PC-505B Manual, Rev. 201013
Warner
Instruments
A Harvard Apparatus Company
31
current on the oscilloscope. An example is shown on the previous page. Since the test pulse
has a known voltage amplitude (100 mV in this case), you can calculate RS by use of Ohm's
law (R=V/I). A good gigaseal resistance is typically in the range 1-10 G
.
Patch or whole cell recording?
At this point you can perform either patch clamp or whole cell recording. The following
sections address additional setup steps to be performed prior to making measurements for
each technique.
Both patch and whole cell recording can be performed with either the
LC-201B
low
current or
HC-202B
high current headstages. In either case, whole cell recordings are
performed with the
PROBE RESISTOR
SELECT
toggle switch set to the
low
position (500 M
or
50 M
, respectively for each headstage).
Patch recording - Leak subtraction
If RS is only a few G
, it may be necessary to
compensate for membrane voltage errors lost to
the leak pathway.
With
TEST PULSE
still active, turn the
LEAK
SUBTRACTION
control clockwise until the I
m
trace on
your acquisition system or oscilloscope becomes
flat except for residual capacitance transients.
(Alternatively, with the
TEST PULSE
off,
run the
holding potential up and down over a range that
doesn't evoke channel currents and adjust the
LEAK SUBTRACTION
control so that the current trace
on the oscilloscope remains fixed.) An example is
shown to the right.
If RS is very high, leak subtraction may not
be needed. If so, turn the
LEAK SUBTRACTION
control
to
0
(
off
).
Patch recording – Excised patch
Watch with the microscope as you excise the patch. To pull a patch, quickly move the
pipette away from the cell with the manipulator, then up and away using the fine vertical
controls. During patch excision, the cell should stay attached to the bottom of the dish. If
the excision is successful, the critical vibration-sensitive phase of the operation is complete.
If the cell remains attached to the pipette, try jiggling the micromanipulator controls, or
tap gently on the micromanipulator, or bring the cell and pipette up briefly into the air for
no more than 1 second. If the patch still does not separate from the cell, you should consider
proceeding with a cell-attached patch recording instead.
