Micro and Nanofluidic Methods for Studying Supramolecular Assembly
Lead Research Organisation:
University of Cambridge
Department Name: Chemistry
Abstract
This project focuses on the development of micro and nanofluidic methods for the study of supramolecular protein assembly. We will aim to identify within a complex mixture proteins and protein complexes by trapping them, using electrophoretic, electrostatic and hydrodynamic forces, at nanojunctions on microfluidic devices fabricated using soft lithographic techniques. This approach will allow the selective enriching of species of interest and will thus allow us to reach very high sensitivities even with dilute analyte solutions, allowing minority populations of misfolded proteins and incorrectly assembled protein complexes to be identified. As part of the project we will develop approaches for effective integration of nano and microfluidic elements on chip. This objective will be achieved through the use of two-photon lithography to overcome the diffraction limit through non-linear optics and to define nanoscale features in the same photoresist used for conventional microfabrication. The lithographically patterned substrate will then be used to imprint polymers in a soft lithography process to yield a scalable avenue to fabricate devices integrating nano and micro scale features. We will detect proteins through deep UV fluorescence, which will allow us to work with fully unlabelled species. These results will open up the path towards quantitative charaterisation of protein self-assembly at high resolution in their native environment.
Organisations
People |
ORCID iD |
| Tuomas Knowles (Primary Supervisor) | |
| Kevin Baumann (Student) |
Studentship Projects
| Project Reference | Relationship | Related To | Start | End | Student Name |
|---|---|---|---|---|---|
| EP/N509620/1 | 30/09/2016 | 29/09/2022 | |||
| 1942963 | Studentship | EP/N509620/1 | 30/09/2017 | 29/09/2020 | Kevin Baumann |
| Description | Proteins are the active molecules in the cell and form the nanoscale machinery of life through self-assembling into complexes. This process underpins biological function and, when assembly doesn't occur in the intended manner, is also the origin of the onset and progression of a number of human disorders, including protein aggregation diseases such as Alzheimer's and Parkinson's diseases. The proteins associated with such diseases are generally only available at inherently low concentrations. Therefore, the detection of these proteins as well as other biomolecules requires the development of assays with high sensitivity, low cost, and minimal sample consumption. We have been working towards an advanced microfluidic platform to selectively trap individual proteins and their complexes. Microfluidic techniques offer the unique benefit of enclosing smallest amounts of samples in micrometre-dimensioned volumes. Reliably trapping biomolecules of interest in their native solution has the potential to open up a new view into biomolecular behaviour. The advantage of the microfluidic platform is the possibility of the direct observation of biomolecular interactions in real time and on single molecule level. To controllably trap biomolecules and effectively enhance their local concentrations, electric methods provide a promising approach as biomolecules can respond to the electric fields by moving or orienting. In the experiments we have conducted, we show successful trapping of a range of common biomolecules, such as DNA, Insulin, Bovine Serum Albumin and the Alzheimer's associated Tau protein by balancing sample and dispersant flows induced by the applied electric forces. We observed these trapping events with conventional fluorescence microscopy as well as single molecule detection methods. Next steps include the analysis of mixtures of analytes, bench-marking the applicability of this technique and how these analytes can be examined quantitatively. |
| Exploitation Route | The trapping platform allows controllable trapping of biologically relevant molecules in a straight-forward approach and by using standard optical detection methods. For a range of diseases, low levels of the associated biomolecules are present in body fluids before the full onset of the disease. The advantage of increasing the concentration in a solution locally and enhancing the signal represents an essential requirement for early detection of diseases associated with these low levels of DNA or proteins. Early detection of diseases is essential for improvement of healthcare and lowering costs. The high-throughput production of the used devices and the ease of handling can pave the way toward large-scale, multiplexed analysis of biomarkers present in low quantities. |
| Sectors | Healthcare Pharmaceuticals and Medical Biotechnology |