Electrochemistry of Biomimetic Membranes - Applications of Electrochemistry and Nanotechnology in Biology and Medicine - page 219

Gold-supported DPTL/DPhyPC tBLMs on microchips have
recently been employed to measure single channel currents of pep-
tides and proteins. To this end, a microelectrode array device con-
sisting of many (100u100 Pm 2 ) sensor pads was employed (see
Fig. 15 ). 179-181 A DPTL monolayer was tethered to the gold-coated
sensor pads from a DPTL solution in ethanol; then, a lipid mono-
layer was formed on top of it by vesicle fusion. In view of the very
small surface area of the pad, the resistance of the resulting tBLM
ranged from 1.5 to 17 G:. This resistance was high enough to
reduce the background electrical noise to the low level required for
the use of the patch-clamp technique. A droplet of a buffered
aqueous solution was applied to a given tBLM-coated gold pad.
Then, a conventional patch pipette containing an Ag/AgCl elec-
trode and filled with the same buffer solution was inserted into the
droplet. The electrical circuit of the patch-clamp amplifier was
closed with a tungsten-tipped electrode that was positioned onto a
gold-coated probe pad electrically wired to the given sensor pad.
This devise has allowed the recording of single channel cur-
rents of gramicin A, 179 the high-conducting Ca 2+ -activated K + (BK
or Maxi-K) channel, the synthetic M2G ion channel 180 and the
mechanosensitive channel of large conductance (MscL) from
Escherichia coli . 181 All these peptides and proteins were incorpo-
rated into lipid vesicles, before fusing them onto the DPTL-coated
sensor pad. With the exclusion of the gramicidin channel, the uni-
tary conductance of the remaining ion channels was found to be
from one third to one tenth of that obtained with conventional
BLMs. In this connection, one cannot exclude the possibility that,
even in this case, the channels responsible for single channel cur-
rents are located in the membrane of adsorbed or partially fused
vesicles. In fact, if this is the case, then the capacitive coupling
between the vesicular membrane and the tBLM is expected to de-
crease the unitary conductance of the channels. The channel life-
times for both open and closed states were normally found in good
agreement with those obtained under similar conditions in conven-
tional patch-clamp experiments. Plots of the current against the
applied voltage are roughly linear and pass through the origin of
the coordinate system, with the only exception of gramicidin.
However, it is not clear whether the voltage is referred to the
Ag/AgCl reference electrode or whether the curves were forced to
pass through the origin.