A-MSystems 2400 User manual

INSTRUCTION MANUAL
FOR
PATCH CLAMP AMPLIFIER
MODEL 2400
Version 14.0
March 2020
A-M Systems, Inc.
PO Box 850
Carlsborg, WA 98324
U.S.A.
360-683-8300 . 800-426-1306
FAX: [phone]
http://www.a-msystems.com
Each Patch Clamp Amplifier is delivered
complete with:
A headstage probe (purchased separately)
Model Cell
5” Model Cell connector cable
Rack Mount Hardware

Disclaimer
THIS EQUIPMENT IS NOT
INTENDED FOR USE WITH
HUMAN SUBJECTS IN ANY
WAY.
Document
The information contained in
this manual was as accurate as
possible at the time of
publishing, but is subject to
change without notice and
should not be construed as a
commitment by A-M Systems,
Inc. Changes may have been
made to the hardware or
firmware it describes since
publications. A-M Systems,
Inc. reserves the right to change
specifications as required. For
the latest information please
check our website
(http://www.a-msystems.com)
or contact A-M Systems, Inc.
directly.
Manual Product Number
880004
Manual Document Number
5027600 Rev 14
All rights reserved. No part of
this document may be
reproduced by any means
without the prior written
permission of A-M Systems,
Inc.
Safety
This instrument is provided with
terminal for protective grounding.
Before applying power, verify that
the correct safety precautions are
taken (see the following warnings).
In addition, note the external
markings on the instrument that are
described under Safety
Symbols. Do not operate the
instrument with its cover removed.
Replace fuse only with specified
type.
Supply Voltage
This equipment is customized at
the factory for line voltages of the
county of destination. The required
line voltage is indicated on the rear
panel.
WARNING
Do not attach a line voltage that
does not match the line voltage
specified on the rear panel.
Before turning on the instrument,
you must connect the protective
earth terminal of the instrument to
the protective earth conductor of
the (mains) power cord. The mains
plug must only be inserted in a
socket outlet with a protective earth
contact.
Service should be performed by
trained personnel only. To avoid
dangerous electric shock, do not
perform any service unless
qualified to do so.
Do not operate the instrument in
the presence of flammable gases or
fumes. Operation of any electrical
instrument in such an environment
constitutes a definite safety hazard.
Safety Symbols
The product is marked with this
symbol when it is necessary for
you to refer to the instruction
manual in order to protect
against damage to the product.
WARNING
The Warning symbol calls
attention to a procedure or
practice, which, if not correctly
performed could result in
injury. Do not proceed beyond
a Warning symbol until the
indicated conditions are fully
understood and met.
CAUTION
The Caution symbol calls
attention to a procedure or
practice, which, if not correctly
performed could result in
damage to the product. Do not
proceed beyond a caution until
the indicated conditions are
fully understood and met.

Table of Contents
Table of Contents....................................... iv
List of Tables and Figures........................... v
About This Manual ..................................... 1
Chapter Overview................................... 1
Conventions ............................................ 1
Quick Overview.......................................... 2
Getting Started ............................................ 4
Basic Connections................................... 4
Functional Test........................................ 5
Single Channel Recordings..................... 6
Whole Cell Recordings......................... 11
Current Clamp Recordings ................... 17
Instrument Operation ................................ 20
Probe..................................................... 20
Power .................................................... 21
Meter..................................................... 21
Mode..................................................... 22
Capacity Compensation........................ 24
Fixed Outputs........................................ 24
Whole Cell Compensation.................... 25
Series Resistance................................... 25
Clamp Signal......................................... 27
Probe Input............................................ 29
Bandwidth............................................. 29
Bath Input.............................................. 30
Telegraph Outputs................................. 30
AC Power Input .................................... 32
Model Cell ............................................ 32
Connecting to a Computer........................ 33
Troubleshooting........................................ 34
Circuit Description.................................... 36
Probe..................................................... 36
Clamp Signal......................................... 36
Current Clamp....................................... 36
Membrane Voltage Output ................... 37
Compensation ....................................... 37
Specifications............................................ 38
Input Impedance.................................... 38
Output Impedance................................. 38
Input Current/Voltage ratings ............... 38
Probe Gain and Bandwidth................... 38
Probe Case ............................................ 38
Maximum Instrument Noise................. 39
Internal Bandwidth................................ 39
Membrane Current Output Range......... 39
Membrane Voltage Output Range........ 39
Filter...................................................... 39
RMS Meter............................................ 39
Capacity Compensation........................ 39
Whole Cell Compensation.................... 40
Series Resistance................................... 40
DC Balance........................................... 40
Hold Command..................................... 40
Offset Command................................... 40
Speed Test Command........................... 40
Vcomp Command................................. 40
Iresist Command................................... 40
External Command: Voltage Clamp..... 41
External Command: Current Clamp ..... 41
Power Supply Requirements................. 41
Physical Dimensions............................. 42
Powered Probe Input............................. 42
Warranty ................................................... 43
Index ......................................................... 45

