Human plasma fibrinogen adsorption on ultraflat titanium oxide surfaces studied with atomic force microscopy

Conformational changes of proteins adsorbing on biomaterial surfaces affect biocompatibility. Titanium is among the most successful biomaterials; howe ver, the mechanisms leading to its biocompatibility are not yet understood. The primary objective of this study was to investigate the conformation of human plasma fibrinogen (HPF) adsorbed on titanium oxide surfaces. A metho d for preparing ultraflat titanium substrates was developed. This allowed h igh-resolution investigation of both the titanium oxide surfaces and the ad sorbed state of HPF. The titanium oxide surfaces were first imaged with an atomic force microscope in air and subsequently incubated in a solution of HPF in phosphate buffer and imaged in fluid with tapping mode AFM. The tita nium oxide surfaces exhibited a root-mean squared (RMS) roughness of (0.29 +/- 0.03) nm over (1.00 x 1.00) mu m(2) areas. Different degrees of molecul ar order were found on the titanium oxide Surface. In crystalline surface a reas square lattices with parameters a(0) = b(0) approximate to 0.5 nm were resolved, which is consistent with the (001) planes of the titanium dioxid e TiO2 rutile. Repetitive scanning of TiO2 surface areas resulted in the ab rasion of the outermost TiO2 layers by the AFM tip to a depth varying betwe en 0.3 and 1.0 nm. When imaging adsorbed HPF, individual molecules and aggr egates, often joined through their D domains, were resolved, and the typica l HPF multiglobular structure was observed. The mean length and height of s ingle molecules were (46 +/- 3) nm and (1.4 +/- 0.2) nm, respectively. Addi tional features (chain segments) adjacent to the D and E domains were resol ved and attributed to the a chains and their C-termini. The combination of AFM and the ultraflat titanium preparation method has proven successful for the high-resolution study of both the TiO2 surface and the adsorbed HPF.