An optics based approach to next generation mass photometry

Thesis

Scattering-based optical microscopy techniques have undergone a significant improvement in sensitivity over the last few decades, recently reaching the single-molecule level.

Mass photometry (MP) is a label-free single-molecule optical microscopy technique, based on the detection of the interference between the light scattered by a protein and the light reflected from a glass-water interface. MP has found rapid adoption to study the stoichiometry of proteins and protein complexes, in order to quantify the kinetics and thermodynamics of protein-protein interactions.

There are four main aims for advancing MP, which are; improving the mass-resolution, lowering the detection limit, increasing the concentration range and extracting more information from an experiment. In this thesis, novel optical designs are introduced with the aim of improving these parameters and understanding the current limitations of the technique.

A minimalistic optical design called inverse mask MP is introduced, which demonstrates an improvement on the state-of-the-art level mass-resolution and detection limit. Furthermore, novel experiments which provide additional structural information on the sample are presented. A major benefit of this design is that it enables the construction and optimisation of more complex MP instruments. This concept is demonstrated by the development of a four-colour illumination, four-colour detection MP instrument, with the aim of further pushing the performance boundaries of MP.

Authors: Peters, JS

• DOI: 10.5287/ora-bp8vvkjez
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: mass photometry; interferometric scattering microscopy; label free; single molecule
• Subjects: Microscopy; Biophysics