Keithley 427

The planar lipid bilayer method has been described previously [36 (link)]. The membranes were formed from a 1% (w/v) solution of asolectin (phospholipids from soybean, Sigma-Aldrich) or diphytanoyl phosphatidylcholine (DiphPC, Avanti Polar Lipids, Alabaster, AL) in n-decane. The lipid membranes had a surface of approximately 0.5 mm² and they were formed using a Teflon loop to spread the lipid across the aperture in the dividing wall. After the membrane had turned black the toxin-containing protein fractions were added to the aqueous phase. The current across the membrane was measured with a pair of Ag/AgCl electrodes with salt bridges switched in series with a voltage source and a highly sensitive current amplifier Keithley 427 (Keithley Instruments, INC. Clevlend, OH). The signal was recorded by a strip chart recorder (Rikadenki Electronics GmbH, Freiburg, Germany). The temperature was kept at 20 °C throughout the experiment.

The methods used for the lipid bilayer measurements have been described
previously in detail (Benz et al., 1978 (link)).
A Teflon chamber divided into two 5 mL compartments. The compartments are
connected by a small circular hole with a surface area of about 0.4
mm² and were filled with 1 M KCl, 10 mM MES, pH 6.0. Black lipid
bilayer membranes were obtained by painting onto the hole solutions of 1% (w/v)
asolectin (phospholipids from soybean, Sigma-Aldrich) or diphytanoyl
phosphatidylcholine (DiphPC; Avanti Polar Lipids, Alabaster, AL) in
n-decane. All salts were analytical grade and the
temperature was maintained at 20°C during all experiments. The current
across the membrane was measured with a pair of Ag/AgCl electrodes with salt
bridges switched in series with a voltage source and a highly sensitive current
amplifier Keithley 427 (Keithley Instruments, INC. Cleveland, OH). The amplified
signal was recorded by a strip chart recorder (Rikadenki Electronics GmbH,
Freiburg, Germany).