Effects of Triton X-100 concentration and incubation temperature on carboxyfluorescein release from multilamellar liposomes

Abstract

Carboxyfluorescein is the most commonly used probe to measure the rate of release of vesicle contents. The validity of the data obtained by this method depends on obtaining an end point based on the complete release of the dye on treatment of the liposomes with a detergent, usually Triton X-100. However, Triton does not completely release entrapped carboxyfluorescein from multilamellar liposomes and the amount and rate of release of marker upon detergent treatment is a function of lipid composition of the liposome, Triton concentration and temperature and duration of detergent incubation. The fluorescence `end point' for distearoyl--[alpha]-phosphatidylcholine/cholesterol (2:1, mol%) multilamellar liposomes treated with 0.5% Triton at 22[deg]C (a condition often used) is only about one-fifth the value for liposomes treated with 5% Triton at 72[deg]C. The conditions of treatment appear to affect the release of carboxyfluorescein from the lipid of the partially or completely disrupted liposome and the subsequent partitioning of the free dye into the aqueous phase. This effect can lead to serious errors in the interpretation of multilamellar liposome stability data. However, Triton allows complete release of entrapped dye from small unilamellar vesicles under all conditions tested.