Binding of biotinated-liposomes to streptavidin is influenced by liposome composition

We describe an 'in vitro' assay which allows rapid quantification of the binding of biotinated-vesicles to streptavidin immobilised on microtitre plates by estimating levels of a liposome encapsulated fluorescent molecule, rhodamine 123. It is shown that optimal vesicle binding to streptavidin occurs when a six carbon biotin spacer arm derivative of distearoylphosphatidylethanolamine (biotin-X-DSPE) is incorporated in liposomes. This alleviates steric hindrance arising due to the inclusion of small amounts of large bulky amphiphiles such as monosialoganglioside (GM1, 5 mol%) in vesicles. In contrast the ability of liposomes containing poly(ethylene glycol) derivatives of DSPE (PEG2000-DSPE, 5 mol%) to bind streptavidin was only marginally better when biotin-X-DSPE was substituted for biotin-DSPE in vesicles. It is further shown that amounts of biotinated-vesicles bound to streptavidin were minimally influenced by the fluidity of the liposome preparation when assayed 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 comparing amounts of biotinated-liposomes bound to streptavidin coated plates using the lipid marker [3H]cholesteryl hexadecyl ether to estimates determined by vesicle entrapped rhodamine 123. The 'in vitro' assay protocol described here is a general method applicable in the optimisation of other targeting protocols. In conclusion our work suggests that liposomes containing GM1 and the spacer arm derivative biotin-X-DSPE bind optimally to immobilised streptavidin which should aid in the use of biotinated-liposomes in 'in vivo' targeted delivery applications.