Lipidation of proteins is used in the pharmaceutical industry to improve the therapeutic efficacy of proteins. However, there is a limited knowledge on the physicochemical properties of these lipidated proteins. They may form irreversible multilayers upon adsorption onto solid surfaces, which in turn may catalyze the formation of surface-associated protein fibrillation. Fibrillation of proteins is considered to be a cause of multiple serious diseases. Insulin detemir, a human insulin analogue, with a fatty acid chain attached to the protein, was used as model compound, and compared to human insulin. Initial AFM and QCM-D results have shown that the lipid chain significant increases the initial adsorption and subsequent formation of fibrils onto hydrophobic surfaces. NR data can give a unique structural insight into the initial adsorbed (multi)-layers, density and thickness of these layers as well as for surface-associated fibrils. This structural insight will be of great significance in exploiting lipidation of proteins as a safe method to improve the therapeutic activity of proteins and understand the impact of the lipid chain in the adsorption process and subsequent fibrillation.