BINDING OF BIOTINATED-LIPOSOMES TO STREPTAVIDIN IS INFLUENCED BY LIPOSOME COMPOSITION

We describe an 'in vitro' assay which allows rapid quantification of t he binding of biotinated-vesicles to streptavidin immobilised on micro titre plates by estimating levels of a liposome encapsulated fluoresce nt molecule, rhodamine 123. It is shown that optimal vesicle binding t o streptavidin occurs when a six carbon biotin spacer arm derivative o f distearoylphosphatidylethanolamine (biotin-X-DSPE) is incorporated i n liposomes. This alleviates steric hindrance arising due to the inclu sion of small amounts of large bulky amphiphiles such as monosialogang lioside (G(M1), 5 mol%) in vesicles. In contrast the ability of liposo mes containing poly(ethylene glycol) derivatives of DSPE (PEG(2000)-DS PE, 5 mol%) to bind streptavidin was only marginally better when bioti n-X-DSPE was substituted for biotin-DSPE in vesicles. It is further sh own that amounts of biotinated-vesicles bound to streptavidin were min imally influenced by the fluidity of the liposome preparation when ass ayed at 4 degrees C. However, at elevated temperatures (37 degrees C) lipid estimates as determined by vesicle entrapped rhodamine 123 were low due to leakage of this marker from vesicles. This was shown by com paring amounts of biotinated-liposomes bound to streptavidin coated pl ates using the lipid marker [H-3]cholesteryl hexadecyl ether to estima tes determined by vesicle entrapped rhodamine 123. The 'in vitro' assa y protocol described here is a general method applicable in the optimi sation of other targeting protocols. In conclusion our work suggests t hat liposomes containing G(M1) and the spacer arm derivative biotin-X- DSPE bind optimally to immobilised streptavidin which should aid in th e use of biotinated-liposomes in 'in vivo' targeted delivery applicati ons.