Biomedical Engineering Reference
In-Depth Information
membrane obtained by osmotic lysis of erythrocyte cells, are
readily adsorbed on alkanethiol-coated gold.
87
The thickness of the
adsorbed erythrocyte membrane, as measured by ellipsometry,
SPR and AFM, ranges from 3 to 4 nm. This is not an unreasonable
estimate for the thickness of a single leaflet of the erythrocyte
membrane, in view of the presence of transmembrane proteins,
which are expected to contribute to the total thickness of the layer.
Acetylcholinesterase, an erytrocyte marker enzyme associated with
the outer leaflet of erythrocyte cells, retains its activity on this al-
kanethiol/erythrocyte hybrid bilayer for at least eight days. The
fact that the enzyme binds to the outer leaflet of the erythrocyte
membrane supports the view that the ghost membrane splits and
spreads onto the thiol SAM exposing the outer leaflet to the bulk
solution, as depicted in
Fig. 8a
. Alkanethiol-coated gold was also
used to adsorb antigens for screening antibody production.
88,89
Usually, epitopes, i.e., peptide sequences representing the major
antigenic sites of the antigen, are adsorbed onto the alkanethiol
monolayer by vesicle fusion or by Langmuir-Blodgett transfer,
thus creating a structured antigen-bearing surface. Antibodies
raised against the peptide sequence show notable preferential bind-
ing to the peptide-covered part of the surface. The resulting in-
crease in the thickness of the gold-supported film is followed by
SPR. This procedure can also be used for studying receptor-ligand
binding events. Thus, for instance, the enzyme pyruvate carbox-
ylase, which catalyzes the carboxylation of pyruvate, requires the
coenzyme biotin for catalytic activity. Biotin is a small molecule
covalently bound to a lysine residue at the active site of the en-
zyme. Avidin is a protein that can bind biotin very tightly, making
it unavailable for its function as a coenzyme. Lipid vesicles con-
taining a biotinylated phospholipid are fused onto alkanethiol-
coated gold. Injection of neutravidin gives rise to specific biotin-
neutravidin interactions. The resulting increase in film thickness is
again followed by SPR.
89
Annexin A1 is a protein that reversibly binds to cellular mem-
branes in a Ca
2+
-dependent manner. It was adsorbed on a
POPC:(palmitoyloleoylphosphatidylserine = POPS) (4:1) lipid
monolayer assembled by vesicle fusion on an octanethiol mono-
layer tethered to the gold surface of a QCM.
91
Adsorption of an-
nexin A1 to this hybrid bilayer in the presence of Ca
2+
ions was
monitored by the decrease in resonance frequency. No adsorption
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