A-M Systems, Inc.
131 Business Park Loop, PO BOX 850, Carlsborg, WA 98324 USA
Telephone: [phone] * 360-683-8300 * FAX: [phone]
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List of Tables and Figures
Figure 1. Model Cell Schematic ................. 7
Figure 2. Electrode in the air....................... 8
Figure 3. Electrode in the bath.................... 8
Figure 4. Offset adjustment trace................ 9
Figure 5. Patch seal trace .......................... 10
Figure 6. Patch seal with transients
compensated...................................... 10
Figure 7. Whole cell recording before
capacity compensation...................... 14
Figure 8. Whole cell recording with fast
transients cancelled........................... 14
Figure 9. Whole cell recording with all
transients cancelled........................... 14
Figure 10. Whole cell recording with series
resistance controls turned on to 90% 15
Figure 11. The Probe................................. 20
Figure 12. Power switch ........................... 21
Figure 13. Meter........................................ 21
Figure 14. Mode........................................ 22
Figure 15. Probe Gain............................... 23
Figure 16. Filter ........................................ 23
Figure 17. Gain ......................................... 23
Figure 18. Mode Output............................ 24
Figure 19. Capacity Compensation........... 24
Figure 20. Fixed outputs ........................... 24
Figure 21. Whole Cell Compensation....... 25
Figure 22. Whole cell recording before
compensation .................................... 25
Figure 23. Whole cell recording after whole
cell compensation.............................. 25
Figure 24. Series Resistance controls ...... 25
Figure 25. Voltage clamp series resistance
controls.............................................. 26
Figure 26. Whole cell recordings with RsPre
at 90% ............................................... 26
Figure 27. Whole cell recordings with
RsComp at 90% ................................ 26
Figure 28. Whole cell recording with
RsComp and RsPre at 90%............... 27
Figure 29. Whole cell recording with
RsComp and RsPre at 90% and Whole
Cell parameters readjusted................ 27
Figure 30. Current clamp Bridge Balance27
Figure 31. Hold and Offset command
control ............................................... 27
Figure 32. Tracking Controls................... 28
Figure 33. Command Filter...................... 28
Figure 34. External Command input........ 28
Figure 35. Probe input.............................. 29
Figure 36. Bath Input............................... 30
Figure 37. Telegraph Outputs.................. 30
Figure 38. AC Power Input..................... 32
Figure 39. Model Cell.............................. 32
Figure 40. Model 2400 Block Diagram... 35
Figure 41. Probe block diagram............... 36
Table 1 Typography Conventions............... 1
Table 2 Minimal equipment........................ 4
Table 3 Front panel test settings ................. 5
Table 4 Typical noise levels ....................... 6
Table 5 Single channel set-up..................... 7
Table 6 Probe Gain and Bath VClamp ....... 9
Table 7 Whole Cell set-up ........................ 12
Table 8 Current clamp set-up.................... 17
Table 9 Probe resistor values.................... 19
Table 10 Probe resistor values.................. 20
Table 11 Probe Pin-out ............................. 21
Table 12 Meter Display settings ............... 22
Table 13 Probe resistor current gain......... 23
Table 14 Mode output units...................... 24
Table 15 External Clamp Signal Scaling.. 29
Table 16 Noise telegraph output............... 30
Table 17 Gain telegraph output................. 30
Table 18 Filter telegraph output................ 31
Table 19 Mode telegraph output............... 31
Table 20 Error telegraph output................ 32
Table 21 Common Problems .................... 34

