Targeted protein adsorption on the surface of biomaterials

Protein adsorption at interfaces has significant importance in the development of materials for applications such as medical devices and biosensors. In this study targeted protein adsorption is investigated. Hydroxyapatite (HA) thin and thick films of ca. 500 nm and 60 μm respectively were fabricated and electrically modified to obtain tailored protein adsorption and to examine their activity. The obtained films were characterised by scanning electron microscopy (SEM) and atomic force microscopy (AFM) to examine their morphology and topography. It is evident from SEM and AFM images that the films possess a highly porous surface with no crack formation. Their chemical composition was analysed by Fourier transform infrared (FTIR) spectroscopy, x-ray diffraction (XRD) and x-ray photoelectron spectroscopy (XPS). The obtained results revealed a surface chemistry with chemical elements of those found in stoichiometry HA. HA thin films were masked by using a transmission electron microscopy (TEM) grid and surface potentials were created by charge injection using the electron beam source found in a SEM. The surface potential domains of the modified HA films were studied using piezoresponse force microscopy (PFM). PFM images confirm the presence of surface potential domains with a charge transition occurring between the exposed and unexposed areas. The HA films with localised electrostatic charges were modified with lysozyme and fibronectin to investigate their activity. Both proteins were preferentially adsorbed onto the created domains. Micrococcus lysodeikticus was used to monitor lysozyme activity on the electrically modified HA films. The immobilised lysozyme was active at different pH values as confirmed by the hydrolysis of Micrococcus lysodeikticus. The influence of fibronectin domains and osteoblast like cells (MC3T3-E1) was evaluated. HA films with patterned fibronectin had lower levels of attached osteoblasts compared to films modified with un-patterned fibronectin. In addition microwave sensing was used to monitor protein binding using SA biotin as a model system. The developed system was successfully used for the label free detection of avidin binding to a biotinylated surface. The limit of detection of the system was examined with various concentrations of avidin on planar and nanoporous gold. While a linear response was obtained for nanoporous surfaces for planar gold the response was not linear. This could be attributed to the higher surface area available on nanoporous gold compared to that in planar gold. Flow measurements were carried out using a flow cell however the results obtained could not detect the specific binding of avidin to biotin under the used conditions (100μL s-1).