Hello,

Did you heat sample? If your phospholipids have phase transition temperature, heating above the phase transition temperature would facilitate the lipid hydration. However, if your liposomes are unstable, liposomes form large aggregation and then precipitate soon. In this case, the problem might be dispersion stability of liposomes.

hi Keitaro Sou ,thanks for your helpful comment and yes i heated them above their phase transition T . i think perhaps reducing the concentration of chol will help and i want to search about it but any other suggestion will be appreciate. do you know how i should make my liposomes more stable to avoid this aggregations ?

i used citrat -Na2HPO4 as buffer .is it possible that this buffer was not

suitable for preparing liposomes ?

I use PEG-lipids and negatively charged lipids to avoid the aggregation of liposomes. I usually use physiological saline or PBS for hydration. Steric hindrance by PEG chain and electrostatic repulsion between liposome surfaces avoid the aggregation of liposomes. Each lipid is effective but both are more effective.

If your liposomes are containing cationic lipids, the citric acid in buffer might be a possible factor to induce liposome aggregation by electrostatically interacting with liposomes.

thanks a lot . i can't use PEG in my project because i want to investigate the interaction of bilayer itself by protein and PEG may interfere .PC and chol are used as lipid .

i want to know is there any way to stop formation of aggregations without changing composition of my liposomes ?

I think you need a hydrophile to balance and solvate the hydrophobic lipids and induce vesicle formation. I may be wrong here, but having just a lipid and no hydrophile (PEG) might not work. Do you have a reference from literature which demonstrates such an approach?

thank you Hemant Vishwasrao and yes there are references and i even myself saw this works .for example i attached one article that used POPC without any PEG .

thank you Yanuar Limasale .i think your suggestion will solve my problem it was very useful.

First, are you sure you have removed absolutely all of the organic solvent from the lipid film before attempting rehydration? This is very important. Overnight under strong vacuum, such as a lyophilizer, is effective.

Second, have you tried extruding your resuspended lipids, such as with a LIPEX extruder? 10 passes through two 0.1-micron membranes at a temperature above the transition temperature usually results in a stable suspension.