A-M Systems, Inc.
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131 Business Park Loop, PO BOX 850, Carlsborg, WA 98324 USA
Telephone: [phone] * 360-683-8300 * FAX: [phone]
E-mail: [e-mail] * Website: http://www.a-msystems.com
About This Manual
Chapter Overview
This manual is designed to provide the researcher with the basic features of
Model 2400 in order to proceed to experiments as quickly as possible. For this
reason we have left the detailed descriptions of each function towards the end of
the manual. Chapter 1 will review the instrument’s capabilities. Chapter 2 will list
the necessary steps to start collecting data, in each of the following situations:
single channel, whole cell, or current clamp mode. Chapter 3 provides
description of the amplifier’s features and controls. Chapter 4 describes how to
interface the Model 2400 with various software packages. Chapter 5 lists some
simple steps to identify and solve common problems encountered in
electrophysiological recording. Chapter 6 provides a description of the amplifier’s
circuitry.
Although this manual provides the basics on using our patch clamp amplifier, it
assumes the researcher has some knowledge of patch clamp techniques. For a
more in-depth discussion of patch clamp recording we refer you to “Single-
Channel Recording”, edited by Bert Sackmann and Erwin Neher (ISBN
030644870X, Plenum Press), and “Voltage and patch clamping with
microelectrodes” edited by Thomas G. Smith, Jr. et al. (ISBN 0683077732,
American Physiological Society).
Conventions
This manual uses certain conventions to indicate elements of the Model 2400’s
user interface. The following table shows some examples:
Named panel controls or
connectors Adjust OFFSET until Im reads zero on the oscilloscope.
Knobs within named outlined
area on the front panel follow
the outlined area after a colon
Set MODE:GAIN to 100, for an overall
gain of 10mV/pA
Values on the meter The meter should display 0.05 pA.
Dual knobs. (i) refers to the
inner knob and (o) refers to the
outer knob
Alternately adjust FAST2-GAIN(i) and
FAST2-LAG(o) until the transient is
minimized
Table 1 Typography Conventions

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1
Quick Overview
The Model 2400 is a low noise, full featured intracellular/extracellular amplifier
designed for voltage or current clamping using patch electrodes on single
channels or whole cells. Its unique design allows fast intracellular current clamp
measurements with sharp electrodes. Advanced circuit design techniques using
field programmable gate arrays eliminated noisy microprocessors.
Amplifier current gain is extremely flexible as it can accept any of four switchable
resistive feedback probes. Each probe has two feedback resistors selected from
the following values: 10M, 100M, 1G, and 10G. The main amplifier automatically
recognizes the feedback resistor and adjusts internal gains to always provide the
correct values to the meter and correct signal magnitude at the outputs. The wide
range of feedback resistors means currents from 1 fA to 1 µA can be recorded
with outputs of 1mV/nA to 10mV/fA.
Unlike most patch clamp amplifiers, the Model 2400 has a voltage follower in the
probe. This allows this amplifier to be a true fast current clamp amplifier with no
instability. An integrated 6 position four pole low pass Bessel filter provides
flexible signal conditioning. Dual fine tuning capacity compensation is available
to eliminate virtually all electrode-induced transients. Calibrated whole cell
compensation provides easy display of membrane capacitance and access
resistance.
A host of command potentials are integrated internally within the model 2400,
including an automatic tracking command to zero the membrane current, manual
controls for offset and holding potentials, and an easily readable digital display.
For signals that are more complicated, an external command input with different
scaling factors is available for use with any signal source.

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Standard stimulation protocols are integrated into the MODE switch. Membrane
voltages can be monitored in 3 modes: Iclamp, Iresist, and Ifollow. Iclamp
is the generic current clamp mode where clamp signals can be customized using
the front panel controls or via the external clamp input. Iresist sends a fixed
amplitude square wave for easy measurement of electrode resistance. Ifollow
configures the probe to be a simple voltage follower to monitor membrane
voltage. Currents can be monitored in 3 modes: Vcomp, Vclamp, and Vtest.
Vcomp is a voltage clamp mode with a fixed amplitude square wave clamping
signal, useful for searching for cells and adjusting the compensation controls.
Vclamp is the generic voltage clamp mode where clamp signals can be
customized using the front panel controls or via the external clamp input. VTest
sends a square wave of current into the probe for testing the frequency response
of the probe.
A digital meter provides accurate values of command signals and membrane
currents or voltages, the true RMS noise of the amplifier and experimental setup,
the cut off frequency of the low pass filter, and the overall gain of the amplifier
plus probe.
Series resistance compensation provides the researcher with the option of
introducing either or both predictive and corrective compensation from zero to
100%. Fine and coarse controls for lag provide sensitive control to minimize
oscillation produced by compensation close to 100%.
Separate compensation controls exist for eliminating transients seen during
current clamp experiments when the bridge balance is used.
Telegraph outputs provide analog voltage equivalents of front panel settings
including error conditions, amplifier mode and gain, Cmembrane, RMS noise, and
low pass filter cut-off value. These telegraphs allow the Model 2400’s front panel
settings to be automatically recorded by your system software.
For those who need extra large bandwidths, the raw bandwidth of the entire
amplifier may be adjusted via a rear panel potentiometer.
A second powered probe input is available for use as a bath signal input.

