In molecular biology and protein engineering the immobilized metal ion affinity chromatography (IMAC) using a NTA-chelator is a very powerful technique in identification and isolation of oligo-histidine-tagged fusion proteins. This concept was transferred to the properties of self-assembling systems with the aim of reversible immobilization, orientation of biomolecules, and functionalization of lipid interfaces. Here we describe the synthesis and the chemical and physical characterization of such metal affinity lipids. The NTA-chelator was coupled either to a phospholipid, DPPE, or to a synthetic lipid, DODA. Metal complex formation was investigated by means of TLC and FTIR techniques. Using film balance techniques the generation of metal sensitive lipid films is demonstrated. In the presence of Ni2+ drastic changes of the area-pressure isotherms were observed. Furthermore, the specific ligand binding of imidazole as a model compound for oligo-histidine-tagged fusion proteins to these functionalized metal-lipid films was investigated.