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Hi all,

I need to make lipid micelles for an experiment. My lipids are dissolved in chloroform. I dry them under argon, resuspend in aqueous buffer (10mM sodium phosphate, 100mM KCl, 1mM EDTA), vortex 1 min to get a cloudy solution, and then sonicate. I use a bath sonicator made by Laboratory Supplies Company Inc, as recommended by Avanti Polar Lipids. My protocol suggests sonicating for about 10-20 min.

I know that the solutions are supposed to get clear (or at least, less cloudy) following sonication and that this change indicates that micelles have formed. However, my solutions remain cloudy even after 45 min of sonication. My questions are:

1) What can I do to get micelles to form?

2) How can I verify the presence of micelles? Could I use a quick measurement (light scattering? CD? turbidity?) to verify that the micelles have formed properly?

Thanks in advance!

flag	asked May 5 at 19:47 Rappley 148●6

WS · Answer 1 · 2010-05-06 07:20:39Z UTC

Dear Irit,

the appearance of micelles depends a lot on their composition and size. You may check the solution before and after sonication with an UV/vis spectrophotometer. Depending on their size, the wavelength dependent light scattering and thus the OD of the solution will change.

If you should have access to a FACS or a particle sizer, this will be a better option, of course.

if your lipids dissolve in a solvent which is miscible with water (like ethanol, maybe DMSO or DMF work, too), injecting the lipid solution (slowly) through a narrow gauge needle (use eg an insulin- or tuberculin syringe) into 10 times the volume of eg PBS while mixing might work well, too (at least it did for me with fluorescently labelled ceramide, where sonication was not satisfactory). If the ethanol is not compatible with your downstram application, you may remove it by dialysis or possibly gel filtration. Cells tolerate small percentages of ethanol quite well, in my experience.

Best Regards,

Wo

Rappley · Accepted Answer · 2010-05-28 20:43:37Z UTC

After much troubleshooting, it turns out that the vessel in which the liposomes are sonicated is important. My original protocol involved drying the lipids in a glass vial, then hydrating and vortexing them in the same glass vial, then transferring the cloudy solution to an eppendorf 1.5mL tube for sonication (because the small glass vials have flat bottoms, which tend to shatter during sonication). The result was a cloudy solution, even after 1 hr of sonication, with liposomes that were on average about 180 nm (LUVs).

It turns out that if I transfer the cloudy solution to a round-bottomed borosilicate glass culture vial (the kind you might use to collect FPLC fractions) for sonication, the solution gets clear within 15 minutes and the resulting liposomes are mostly SUVs (57% of the population averages around 19 nm, with only 32% averaging around 126 nm).

When sonicating in the large glass vial, it is important that the level of liquid inside the vial is flush with the level of liquid outside the vial.

help with lipid micelles

2 Answers