A-M Systems, Inc.
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131 Business Park Loop, PO BOX 850, Carlsborg, WA 98324 USA
Telephone: [phone] * 360-683-8300 * FAX: [phone]
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2
Getting Started
Patch clamp amplifiers can seem complicated. This chapter is designed to take
you through an experiment with the minimal amount of instructions necessary. If
you are familiar with patch clamp recording already, you may want to skip ahead
to Chapter 3 where each feature of the Model 2400 is described.
When you receive your Model 2400 confirm that everything in the packing list is
included. Make sure there are no obvious signs of internal damage, such as
rattling. Pick up the amplifier and tilt in gently from side to side, and listen for
anything that might be loose.
Check that the correct voltage for your country is shown on the back of the unit.
The Model 2100 should be delivered with the appropriate mains power voltage
set, at either 100-120 volts or 220-240 volts. The setting is indicated on the back
panel to just to the side of the power input. If the wrong voltage or no voltage is
indicated on the rear panel contact A-M Systems, Inc. immediately
In this chapter you will need the equipment listed in the following table.
A-M Systems, Inc. Model 2400 Patch Clamp Amplifier
A-M Systems, Inc. Model Cell
An oscilloscope
Basic Connections
Connect the power cable to the AC POWER INPUT on the rear of the unit and
to an approved outlet. Do not turn the Model 2400 on. Connect the probe cable
to the PROBE input on the rear of the unit. Connect a BNC cable from the
MODE OUPUT to an oscilloscope.
Be certain to properly ground yourself whenever you handle the probe as it is
susceptible to electric static discharge (ESD). ESD can cause immediate or subtle
damage to sensitive electronic parts. You can reduce the chances of ESD damage
by:
·Only connecting the probe to the amplifier when the amplifier is off.
·Always grounding yourself by touching the handle of the amplifier, or
another grounded piece of metal prior to handling the probe.
·Avoiding extraneous walking around while holding the electrode holder
or model cell, especially if you are on carpet or during conditions of low
temperature and low humidity.
Table 2Minimal equipment
WARNING
CAUTION

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Functional Test
This simple test will check to see if the most essential components of the
amplifier are working.
With the amplifier turned off, set the front panel controls as follows:
METER (knob) Noise
MODE Vclamp
MODE PROBE GAIN LOW
MODE: GAIN 1
MODE: FILTER Open
CAPACITY COMPENSATION: FAST1: Gain Fully counterclockwise
CAPACITY COMPENSATION: FAST1: Lag Fully counterclockwise
CAPACITY COMPENSATION: FAST2 Fully counterclockwise (inner and outer knob)
WHOLE CELL COMPENSATION OFF
SERIES RESISTANCE OFF
CLAMP SIGNAL (Hold/Offset Toggle) OFF
CLAMP SIGNAL: 3kHz OFF
CLAMP SIGNAL (tracking) OFF
CLAMP SIGNAL: EXTERNAL OFF
Place the probe in a Faraday cage (this can be as simple as surrounding the probe
with aluminum foil and grounding the foil). Note: the BNC shields on the front
panel make good ground connectors.
Depress the Power switch.
The power switch should light up and the meter should display the noise value.
Step the FILTER knob counterclockwise through the decreasing cut-off
frequency positions and the noise value should drop. Select the 1.0kHz position.
The meter now displays the RMS noise at 1kHz.
Switch the PROBE GAIN between HIGH and LOW. You should see the noise
value on the meter alternate between two different values. The values will depend
on the type of probe you have. The following table gives expected values of
noise for each type of feedback resistor. At any time, you can change METER to
GAIN, and MODE: GAIN to 1, then the meter will display the current gain of
the probe. You can then refer to the table to determine the value of resistor in
each PROBE GAIN setting.
Table 3Front panel test settings

