Hi, I'm doing experiments reconstituting membrane proteins to liposome.
And have a few questions.
1. If I use buffer while hydration of lipid, buffer can encapsulated into liposome. Then, detergent treatment for membrane protein reconstitution (I usually use 0.75% OG, n-octyl-beta-D-glucoside) results in leakage of buffer from liposome?
2. Will buffer leak from liposome during or after detergent removal by dialysis?
0.75% OG is approximately equal to the critical micellar concentration, so I would expect there to be some permeabilization and even disruption of the liposome membranes. This will cause the internal and external liquids to equilibrate.
Once the detergent is removed by dialysis, the internal compartment should become isolated from the external compartment again (assuming that the lipids chosen are capable of forming sealed liposomes).
If you need to create a situation in which there are different aqueous compositions inside and outside, you can dialyze the liposomes against a different buffer after the detergent removal stage, or pass the liposomes over a gel filtration or desalting column (e.g. PD-10) equilibrated with the desired external buffer.
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