I am trying to perform dye leakage assays using carboxyfluorescein (CF) containing liposomes. I have been observing this strange phenomena where CF will passively leak out of the vesicles over time. The liposomes are prepared by rehydrating the lipid film (POPG/POPE) in 20 mM CF in PBS, followed by 10 freeze thaws, extrusion through 0.2 µm polycarbonate filter, and separation by size exclusion chromatography. Has anyone experienced anything similar and/or know how to avoid this issue?
Dear Imran Rahman ,
As you know leakage of carboxyfluorescein (CF) occur round the transition temperatures of the membrane lipids used. According to:
https://avantilipids.com/tech-support/physical-properties/phase-transition-temps
The transition temperatures (Tm) are:
POPE 25 oC
POPG -2 oC
So, working at room temperature is rather close to the Tm of POPE, I suspect that this might be a reason of your observation. Not sure about the reason for using POPE/POPG but using DOPE/DOPG might be a better combination.
Best regards.
Rob Keller Thank you! I will try using the alternative acyl chain length lipids.
Liposomes are sort of 'liquid-capsules'. So they are not perfect solid containers. Therefore, the leakage of drugs/dyes from liposomes naturally occur. Additionally, liposomes also disintegrate /denature over a period of time. That's why there is a shelf life of drug loaded liposomes.
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