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Feedback Resistor Typical RMS noise (@1kHz) Probe Gain
10M 0.003 nA 10mV/nA
100M 0.5 pA 100mV/nA
1G 0.17 pA 1mV/pA
10G 0.08 pA 10mV/pA
If you were able to complete the preceding procedures, then most of the main
amplifier is functioning and you have a valid connection to the probe. If the
noise is much larger than the values displayed in Table 4, confirm that the input
connector is not touching a ground source. If the noise remains excessive, you
may have a broken probe, continue with the rest of this section to see if any other
features are malfunctioning. If the noise is much lower than listed in the table,
confirm that the probe is connected properly. If the probe is not connected to
the amplifier the PROBE error light will be lit.
The following will test the components within the probe. Change the MODE
knob to Vtest. This will inject a biphasic 4nA peak to peak (p-p) square wave at
60Hz (0.2nA p-p with probe gain high).
Set your oscilloscope to 1V/Division, 5ms/div, and triggered on channel 1. On
your oscilloscope, you should see a square wave reflecting the gain of your probe.
If you do not see a square wave, check to see that the probe cable is firmly
screwed into the rear panel. You may need to reduce the vertical sensitivity on
the oscilloscope.
If you see a square wave, then the probe is working. If you do not have a square
wave on the output, re-check all connections, and confirm there is no error
indication on the front panel. If problems persist please call customer service at
A-M Systems, Inc.
Single Channel Recordings
This section will walk you through the features of the Model 2400 that are
commonly used in a typical single channel recording experiment. In this section
we will be using the model cell, but after you become familiar with the controls
on the Model 2400, you will be able to substitute a real electrode and cell for the
model cell.
Table 4 Typical noise levels

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The model cell is an electrically equivalent circuit of a simplified
electrode/bath/cell interface. Figure 1 shows the schematic diagram of the
model cell. The component names indicate which part of the electrode, bath, or
cell they represent.
Equipment you will need in this section is the Model 2400 with probe, the model
cell, and an oscilloscope. In a real experiment, you would also need a pulse
generator, a cell chamber (bath), a cell, a manipulator, and something to record
your data.
Set-Up
Set the controls on the front panel as follows:
METER (knob) MODE
MODE Vclamp
MODE: PROBE GAIN HIGH
MODE: GAIN 1
MODE: FILTER Open
CAPACITY COMPENSATION: FAST1: Gain Fully counterclockwise
CAPACITY COMPENSATION: FAST1: Lag Fully counterclockwise
CAPACITY COMPENSATION: FAST2 Fully counterclockwise (inner and outer knob)
WHOLE CELL COMPENSATION OFF
SERIES RESISTANCE OFF
CLAMP SIGNAL (Hold/Offset Toggle) OFF
CLAMP SIGNAL: 3kHz OFF
CLAMP SIGNAL (tracking) OFF
CLAMP SIGNAL: EXTERNAL OFF
Connect the MODE: OUTPUT to channel 1 on your oscilloscope. Set the
oscilloscope to 1V/division, 2ms/division, and trigger on channel 1. Attach the
probe to your micromanipulator located within the recording (Faraday) chamber.
Attach the model cell to the probe’s BNC connector, but leave the ground pin
unconnected.
Figure 1. Model Cell Schematic
R_Patch_Seal
10G
To Probe GND
BATH
11
Cpatch
3.3pF
Rmembrane
500M
To Probe GND
Patch
11
Celectrode
2.7pF
To Probe
BNC
1
2
To Probe GND
WholeCell
11
Cmembrane
33pF
Relectrode
10M
Relectrode
10M
Celectrode
2.7pF
Table 5 Single channel set-up

