PC-505B Manual, Rev. 201013
Warner
Instruments
A Harvard Apparatus Company
29
wave to the pipette. Adjust your acquisition system or oscilloscope and amplifier so as to
clearly display 1-2 complete cycles of the output current signal. An example trace is shown
to the right.
Note:
If needed, the amplitude of the
TEST PULSE
can be set to a different value (
1
,
10
, or
100
mV
) as needed to see a clear signal. This is achieved by adjusting the
SENSITIVITY SELECTOR
(for example, x0.01 = 10 mV).
Read the amplitude of the current (peak-
to-peak on the square wave in pA or nA) on
your oscilloscope or acquisition system. From
the voltage amplitude of the test pulse and the
output current you can calculate RP in M
from Ohm's law (V=IR).
Notes
:
(1) If reading from an oscilloscope, be sure to
take into account the
GAIN MULTIPLIER
just above
GAIN SELECTOR
switch.
(2
) 1 mV/pA = 1 G
Voltage clamp
Attach to the cell and form gigaseal
Leave the
TEST PULSE
active and advance the pipette tip to contact the cell. Once the
pipette begins to dimple the cell membrane, the amplitude of the test current should begin
to decrease. When this happens, release the applied positive air pressure in the electrode
holder and watch the test pulse for seal formation.
If a seal does not immediately form, then apply a pulse of gentle suction (2-3 kPa or
less) to the electrode holder and watch the test current closely for seal formation (usually
10-30 s). Repeat as necessary and once a seal forms, quickly release suction to avoid going
into whole-cell.
You’ll know a seal has formed when the test current quickly decreases, then goes flat.
The signal appears flat since the current is now too small to see at the scale your recording
system is set to. You can increase the signal amplitude by making the following adjustments
in order: (1) increase the
COMMAND SENSITIVITY
, (2) increase the amplifier
GAIN
, and (3) increase
the y-axis scaling on your recording system.
Notes
:
1.
If a seal doesn't form within about 30 seconds, try alternately releasing and re-applying
suction for about 10-30 s each.
2.
If a seal appears but forms very slowly, continue alternating the suction, or advance the
pipette very slightly against the cell.
