Experimental approach to a biological characterization of materials

Abstract

Titanium oxide surfaces with thin (4 to 5 nm) or thick (30–40 nm) TiO2 layers and rough (Rrms 1.9 μm) or smooth (Rrms 0.45 μm) surface structure were exposed to capillary blood for 5 s through 64 min. The adsorption of main plasma proteins (albumin, IgG, fibrinogen), binding of cascade enzymes (serine proteases) and adhesion and activation of blood cells was measured with immunofluorescence techniques using specific antibodies, the binding of which was quantitated by computer-aided image analysis. The activation of cascade enzymes was detected with antibodies against prothrombin/thrombin (terminal step in the coagulation cascade), C1q and C3c (initial complement activation), terminal complement complex (TCC), and plasmin (terminal step in fibrinolysis). The activation of cells was measured by expression of selectin (platelet CD62), expression of integrins (neutrophil CD11b), or respiratory burst (NBT test) of peritoneal leucocytes adhering to the surfaces after in vivo exposure. The results show different profiles of protein adsorption at the four surfaces. Fibrinogen was the dominating protein on all surfaces, significant amounts of IgG was found on the rough surfaces only. Prothrombin/thrombin and C1q were found initially on the thick-rough surface only. No TCC or plasmin levels were detected during the exposure time studied. Platelets and neutrophils were activated to a significantly higher degree on rough surfaces than on smooth, and neutrophils more on the rough surface with a thin oxide layer than on the thick one. The ensemble of antigens and enzyme activity detected shows a fingerprint which is unique for each material investigated.