Preparation of multilayer plasma protein films on silicon by EDC/NHS coupling chemistry

Crosslinked multilayer protein films were prepared from fibrinogen, albumin, IgG, a combination of fibrinogen and catalase, and blood plasma on silicon by ethyl-dimethyl-aminopropylcarbodiimide and N-hydroxy-succinimide coupling chemistry. The 4–70 nm thick films were placed in blood plasma and the additional protein deposition measured by null ellipsometry after 5 or 60 min of incubation. The activation of the complement system and intrinsic pathway of coagulation were indicated through the subsequent binding of anti-C3c, anti-C3d, anti-properdin and anti-HMWK on top of the surface bound blood plasma. The proportion of Annexin V, Propidium Iodide and 4,6-diamidino-2-phenylindole positive cells, and the secretion of tumor necrosis factor (TNF-) and interleukin-10 (IL-10) were analysed in a monocyte culture. The results show that well known protein coupling techniques can be used for the preparation of protein layers with well controlled thickness. The layers possess low contact activation of blood plasma and induce different release of TNF- and IL-10 in monocyte cultures.