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Entering the Bath
Normally as you approach the cell you would provide a square wave voltage
clamp signal (for example, 10mV, 60Hz). In theory, before you enter the bath the
current resulting from the square wave is essentially zero since you are clamping
an extremely large resistance (from the electrode through the air to your reference
electrode) (Figure 2). Small transients at the frequency of the clamping signal
might be observed. When the electrode enters the bath, the resistance seen by the
amplifier will suddenly drop to the resistance of the electrode. This will mean
there will be a large increase in current and you should see a square wave of
current on the oscilloscope (Figure 3). When the electrode contacts the resistance
seen by the amplifier will increase, so the current will decrease. As the patch seal
is formed (through suction) the current will decrease further. The better the
patch seal the smaller the current waveform will be.
In our case we will be using the model cell to simulate these conditions. You
have two choices for producing the square wave clamping signal. You can either
use the built in signal by switching the MODE switch to Vcomp, or you can
provide your own signal through the EXTERNAL CLAMP SIGNAL BNC. If
you are using Vcomp, then the amplifier will generate a 10mV p-p 60Hz signal
internally (+/- 5mV).
If you are using an external signal, turn the CLAMP SIGNAL:EXTERNAL
toggle switch to ¸10 or ¸50 depending on the scale factor you want for your
signal. If you want to use the internal signal, change MODE to Vcomp.
The signal you see on the oscilloscope should be essentially a flat line at 0VDC
with sharp transients at 60Hz.
Connect a ground cable between the ground pin on the probe and the Bath pin
on the model cell.
If you are using a 100Meg/10Gig Headstage Probe, the signal you see on the
oscilloscope should be a 60 Hz square wave with amplitude of 10Vp-p when
PROBE: GAIN = HIGH. Switching PROBE: GAIN = LOW selects the
smaller feedback resistor. If that resistor value is 100Meg, then the amplitude of
the square wave should be 100mVp-p. Expected observed values are listed in
Table 6.
Figure 2. Electrode in the air
0.m 2.m 4.m 6.m 8.m 10.m
-1.
0.
1.
V(VIB)
T
V(10Vc)*10
Figure 3. Electrode in the bath
0.m 2.m 4.m 6.m 8.m 10.m
-1.
0.
1.
V(VIB)
T
V(10Vc)*10

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Feedback Resistor Probe Gain Observed Signal at OUTPUT
10Meg 10mV/nA 10mV
100Meg 100mV/nA 100mV
1Gig 1000mV/nA 1V
10Gig 10000mV/nA 10V
This signal is similar to those you would see once you have a patch electrode in a
bath. Return MODE switch to Vclamp.
Offset Zero
There can often be offsets due to different conductances between your reference
electrode and your recording electrode (Figure 4). These offsets can be adjusted
to zero automatically using tracking, or manually using the OFFSET control.
The following procedure will enable you to set the value of offset manually. First,
switch the METER knob to Voffset in order to display the voltage value
determined by the knob OFFSET. Slowly rotate OFFSET clockwise and watch
the value on the meter increase. Set the offset value to 2mV . Notice that as you
moved OFFSET, the current trace on the oscilloscope did not move. This is
because the offset voltage is not yet connected to the clamping potential. To turn
on the offset, set the hold/offset switch to Ofor offset only (Figure 31, Page 27).
Notice that the current trace is now offset vertically on the oscilloscope. We can
now use this artificial offset to see how automatic tracking works.
The tracking control monitors the current output, and automatically adjusts the
clamping voltage until the current equals zero. To use tracking, set the tracking
switch to TRACK.Notice that the trace slowly adjusts until the output is
centered on zero volts. To fix the offset at this value, switch the tracking knob to
SET. This will fix the offset value and you will be able to change the clamping
value again. Try adjusting OFFSET when the tracking switch is in the TRACK
position and when it is in the SET position. If you switch the tracking switch to
OFF this will disconnect the tracking circuit from the clamping circuit.
Return the tracking switch to OFF and the offset/hold switch to OFF. If you
are using an external signal, turn the CLAMP SIGNAL:EXTERNAL toggle
switch to ¸10 or ¸50 depending on the scale factor you want for your signal. If
you want to use the internal signal, change the MODE switch to Vcomp.
Table 6 Probe Gain and Bath VClamp
Figure 4. Offset adjustment trace
0.m 2.m 4.m 6.m 8.m 10.m
-1.
0.
1.
V(VIB)+50m
T
V(10Vc)*10

