Diffusing colloidal probe measurements of nanoparticle-surface and protein-carbohydrate interactions.

摘要

This dissertation presents measurements of potentials of mean force between nanoparticles (and colloids) and surfaces with and without adsorbed biomacromolecules toward measuring weak single protein interactions on the order of kBT. The measurements presented utilize Diffusing Colloidal Probe Microscopy (DCPM) to track particles in 3D in real time as they experience Brownian motion above a planar surface. By tracking particles to gather statistical sampling of the Brownian motion I measure particle-wall potential energy interactions, as well as, particle dynamics. Results from experimental measurements are verified by fits to theoretical predictions and Brownian Dynamics simulations with no adjustable parameters.;First, DCPM measurements of bare gold nanoparticles (AuNPs) with diameters of 50nm, 100nm, and 250nm electrostatically confined in sub-micron gaps between silica coverslips are shown. Results demonstrate excellent agreement with theoretical predictions for potential energy interactions while diffusion measurements demonstrate electroviscous drag due to overlap of large electrostatic double layers. Also presented are measurements of electrostatically confined multi-walled carbon nanotubes (mwCNT) where effects of pH and ionic strength on potential energy profiles, diffusion, and stability are studied. Using DCPM in these nanoparticle/mwCNT experiments shows the first measurements of their kind where particle scattering is used to continuously track sub-diffraction limit sized particles and quantitatively measure potentials of mean force and diffusion.;Next, measurements at physiological ionic strengths with 100nm AuNPs where the particles and confining surfaces are functionalized with bovine serum albumin for non-specific protein-protein interactions are presented. These results show the ability to account for all system parameters including CCD camera exposure time and system noise through simulations to directly compare experimental findings to theoretical predictions. These measurements demonstrate the first DCPM measurements of non-specific protein-protein interactions using AuNPs. Finally, measurements of competitive protein-carbohydrate interactions measured by DCPM using micron-sized silica colloids are presented. These results show quantitative measurements of specific protein interactions where changes in particle motion above the planar surface can be directly correlated to protein interactions mediated by competing ligand. These results provide the basis for scaling down to measurements with gold nanoparticles where weaker binding events can be observed leading to future measurements of single protein-receptor interactions.