Periodic Reporting for period 5 - NanoPD (Single Molecule Nanoscale Sensors for Improved Therapies and Diagnostics)
Período documentado: 2023-10-01 hasta 2024-12-31
One of the most significant challenges facing analytical scientists is the accurate detection and quantification of trace levels of analyte. This can have enormous impact on a local environment, for example, early stage detection of disease, identifying disease markers, drug screening, water quality control, and chemical agents amongst others ultimately come down to the work of analytical scientists. These challenges are compounded when the analysis of real samples or even complex mixtures are required. With this in mind, a currently hotly pursued topic in nanoscale sciences is in the development of single molecule or near single molecule methods to improve quality of life by removing the “clouding” associated with ensemble averaging. Although strategies for detecting single molecules have existed for a number of years, efficient label-free methods without the need for chemical modification are lacking especially when considering smaller molecules directly in unprocessed clinical samples.
As Part of NanoPD we developed a number of nanopore-based systems for multiplexed detection directly in unmodified clinical samples. We have made substantial progress in technological developments, key achievments are as follows:
1. We have developed a strategy for detecting sepsis biomarkers, including PCT and let-7a miRNA, using nanopore sensing and AuNP/DNA molecular probes. Our results demonstrate that this strategy exhibits high sensitivity and selectivity in preclinical samples.
2. We successfully performed multiplexed detection of proteins and nucleic acids from patient samples for different variants of SARS-CoV-29 and Mpox virus. These results were achieved with minimal sample processing and ML-enabled classification using up to three barcoded probes.
3. By integrating dielectrophoretic trapping with nanopore sensing, we have enabled detection down to 5 fM concentrations, up to a 1000-fold improvement in the detection limit compared with existing methods while maintaining efficient throughput.
4. Nanopore sensing has been applied directly to patient samples to detect biomarkers for diseases such as prostate cancer and heart failure. We demonstrated proof-of-concept for the multiplexed detection of miRNAs, proteins, and small molecules, including the simultaneous detection of 40 miRNAs in human serum.
Multiplexed biomarker detection represents a transformative advancement with far-reaching implications for patients, healthcare providers, and society as a whole.
As Part of NanoPD we developed a number of nanopore-based systems for multiplexed detection directly in unmodified clinical samples. We have made substantial progress in technological developments, key achievments are as follows:
1. We have developed a strategy for detecting sepsis biomarkers, including PCT and let-7a miRNA, using nanopore sensing and AuNP/DNA molecular probes. Our results demonstrate that this strategy exhibits high sensitivity and selectivity in preclinical samples.
2. We successfully performed multiplexed detection of proteins and nucleic acids from patient samples for different variants of SARS-CoV-29 and Mpox virus. These results were achieved with minimal sample processing and ML-enabled classification using up to three barcoded probes.
3. By integrating dielectrophoretic trapping with nanopore sensing, we have enabled detection down to 5 fM concentrations, up to a 1000-fold improvement in the detection limit compared with existing methods while maintaining efficient throughput.
4. Nanopore sensing has been applied directly to patient samples to detect biomarkers for diseases such as prostate cancer and heart failure. We demonstrated proof-of-concept for the multiplexed detection of miRNAs, proteins, and small molecules, including the simultaneous detection of 40 miRNAs in human serum.
Multiplexed biomarker detection represents a transformative advancement with far-reaching implications for patients, healthcare providers, and society as a whole.
This project involves the development of a new class of analytical sensors which can be used for diagnosis in neurodegenerative disorders. Ultimately I envisage that the technology will be useful in regulating drug administration. Apart from the clinical relevance, the proposed technology will be invaluable in broadening the fundamental understanding of nanopore sensing, encouraging the construction of a platform capable of detecting multiple chemicals in parallel, which will dramatically expand the value of the device to clinical diagnostics.
To date we have successfully performed the following tasks:
1. Fabrication of nanopore sensors (WP1)
2. Commenced optimization of the recognition chemistry (WP2) including aptamer functionalisation.
3. Performed experiments associated with the detection of alpha-synuclein, Mpox, SARS-CoV-2, miRNA, prostate canceer marker (WP3)
4. Fully developed the electrophoresis platform that can be used with clincial samples (WP4).
5. Extensive work has been performed on detecting biomarkers directly from clinical samples (WP5). Furthermore, the technology has been bencmarked against conventional methods.
The output from this work has been published in high impact journals, a number of patents have also been obtained in relation to this work, collaborations with industrial partners have been setup, and follow on translational funding has been secured.
To date we have successfully performed the following tasks:
1. Fabrication of nanopore sensors (WP1)
2. Commenced optimization of the recognition chemistry (WP2) including aptamer functionalisation.
3. Performed experiments associated with the detection of alpha-synuclein, Mpox, SARS-CoV-2, miRNA, prostate canceer marker (WP3)
4. Fully developed the electrophoresis platform that can be used with clincial samples (WP4).
5. Extensive work has been performed on detecting biomarkers directly from clinical samples (WP5). Furthermore, the technology has been bencmarked against conventional methods.
The output from this work has been published in high impact journals, a number of patents have also been obtained in relation to this work, collaborations with industrial partners have been setup, and follow on translational funding has been secured.
Biosensors are an integral and increasingly important part of modern biomedicine and new developments in biosensor technologies are increasingly seeing their application in the process industry, security, environmental and biodefense application markets. One of the limitations in developing biosensor technology is the coupling of biological recognition to an electronic output. The most successful commercial technologies use enzymatic activities that exploit pH or oxidation state, but these cannot be applied in a generic format as they are highly specific to the enzyme in question, such as glucose oxidase which is commonly used in blood glucose testing. Furthermore, most technologies cannot currently cope with a large abundance of similar molecules often masking the presence of distinct molecular anomalies that are markers in the early stages of disease. Thus the ability to effectively detect specific markers at the molecular level provides new opportunities for analytical measurements in complex systems. While there are some moves to develop nanotechnology approaches to biosensing, these have yet to make a mark and there remains an unmet need in the development of lab-on-chip biosensors that are affordable, integrated, fast, capable of multiplexed detection and monitoring, and highly sensitive which can detect trace levels of analyte. Accordingly, my current proposal aims to define a new nanoanalytical technology that aims to address these key issues and in the process define a new analytical technique for diagnostic and therapeutic applications.