I'm new in the field and I prepared a liposomal preparation using microfluidics (Nano assembler) DPPC:CHOL (2:1), solvents ethanol: distilled water. No drug loaded since I'm training. How do I separate the ethanol to try the zeta potential..etc I have been told I shouldn't leave my liposomes in water or ethanol either during storage or for other characterization tests.??
Also later on when I load a drug how do I choose the right purification methods (on what basis) I'll be using same lipid with ascorbic acid??
Please if there's any papers or article that explain it thoroughly.
Hi Tia Mo,
These articles may help you.
Article A membrane filtering method for the purification of giant un...
Article Rapid purification of giant lipid vesicles by microfiltration
Article Low cost non-electromechanical technique for the purificatio...
Hello,
you can remove the ethanol by dialysis or you can use higher FRR to reduce the ethanol content in final suspension to a concentration which will not interfere with other measurements. Is there any reason why are you using FRR 2:1?
Also, check the PrecisionNanosystems website for application notes.
https://www.precisionnanosystems.com/docs/default-source/pni-files/app-notes/pni-app-bt-011.pdf?sfvrsn=588bbd82_2
Thank you Jan Kotouček for your answer. The 2:1 is the molar ratio between the lipids. the FRR was also 2:1for a moderate size liposomes according to the PrecisionNanosystems papers. Would it be okay If I measure the zeta potential without the ethanol removal ?
Also would it be okay if I centrifuge and replace the ethanol with PBS?
Thanks,
Well, from my experience, there are several problems. First one is the viscosity of water/EtOH mixture. You need to know the exact viscosity for the DLS to obtain correct values (values of size and zeta potential). The second problem is the medium itself, for correct zeta potential measurement, the medium should have higher conductivity than the pure water (salt addition is needed). The third problem is the FRR 2:1, the vesicles are not fully formed, due to the high ethanol content in the final mixture. If you measure the size of the liposomes after the dialysis or dilution you will probably get worse distribution.
The DPPC lipid is a neutral lipid, so you will get slightly negative zeta potential near to zero value.
You could use higher FRR (1:7, 1:9) with appropriate stock lipid concentration, and instead of pure water, you could use PBS.
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