Immunologic response to protein immobilized on the surface of liposomes via covalent azo-bonding

A new method for immobilizing protein on the surface of liposomes is described. Inclusion of N-(p-aminophenyl) stearylamide in the lipid composition of vesicles resulted in liposomes that could be 'activated' by diazotization with NaNO2/HCl, and subsequently coupled with protein. Using this method 39.7 +/- 7.5 micrograms egg albumin/mumol phospholipid has been coupled to multilamellar vesicles composed of phosphatidylcholine, cholesterol, and N-(p-aminophenyl) stearylamide in a molar ratio of 15:75:1.1. Furthermore, when the immunologic response of mice to egg albumin that was encapsulated in, nonspecifically absorbed, or covalently linked to liposomes was investigated, only the covalent protein-liposome conjugates elicited pronounced and sustained elevations in antibody titers. These results suggest that the immunoadjuvant effects of liposomes can be maximized by covalently linking protein antigens to their surface.