and dimeric aptamer-modified DNA carriers
Author(s)
Wang, Xiaoyi
Yaxian, Liu
Ren, Ren
Edel, Joshua
Ivanov, Aleksandar
Type
Journal Article
Abstract
Single-molecule protein profiling offers unparalleled sensitivity for detecting rare biomarkers, with
significant potential for early disease diagnosis and monitoring. Nanopore sensors enable the detection of proteins in solution and are well-suited for clinical applications due to their high sensitivity and throughput. Here, we introduce a nanopore-based detection strategy utilising an aptamer-protein-aptamer sandwich structure to enhance protein detection further. The aptamer probe is extended with a double-stranded DNA carrier to enhance protein transport and detection efficiency. Upon protein binding, dimerization of the DNA carriers occurs, allowing charge-based classification of bound versus unbound states. Additionally, the sandwich complex produces a characteristic subpeak within individual nanopore events, with subpeak height correlating to protein size. This approach enables selective detection of dimeric proteins, including vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF), with sub-picomolar detection limits. In principle, it can be adapted for any protein with two distinct aptamer-binding sites, as demonstrated with thrombin. This method also facilitates real-time monitoring of ligand-induced receptor dimerization in complex biological fluids, offering a powerful tool for potential single-molecule-based diagnostics.
significant potential for early disease diagnosis and monitoring. Nanopore sensors enable the detection of proteins in solution and are well-suited for clinical applications due to their high sensitivity and throughput. Here, we introduce a nanopore-based detection strategy utilising an aptamer-protein-aptamer sandwich structure to enhance protein detection further. The aptamer probe is extended with a double-stranded DNA carrier to enhance protein transport and detection efficiency. Upon protein binding, dimerization of the DNA carriers occurs, allowing charge-based classification of bound versus unbound states. Additionally, the sandwich complex produces a characteristic subpeak within individual nanopore events, with subpeak height correlating to protein size. This approach enables selective detection of dimeric proteins, including vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF), with sub-picomolar detection limits. In principle, it can be adapted for any protein with two distinct aptamer-binding sites, as demonstrated with thrombin. This method also facilitates real-time monitoring of ligand-induced receptor dimerization in complex biological fluids, offering a powerful tool for potential single-molecule-based diagnostics.
Date Issued
2025-06-29
Date Acceptance
2025-06-20
Citation
Angewandte Chemie International Edition, 2025
ISSN
1433-7851
Publisher
Wiley
Journal / Book Title
Angewandte Chemie International Edition
Copyright Statement
© 2025 The Author(s). Angewandte Chemie published by Wiley-VCH GmbH This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
License URL
Publication Status
Published online
Article Number
e202505902
Date Publish Online
2025-06-29