Chemistry Reference
In-Depth Information
Electrolyte
150mM NaCl in Millipore water as electrolyte (8.8mg/mL NaCl in water) is used.
Agar Salt Bridge
0.5% (i.e. 0.5 g/ 100mL) agarose in electrolyte solution is used to form a gel to fill
the capillaries used to form the electrodes.
Pulling Micropipettes
A ramp test was done on the Sutter P-97 Flaming Brown pipette puller and the
procedure for Program 1 as recommended in the manual was followed and the results
are as below: (The ramp test is done to identify the temperature at which the glass
will melt and can be pulled into a micropipette)
Ramp test results:
Heat: 791 (R)
So the following program was used:
Heat: R
+
=
15
806
Pull: 0
Velocity: 50
Time: 150
For the borosilicate glass (1
.
0
×
0
.
5mm): heating time was 7.44 s (1 loop)
Liposome Preparation
50mg of 1,2-diphytanoyl-sn-glycero-3-phosphocholine (DPhPC, Avanti Polar
Lipids, Alabaster, AL) is taken in a glass vial and Buffer (10mM HEPES, 200mM
KCl, pH 7.06; 5mL) is added to the lipid and vortexed for 10 s. The solution was
subjected to five freeze-thaw cycles. The lipid-buffer suspension was then pushed
through a 0
m PTFE filter (VWR). The extruded sample was diluted with buffer
to a volume of 12mL, yielding a 2mg/mL lipid solution. The vesicle solutions were
divided into aliquots of 1mL and stored at 4
◦
C.
.
2
µ
