Proteins at surfaces

Understanding protein adsorption is of vital importance in many fields of medicine and industry that can be divided into two categories: those in which it is desired to minimize adsorption, and those in which protein adsorption is desired. The first category covers materials for kidney dialysis membranes, implants in the bloodstream, vessels for storing pharmaceutical products, marine antifouling paints, etc. The second covers mainly materials for surgical implants, for example, bone replacement, teeth implants, which must be assimilated with the living tissue. In between these two categories lies a work to understand the fundamental processes of protein adsorption onto solid surfaces. Proteins have colloidal type of interactions, with the solid surfaces and between each other, due to their large size. The electrostatic interaction is an important driving force for protein adsorption onto hydrophilic solid surfaces that was thoroughly investigated in this thesis. Furthermore, proteins have additional complexity because they are largely inhomogeneous in their charge, shape, interactions and they can change their conformation upon adsorption. In order to characterize and predict protein adsorption, the information about adsorption kinetics in situ, conformation of adsorbed proteins and the physical parameters describing the adsorbed protein layer are required. All this information can be obtained using different experimental techniques. When combined, such information can answer some questions about the mechanism of protein adsorption onto surfaces. In the present thesis several experimental techniques were applied to study the protein adsorption on planar surfaces. Attenuated total internal reflection Fourier transform infrared spectroscopy is used to determine and follow the protein structural changes during adsorption and under different experimental conditions. Attenuated total internal reflection Fourier transform infrared spectroscopy also allows the protein adsorbed mass as a function of time to be measured. Obtained protein adsorption curves were fitted by a kinetic model proposed, which takes into account the possibility of two protein adsorption states with different orientations, side- and end-on, and transitions between them. The architecture, density profile, of the adsorbed protein layer and adsorbed amount was studied by neutron reflectometry and gave a consistent results with attenuated total internal reflection Fourier transform infrared spectroscopy measurements. Finally, a new thin gap technique was developed to follow the kinetics of protein adsorption using proteins labelled by 125I. It also allowed to study the protein exchangeability by exchange experiments with nonlabelled proteins. First results of this new technique are discussed