Aptamer-based sensing of disease biomarkers with nanopores

Nanopore sensing have demonstrated great potential for biosensing given its exceptional sensitivity at the single-molecule level which enables direct detection of individual proteins and other biomolecules. Nanopore based sensing have been successfully applied to the sensing of proteins, RNA, DNA and other related targets. In particular biological nanopores have achieved huge success in direct sequencing and identification of disease-causing pathogens such as viruses and bacteria. However, in many clinical settings, full sequencing of the pathogen is not needed for positive identification. Instead, direct detection of the specific biomarkers such as the presence of disease-associated proteins can provide sufficient information for diagnosis and for the treatment to begin promptly. In principle the entire process can be done in less than an hour, enabling rapid detection of diseases in clinical settings. Our group have previously demonstrated several relevant proof-of-approaches including CRISPR-based DNA fingerprinting, aptamer capture and sensing using nanopores and other related techniques (see attached literature). We propose extending this proof of concepts experiments towards real world application using patient samples for a range of infectious disease. Platform/idea: The proposed sensing concept uses DNA carriers functionalized with aptamer binding sites in order to ‘fish’ disease related proteins from serological samples. Following target capture, the DNA carriers are then translocated through a solid state nanopore. During the passage of the DNA carrier, the flow of ions is blocked, thereby generating a current blockage signature that can positively identify the captured proteins during its passage through the nanopore. A key feature of this proposed concept is its versatility. By employing different pre-prepared DNA carriers with different aptamer binding sites, we can target and capture different sensing targets – all while using the same nanopore device. This enables repeated use and a high user-programmability.

Identification of relevant proteins and biomarkers. Search and screen aptamers for binding efficiency and selectivity. Design and production of DNA carriers. Preliminary experiments with spiked samples. Extend sensing and detection of patient derived samples. Optimised clean up and protocols for recovery of nanopores for repeated use.