Biomedical Engineering Reference
In-Depth Information
sphingolipids and cholesterol, which are present in the outer leaflet
of plasma membranes. Rafts are receiving increasing attention
since they are considered to regulate the
membrane function in
eukaryotic cells. The presence of microdomains was directly visu-
alized from the image of the distal lipid monolayer, by using two-
photon fluorescence lifetime imaging microscopy (2P-FLIM).
39
The distal monolayer consisted of a sphingomye-
lin:DOPC:cholesterol (47:47:6) mixture, with the addition of 1
mol% laurdan. Laurdan is a fluorophore that is known to distribute
uniformly in coexisting gel, liquid-ordered and liquid-disordered
phases. It emits in the blue region of the optical spectrum and has a
longer lifetime in the gel phase, while it emits in the green region
and has a shorter lifetime in the liquid-disordered phase. The fluo-
rescence lifetime image of the distal lipid mixture shows a high
degree of heterogeneity and is characterized by the coexistence of
gel, liquid-ordered and liquid-disordered phases. Microdomains
with an irregular percolative-like shape and lifetimes of 3450±50
ps are ascribed to the gel phase. Small roundish microdomains
immersed in the surrounding matrix have lifetimes of 2830±50 ps
and are attributed to the liquid-ordered phase. Finally, the matrix,
with intermediate fluorescence lifetimes, is ascribed to the coexist-
ence of the liquiddisordered phase and of liquid-ordered micro-
domains of size below the resolution of the microscope (about 0.4
Pm radial).
(
iii
)
(
Thiolipid-Spacer
)
-Based tBLMs
A mixture of a short hydrophilic spacer anchored to gold via a
sulfhydryl or disulfide group (e.g., mercaptoethanol or dithiodi-
glycolic acid) and of a thiolipid with its terminal hydrophobic
group (e.g., a cholesteryl
13,14
or a phytanyl group
34
) capable of
pinning a lipid bilayer gives rise to a practically aqueous region
sandwiched between the short spacer and the lipid bilayer (see
Fig.
19
); this may favor the accommodation of the extramembrane do-
mains of relatively bulky membrane proteins. Deposition of a lipid
bilayer onto such a mixture of a short spacer and of a thiolipid is
usually carried out by vesicle fusion or by rapid solution exchange.
The resulting lipid bilayer has a capacitance of about 0.5-0.6 PF
cm
-2
and a resistance higher than 5 M:cm
2
. These values com-
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