Protein Adsorption On Poly(N-vinylpyrrolidone)-Modified Surfaces

Protein adsorption is believed to be the initial event when a material comes into contact with a biological environment. The properties of the adsorbed protein layer, including the type, amount, and conformation of adsorbed proteins, determine the biocompatibility and biofunctionality of the material. Therefore, investigation of the interactions between proteins and biomaterials and control over these interactions is a key consideration in developing improved materials.Since it was first synthesized in1938, poly(N-vinylpyrrolidone)(PVP) has been widely used in the biological, pharmaceutical, food, materials, and other fields due to its high water solubility and reasonably good biocompatibility. The main work of this thesis is the development of PVP-modified surfaces and the systematic investigation of their protein adsorption behavior. The relationship between the conformation and bioactivity of lysozyme adsorbed on gold chips modified using different chemistries was also studied. The work has four major components:1. Two surfaces were formed by the self-assembly of (a) octadecanethiol (hydrophobic) and (b) thiolated PEG (hydrophilic) on gold chips (Au-C18and Au-PEG, respectively). The adsorption of a model protein, lysozyme, on the modified surfaces was evaluated using fluorescein isothiocyanate (FITC) labelling. The specific activity of lysozyme adsorbed on the modified surfaces was measured using a fluorescence-based EnzChek(?) Lysozyme Assay Kit. Conformational changes of adsorbed lysozyme were investigated using an improved SERS technique. It was shown that changes in bioactivity were well correlated with conformational changes: the smaller the conformational change the greater the specific activity of the adsorbed lysozyme. The improved SERS technique provides a simple and effective method for exploring the conformation of proteins adsorbed on material surfaces.2. PVP-modified surfaces were prepared by surface-initiated atom transfer radical polymerization (SI-ATRP) and the interactions between various proteins (especially plasma proteins) and PVP-modified surfaces were investigated using radiolabelling methods. It was shown that the PVP-modified surfaces can repel most plasma proteins, especially those that are important in blood coagulation (e.g. fibrinogen). At the same time, the modified surfaces showed preferential adsorption of albumin, a desirable property since albumin layers are generally found to be anti-biofouling. The excellent protein resistance of the PVP-modified surfaces may make them interesting candidates for blood contact applications.3. Poly(N-vinylpyrrolidone)-block-polystyrene (PVP-b-PS) brushes were prepared via surface-initiated consecutive ATRP on initiator-immobilized surfaces and their interactions with proteins and cells were investigated. The quantities of adsorbed proteins (lysozyme, fibrinogen, and vitronectin) were different on the PVP-b-PS modified surfaces compared to the PVP-modified surfaces. However, on addition of the PS segment to the initially tethered PVP, the surface showed increased cell adhesion and spreading. Thus it appears to be possible to regulate protein adsorption and cell adhesion by tuning the chemical compositions of tethered diblock copolymers.The influence of chemical composition and microphase separation behavior of tethered diblock copolymers on protein adsorption and cell adhesion was also investigated. PVP-b-PS brushes and PS-b-PVP brushes were prepared via surface-initiated consecutive ATRP on initiator-immobilized surfaces. It was found that protein adsorption and cell adhesion can be regulated by precise control of surface properties such as the distribution of hydrophilic and hydrophobic domains, surface chemical domains etc.4. Well-controlled polymerization of N-vinylpyrrolidone (NVP) on gold surfaces by SI-ATRP was carried out at room temperature by a silanization method. The modified gold surfaces showed excellent thermal stability as suggested by reflectance FTIR spectra. Thus, post-functionalization of polymer brushes on gold surfaces at elevated temperatures is possible. Moreover, data from radiolabelled protein experiments showed that the PVP-modified surfaces have excellent protein resistance.This method was also used to modify SPR chips. Protein adsorption on PVP-modified surfaces was monitored in real time by SPR and the results were in accord with the radiolabelling data. The silanization method thus shows potential for applications in biosensors and biochips.In conclusion, surface chemical modification is one of the efficient methods of regulating protein adsorption and cell adhesion and improving biocompatibility of material surfaces. Our work provides multiple ways of designing new biomaterial surfaces and has potential applications in tissue engineering, biosensors and biochips.