Compensation Procedures
EPC9 Manual
63
alternatingly to arrive at settings for which the remaining transient is minimal.
Settings for
C-fast
are left at their respective values whenever an
Auto C-slow
is
performed and vice versa. This way, a combined fit can be obtained by alternatingly
executing
Auto C-slow
and
Auto C-fast
. It is better, however, to perform
Auto C-fast
in
the cell-attached recording configuration and not to touch it any more when
proceeding to whole-cell.
During
Auto C-fast
and
Auto C-slow
, certain settings are changed temporarily and
restored upon completion. These are:
• R
s
-compensation is switched off
• The gain is set to 0.2, 10 or 50 mV/pA, depending on the gain range in use
• External stimuli are disabled
• Filter 1 is set to 10 or 30 kHz, depending on the required bandwidth
These changes are not displayed on the screen, since they usually are only effective
for fractions of a second. In
Cap. Track
, however, the changed values are effective for
longer times, since the
Auto C-slow
procedure is called periodically. It should be kept
in mind that under this condition the parameters on the screen may not represent the
hardware settings.
The relationship between the values of the compensation network (
C-slow
and
R-
series
) and those of the pipette-cell assembly (
C
m
and
R
s
) is straightforward, if the
membrane conductance is negligible. In this case, perfect compensation will leave no
residual current and
C-slow
will be equal to
C
m
. If, however, there is a finite
membrane conductance, then some ambiguity exists, because a three-component
network is being approximated by a two-component compensation network. Details
of the compensation procedure will then determine the residual current (some
filtered version of the command waveform) and
C
m
is likely to be underestimated.
The
Auto C-slow
procedure is designed to provide unbiased estimates of the actual
membrane capacitance and series resistance (Sigworth, Neher & Affolter, 1995).
Offset Compensation
In all patch-clamp configurations a number of offsets have to be taken into account.
These include amplifier offsets (± 30 mV), electrode potentials (± 200 mV, depending
on Cl
-
concentration of pipette and reference electrode), liquid junction potentials,
and potentials of membrane(s) in series with the membrane under study. Some of
these offsets are fixed during an experiment (like amplifier and electrode offsets),
some are variable.
