Advances in biophysical methods for protein detection and characterisation

Proteins are the building blocs of life and mediate nearly every function in the cell. They are therefore a major and incredibly wide research topic. Their functions and malfunctions have serious impacts on a wide range of diseases. Proteins may be used as extremely versatile tools for biology such as for gene editing or biological medical products. Developing novel methods for protein detection and characterisation may thus have a tremendous impact on modern medicine and research. The present thesis discusses advances in biophysical methods for protein detection and characterisation. First, the possibility of detecting proteins label-free is addressed. A label may change the behaviour of the target protein. Two approaches are investigated: An ultraviolet light based autofluorescence microscope is described; and scattering based detection is explored by expending on the existing interferometric scattering (iSCAT) technique. An oblique illumination approach helps with increasing the contrast of the data, and a time correlation technique is used for local sizing on chip. Second, microfluidic techniques are routinely used to create protein assays. These assays minimise the amount of sample required and the absence of turbulences enable new techniques. A method to easily add nanofluidics elements to microfluidic designs is discussed. Finally, three characterisation methods are described. First, diffusional sizing uses a microfluidic chip to create a concentration gradient. The protein diffusion coefficient is extracted from the time evolution of this gradient. Second, the diffusiophoretic coefficient of the protein can be extracted by diffusiophoresis, which is the motion of proteins driven by the concentration gradient of another solute. This could be an important protein motion mechanism, as many gradients are present in cells and in living beings. Finally, the spatial propagation of the protein amyloid-beta 1-42, a protein associated to neurodegenerative disorders, is observed in a capillary.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2021

Author dc:creator

• Peter, Quentin

Advisor dc:contributor.advisor

• Knowles, Tuomas

• microfluidics
• label-free detection
• characterisation

Licence

• https://www.repository.cam.ac.uk/bitstreams/fd49cb3b-e4cf-440e-a1aa-42931ebc64eb/download
• https://creativecommons.org/licenses/by-sa/4.0/

Language dc:language

eng

DOI dc:identifier.doi

https://doi.org/10.17863/CAM.74582

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/327133