Direct Visualization of Single-Molecule Protein Dynamics on Silica Nanoparticle Coating Surfaces Using High-Speed Atomic Force Microscopy

Controlling protein adsorption at the material-biological interface is essential in many biomedical applications, including blood-contacting implantable medical devices, biosensors, microfluidic devices, protein purification and diagnostics assays. Nonspecific protein adsorption is rapid and may trigger other unfavourable biological interactions. This is particularly life-threatening for material surfaces in contact with blood, often resulting in inflammation, blood coagulation, and thrombosis. The protein adsorption phenomenon is shared across different fields, and a plethora of studies have provided an immense understanding of protein-surface interactions, yet the fundamental microscopic details of this process are still lacking. Most current models for protein adsorption are based on macroscopic or bulk averaged observations and neglect the microscopic (single protein) dynamics, making it difficult to determine the fundamental protein-surface interactions. That said, directly observing single protein dynamics on material surfaces is very challenging. In particular, studies are often undertaken on materials with inherently rough, opaque or fluorescent quenching surface properties that are not amenable to high-resolution optical/fluorescence techniques for imaging single molecule dynamics. This is where the recent emergence of High-Speed Atomic Force Microscopy (HS-AFM) is providing exciting opportunities for visualising single protein dynamics on surfaces. In this thesis, HS-AFM was utilised for visualising single molecule fibrinogen (FG) and bovine serum albumin (BSA) protein adsorption onto muscovite mica substrates and silica nanoparticle (SiNP) coatings to investigate the fundamental structural dynamics of individual proteins during adsorption. Further, quartz crystal microbalance with dissipation monitoring (QCM-D) was utilised to investigate the bulk protein adsorption properties on SiNP coatings and attempts made to rationalize these bulk adsorption characteristics using the single molecule observations acquired from HS-AFM.