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Nanopore sensing for environmental and biomarker analysis
Title
Nanopore sensing for environmental and biomarker analysis
Creator
Arora, Pearl
Date
2024
Description
Nanopore stochastic sensing is a powerful analytical tool for detecting target molecules through a nanoscale pore. The analyte and electrolyte...
Show moreNanopore stochastic sensing is a powerful analytical tool for detecting target molecules through a nanoscale pore. The analyte and electrolyte ions are subjected to a voltage bias which drives them to translocate through the nanopore, resulting in disruptions in the ionic current. These disruptions are translated to blockage events which can serve as a signature of the analyte. Owing to its unique features of single-molecule and label-free sensing, nanopore technique has been exploited in a wide array of applications such as detection of metal ions, proteins, DNA, microRNA, toxic agents etc. In this dissertation, projects showcasing nanopore’s sensing capability of different biomarkers and in the detection of a wide range of target molecules based on non-covalent interactions are presented. Particularly in the first two projects, nanopore detection of ferric ions relevant to environmental regulation as well as a biomarker for human health and a miRNA-based biomarker for oral cancer and oral related diseases are summarized. Ferric ions, which are benign if present in balanced quantities but can be toxic otherwise, are detected by using an engineered multifunctional nanopore and a chelating organophosphonic acid ligand. The chelate complex formed after ferric ions bind to ligand gives significantly different event signatures than the free ligand in the solution enabling ferric ion detection. Even in the presence of interfering ions, the ferric ions could be recognized easily because of the conformational changes brought in the nanopore lumen by the interaction of the interfering metal ions with the His-tags of the nanopore which in turn resulted in variations in the characteristics of blocking events. In the second project, miR31, an oral cancer biomarker, is selectively detected with the help of an engineered nanopore, and a DNA based probe. Several probes with variations in length, composition and position of the overhangs or probes with no overhangs were compared and studied as the probes play a crucial role in capturing the target of interest with high specificity. Our strategically designed probe emerged as the most effective in capturing the target even in presence of large background from human saliva samples and enhanced the sensitivity of the system. In the first two projects, nanopores are utilized for selective and specific detection of certain target molecules. However, in order to analyze diverse range of analytes, numerous sensing systems have to be constructed which can be a time-consuming and challenging task. To circumvent this limitation, in the third project, diverse recognition sites based on various non-covalent interactions are incorporated into the α-hemolysin protein pore to achieve detection of not just a single analyte but broad category of molecules such as cations, anions, aromatic and hydrophobic compounds.
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