Protein-liposome conjugates with defined size distributions.

Conjugation of protein to liposomes by two coupling protocols is shown to result in vesicle aggregation. The degree of aggregation is directly related to the levels of protein conjugated to the liposomes. In an attempt to develop a method of generating stable, homogeneously sized protein-conjugated vesicles, highly aggregated liposome-protein conjugates were extruded through filters of defined pore size distributions, with no loss of protein binding. The extruded samples are relatively stable with respect to size and are easily prepared for various protein to lipid ratios. Liposome size has been shown to be a major factor in determining the in vivo blood circulation times of liposomes. A corresponding, significant enhancement in the blood circulation lifetimes for extruded versus aggregated streptavidin-liposome conjugates is observed. Furthermore, the stability of streptavidin-liposome conjugates in vivo was shown by the binding of biotin to liposomes isolated from plasma 1 and 4 h post-injection. In conclusion, extrusion of the aggregated systems obtained on coupling proteins to liposomes provides a convenient and general method for generating homogeneously sized protein-liposome conjugates.