Highly efficient immobilisation of antibody fragments to functionalised lipid monolayers

The covalent attachment of Fab' fragments of polyclonal anti-human IgG to a lipid with a terminal linker group was examined by means of quartz crystal microbalance and surface plasmon resonance measurements. The linker lipid was embedded in binary or ternary monolayers of dipalmitoylphosphatidylchol ine (DPPC) and cholesterol. Atomic force microscopy images of the films dep osited on silanised SiO2 substrates showed that Fab' fragments take a stand ing position, thus giving site-directed immobilisation. Human IgG forms a n etwork on interaction with the antibodies. Non-specific binding of bovine s erum albumin was found to be very low when DPPC was used as the host matrix . At an optimal Fab' fragment concentration a binding capacity above 60% wa s obtained. However, if the surface concentration of the immobilised antibo dies was too high, the binding capacity decreased due to steric hindrance. The results demonstrate that the covalent coupling of Fab' fragments to N-( epsilon-maleimidocaproyl)-dipalmitoylphosphatidylethanolamine (DPPE-EMC) em bedded in a host monolayer matrix of DPPC is a promising approach to achiev e a site-directed immobilisation of antibodies with high antigen-binding ef ficiency. (C) 1999 Elsevier Science B.V. All rights reserved.