Nanopores have been proved to be promising analytical platforms for label-free single-molecule biosensing. However, the practical application of nanopores, such as early stage diagnosis tools or the DNA sequencing tools, is restricted by several functional defects. The main aim of this work is to explore multiple approaches to improve the sensitivity and selectivity of a nanopipette based nanopore sensing, by a combination of field-effect transistors or molecular carriers.
In the first part of the thesis, a new class of nanoscale sensors – dubbed a nanopore extended field-effect transistor (nexFET) – is fabricated to control and selectively sense the biomolecules. Through real-time feedback controlling electrodeposition, a tuneable polypyrrole nanopore can be formed on a carbon-coated double-barrel nanopipette tip with one barrel filled with carbon nanoelectrodes. The generated nexFET displays outstanding FET performance and pH sensitivity. Combining the advantages of nanopore single-molecule sensing, field-effect transistors, and recognition chemistry, the nexFETs are subsequently employed to probe DNA and protein molecules. By controlling the gate voltage of the nexFET, the transport of DNA can be improved comprehensively; for example, a higher throughput, a longer dwell time, and a higher signal-to-noise ratio can be achieved. The selectivity of the nexFET can be further improved by functionalising with embedded receptors.
Attention is then focussed to explore novel strategies for selective biomolecular sensing using nanopipettes, and molecular carriers and probes. Here, I design probes based on the dimerisation of gold nanoparticles (NP), which can be used for the detection of biological targets. Two strategies are employed: 1) aptamer-modified NP dimers are used to target biomarkers, and 2) antibody-modified NP monomers dimerise and self-assemble based on the binding of an antigen target. Importantly, we demonstrate that nanopore sensing can be utilised with both strategies to quantify bound analytes from unbound ones and, in the process, perform single-molecule binding assays.
In summary, multiple approaches including functionalised nanopores, novel molecular carriers, and novel molecular probes were designed and developed. As they can address various limitations of nanopore-based biosensors, these improved methods are expected to pave the way for the practical application of nanopores in early diagnostics and therapeutics.