Separation of proteoliposomes from proteins and liposome-encapsulated macromolecules from liposomes
After protein has been incorporated into some form of liposome, it is necessary to resolve the proteoliposomes from any unincorporated protein. The most widely used strategy is described in Figure 1: the sample is adjusted to a density of 1.17-1.22 g/ml by mixing with a high-density stock solution and layered beneath two lower density solutions (the top layer is sometimes the isolation buffer).
During centrifugation the proteoliposomes float upwards to band at the top interface. The big advantage of this strategy is that the unincorporated protein remains in the sample zone and will even tend to sediment in the opposite direction (Figure 1).
If the sample is on top of a
density barrier the proteoliposomes and the free proteins sediment in the same direction; separation of
the two types of particle may be less clear; such separations also require longer centrifugation times
(see Section 5b).