Electrochemistry of Biomimetic Membranes - Applications of Electrochemistry and Nanotechnology in Biology and Medicine - page 165

in the case of SPR, or of its dielectric constant in the case of EIS.
However, it must be borne in mind that the two techniques are
sensitive to different features of a film.
2. Vesicle Fusion
Vesicles (or, more precisely, unilamellar vesicles) are spherical
lipid bilayers that enclose an aqueous solution. To obtain vesicles,
lipids are usually dissolved in an organic solvent. The solvent is
then evaporated using a nitrogen stream or vacuum, so that a thin
lipid film is produced on the glass surface of a vial. The lipid film
is hydrated with an aqueous solution, whose temperature should be
above the gel to liquid-crystalline transition temperature of the
lipid with the highest transition temperature in the mixture. Giant
unilamellar vesicles (GUVs) have a diameter ranging from 1 to
300 Pm. They are usually produced by an electroformation ap-
proach. Multilamellar vesicles (MLVs) are quickly generated by
the general protocol described above. Starting from MLVs, large
unilamellar vesicles (LUVs, 100 to 1000 nm in diameter) with a
narrow size distribution around a desired value are produced by
freeze-thaw cycling the vesicles, followed by extrusion; this con-
sists in pressing the vesicle suspension repeatedly through a mem-
brane of defined pore size. Small unilamellar vesicles (SUVs, 20
to 50 nm in diameter) are prepared by extrusion through mem-
branes with smaller pore size (about 30 nm) or by supplying ultra-
sound energy to the MLV suspension by using an ultrasonic bath
or an ultrasonic probe (sonication).
The procedure for vesicle fusion consists of adsorbing and
fusing SUVs on a suitable substrate from their aqueous dispersion.
If the substrate is hydrophilic, vesicle fusion gives rise to a lipid
bilayer by rupture of the vesicles and their unrolling and spreading
onto the substrate. Conversely, if the substrate is hydrophobic, a
lipid monolayer with the hydrocarbon tails directed toward the
substrate is formed by rupture of the vesicles, splitting of the ve-
sicular membrane into its two monomolecular leaflets and their
spreading 12 , as shown schematically in Fig. 8 . This is confirmed by
the different thickness change following vesicle fusion on a hydro-
phobic substrate (2-2.5 nm) with respect to a hydrophilic substrate
(4.5-5 nm), as estimated by SPR. 13