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Getting A Seal
To simulate a patch seal with the model cell, change the ground connector on the
model cell from BATH to PATCH. (If only it was this easy with a real cell.
With a real cell you would have to monitor your approach to the cell by watching
until the current pulses become about half as large. Application of suction, if
applied properly (see Single-Channel Recording by Sakmann and Neher), will
form a giga-seal seen by the current trace becoming essentially flat, with capacitive
transients at rising and falling edges of the clamp signal).
Capacity Compensation
Once you have switched the model cell connection from BATH to PATCH the
trace on the oscilloscope should be flat with small transients at the frequency of
the clamping signal (Figure 5). These transients are due to the total capacitance of
the amplifier, holder, and electrode. You should verify that PROBE GAIN is
HIGH in order to see clearly the transients. These transients can be cancelled out
using CAPACITY COMPENSATION knobs FAST 1 and FAST 2. Use
both the FAST 1:GAIN and FAST 1:LAG to minimize the transient. You will
need to alternate between GAIN and LAG until the trace looks similar to Figure
6. While working with a real cell, you may need to use FAST2 if there is more
than one time constant to the transient.
Filtering
You can further optimize the output signal available at MODE OUTPUT by
using the four-pole low pass Bessel filter. This filter affects only the MODE
OUTPUT and does not affect the x10Vm or Im outputs. To adjust the filter
frequency, turn the MODE: FILTER knob to the desired filter frequency.
Changing the METER knob to Noise will result in the display indicating the
noise levels with the selected filter applied. When you change the METER knob
to MODE the meter will display the membrane current.
Holding and Command Signals
CLAMP SIGNAL: HOLD provides a holding potential to the cell. This knob
works similar to OFFSET, except a change in the holding potential is seen as a
change in both membrane voltage and current, while OFFSET only affects the
membrane current.
Switch the METER knob to Vhold. Using HOLD, adjust the value to 0mV.
Switch the METER knob to Voffset. Using OFFSET, adjust the value to
0mV. Connect a BNC cable from the x10Vm output to Channel 2 of your
oscilloscope. Set the voltage range on channel 2 to 1V/division.
Figure 5. Patch seal trace
0.m 2.m 4.m 6.m 8.m 10.m
-1.
0.
1.
V(VIB)
T
V(10Vc)*10
Figure 6. Patch seal with transients
compensated
0.m 2.m 4.m 6.m 8.m 10.m
-10.m
0.m
10.m
V(VIB)
T
V(10Vc)
Make certain you
ground yourself before
handling the probe
CAUTION

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Adjust the hold/offset switch to O. This will turn on the OFFSET control.
Adjust the OFFSET control and watch as both the value on the meter and the
output change, but the x10Vm does not (oscilloscope channel 2). Switch the
METER knob to Vhold and the hold/offset switch to (H+O). This will turn on
both the OFFSET and HOLD controls. Adjust HOLD and watch how the
meter, MODE OUTPUT, and x10Vm output all change.
Any command potential that is not a constant DC command must be created
with an external device and connected to the EXTERNAL input. The external
input is then divided by 10 or 50 before it reaches the cell, depending on the
setting of the EXTERNAL switch.
Recording
Typically in channel recordings you would record both the membrane voltage and
current. You can get both of these outputs from the front panel of the 2400. It
is advantageous to monitor membrane current at the MODE OUTPUT , rather
than Im, because MODE OUTPUT can be filtered using FILTER and amplified
using GAIN, whereas the Im output is an unfiltered, un-amplified output straight
from the probe. The membrane voltage is recorded from the x10Vm output.
Outputs can be recorded onto a computer using any number of analog to digital
converters available from a variety of manufacturers. See Chapter 4 “connecting
to your computer” for more information.
Whole Cell Recordings
This section will walk you through the features of the Model 2400 that are
commonly used in a typical whole cell recording experiment. In this section, we
will be using the model cell, but after you become familiar with the controls you
will be able to substitute a real electrode and cell for the model cell.
The model cell is an electrical equivalent circuit of a simplified
electrode/bath/cell interface. Figure 1 shows the schematic diagram of the
model cell. The component names indicate what part of the electrode, bath, or
cell they represent.
Equipment you will need in this section is the Model 2400 with probe, the model
cell, and an oscilloscope. In a real experiment, you would also need a pulse
generator, a cell chamber (bath), a cell, a manipulator, and something to record
your data.

