The loosely and firmly adsorbed amount of bovine serum albumin (BSA) on dental titanium and dental gold was investigated by bicinchoninic acid assay (BCA assay). This method does not require special properties of the sample like a high reflectivity or conductivity and turns the BCA assay into a valuable tool to investigate a widespread spectrum of different substrate materials, e.g., natural enamel. The experiments in this work showed that it is possible to distinguish between loosely and firmly bound protein with the BCA assay. On the here investigated dental titanium and dental gold samples the amount of adsorbed BSA was dominated by the loosely bound part. On dental gold the total amount of adsorbed BSA was significantly higher than on titanium. The amount of loosely bound BSA was high near the isoelectric points of dental titanium and dental gold while the amount of firmly adsorbed BSA on dental titanium followed the electrostatics. At high pH values (pH 9.0) conformational changes of the BSA molecules played a key role in the adsorption process.
Plain and microstructured cp-titanium samples were studied as possible biofilm reactor substrates. The biofilms were grown by exposition of the titanium samples to bacteria in a flow cell. As bacteria the rod shaped gram negative Pseudomonas fluorescens and the spherical gram negative Paracoccus seriniphilus were chosen. Afterward, the samples were cleaned in subsequent steps: First, with a standard solvent based cleaning procedure with acetone, isopropanol, and ultrapure water and second by oxygen plasma sputtering. It will be demonstrated by means of x-ray photoelectron spectroscopy, fluorescence microscopy, and confocal laser scanning microscopy that oxygen plasma cleaning is a necessary and reliant tool to fully clean and restore titanium surfaces contaminated with a biofilm. The microstructured surfaces act beneficial to biofilm growth, while still being fully restorable after biofilm contamination. Scanning electron microscopy images additionally show, that the plasma process does not affect the microstructures. The presented data show the importance of the cleaning procedure. Just using solvents does not remove the biofilm and all its components reliably while a cleaning process by oxygen plasma regenerates the surfaces.
The total amount of albumin and lysozyme adsorbed on different dentally relevant materials like titanium, gold, ceramics, poly(methyl methacrylate) (PMMA) and polytetrafluoroethylene (PTFE) was measured by bicinchoninic acid assay (BCA assay) after detachment of the proteins from the samples. The total amount of protein was separated into the amount of loosely and firmly adsorbed protein. The parts of loosely and firmly bound protein differ remarkably depending on the material. For albumin, the loosely bound protein dominates the total adsorption. In contrast, for lysozyme the amount of loosely and firmly bound protein is in some cases equal and sometimes there is more firmly than loosely bound lysozyme. In general, the firmly bound protein masks the electrostatics of the surface. The amount of loosely bound protein follows no strict tendency, whereas the amount of firmly bound protein is mostly governed by entropic and hydrophobic interactions, but modulated by electrostatics and thus by the isoelectric points of the surface and of the proteins. Most protein adsorbs on the metallic surfaces, if compared to the investigated non-metallic surfaces. For the adsorption on gold, the number of cysteines in the amino acid sequence influences the amount of protein on the gold surface. Least albumin adsorbs on PMMA, least lysozyme adsorbs on PMMA and PTFE. For albumin, pH depending conformational changes play a key role for the adsorption at pH 9.0.
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