Linker lipids were embedded in a phosphatidylcholine monolayer matrix prepared at the air-water interface. The covalent coupling of antibody fragments, non-specific adsorption of bovine serum albumin and specific binding of antibodies was monitored in situ with a 10-MHz quartz crystal microbalance. The attachment of antibody fragments and the activity of the layers was also showed with standardized radioimmunoassay. The results demonstrate that the coupling of Fab'-fragments to linker lipids in a monolayer matrix is a promising approach to achieve a highly oriented layer of antibody fragments with a high density of binding sites on the sensor surface for immunological measurements.