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Set-Up
Set the controls on the front panel as follows:
METER (knob) MODE
MODE Vclamp
MODE: PROBE GAIN LOW
MODE: GAIN 1
MODE: FILTER Open
CAPACITY COMPENSATION: FAST1: Gain Fully counterclockwise
CAPACITY COMPENSATION: FAST1: Lag Fully counterclockwise
CAPACITY COMPENSATION: FAST2 Fully counterclockwise (inner and outer knob)
WHOLE CELL COMPENSATION OFF
SERIES RESISTANCE OFF
CLAMP SIGNAL (Hold/Offset Toggle) OFF
CLAMP SIGNAL: 3kHz OFF
CLAMP SIGNAL (tracking) OFF
CLAMP SIGNAL: EXTERNAL OFF
Connect the MODE: OUTPUT to channel 1 on your oscilloscope. Set the
oscilloscope to 1V/division, 2ms/division, and trigger on channel 1. Place the
probe in your recording chamber attached to your micromanipulator. In a
normal experiment, you would then attach the electrode holder and patch pipette
to the BNC connector on the probe. Instead, attach the model cell to the probe’s
BNC connector, but leave the ground pin unconnected.
Entering the Bath
Normally as you approach the cell you would provide a square wave voltage
clamp signal (10mV, 60Hz). In theory, before you enter the bath the current
resulting from the square wave is essentially zero since you are clamping an
extremely large resistance (from the electrode through the air to your reference
electrode) (Figure 2). Small transients at the frequency of the clamping signal
might be observed. When the electrode enters the bath the resistance seen by the
amplifier will suddenly drop to the resistance of the electrode. This will mean
there will be a large increase in current and you should see a square wave of
current on the oscilloscope (Figure 3). When the electrode contacts the cell the
resistance seen by the amplifier will increase, so the current will decrease. As the
patch seal is formed (through suction) the current will decrease further. The
better the patch seal the smaller the current waveform will be.
In our case we will be using the model cell to simulate these conditions. You
have two choices for producing the square wave clamping signal. You can either
use the Model 2400’s built-in signal by switching the MODE switch to Vcomp,
or you can provide your own signal through the EXTERNAL CLAMP
SIGNAL BNC.
Table 7 Whole Cell set-up

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If you are using an external signal, turn the CLAMP SIGNAL:EXTERNAL
toggle switch to ¸10 or ¸50 depending on the scale factor you want for your
signal. If you want to use the internal signal, change the MODE switch to
Vcomp.
Turn on the model 2400.
The signal you see on the oscilloscope should be essentially a flat line at 0VDC
with sharp transients at 60Hz.
Connect a ground cable between the ground pin on the probe and the Bath pin
on the model cell.
If you are using a 100Meg/10Gig Headstage Probe, the signal you see on the
oscilloscope should be a 60 Hz square wave with amplitude of 100mVp-p when
PROBE: GAIN = LOW. Expected observed values are listed in Table 6.
This signal is similar to those you would see once you have a patch electrode in a
bath. Return MODE switch to Vclamp.
Offset Zero
There can often be offsets due to different conductances between your reference
electrode and your recording electrode. These offsets can be adjusted to zero
automatically using tracking, or manually using the OFFSET control.
The following procedure will enable you to set the value of offset manually. First,
switch the METER knob to Voffset in order to display the voltage value
determined by the knob OFFSET. Slowly rotate OFFSET clockwise and watch
the value on the meter increase. Set the offset value to 50mV . Notice that as you
moved OFFSET, the current trace on the oscilloscope did not move. This is
because the offset voltage is not yet connected to the clamping potential. To turn
on the offset, set the hold/offset switch to Ofor offset only (Figure 31, Page 27).
Notice that the current trace is now offset vertically on the oscilloscope. We can
now use this artificial offset to see how automatic tracking works.
The tracking control monitors the current output, and automatically adjusts the
clamping voltage until the current equals zero. To use tracking, set the tracking
switch to TRACK. Notice that the trace slowly adjusts until the output is
centered on zero volts. To set the offset value at this value switch the tracking
knob to SET. This will fix the offset value and you will be able to change the
clamping value again. Try adjusting OFFSET when the tracking switch is in the
TRACK position and when it is in the SET position. If you switch the tracking
switch to OFF this will disconnect the tracking value from the clamping circuit.
Return the tracking switch to OFF and the offset/hold switch to OFF. If you are
using an external signal, turn the CLAMP SIGNAL:EXTERNAL toggle switch
to ¸10 or ¸50 depending on the scale factor you want for your signal. If you
want to use the internal signal, change the MODE switch to Vcomp.

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Attaching To A Cell
To simulate a seal with the model cell you need to do is change the ground
connector on the model cell from BATH to PATCH. With a real cell you would
have to monitor your approach to the cell by watching until the current pulses
become about half as large. Application of suction, if applied properly (see
Single-Channel Recording by Sakmann and Neher), will form a giga-seal seen by
the current trace becoming essentially flat, with capacitive transients at rising and
falling edges of the clamp signal. After the seal is formed, access to the cell can
be achieved by a number of methods, including applying more suction. In the
model cell access to the cell is made by changing the connection from PATCH to
CELL.
Capacity Compensation
Once you have switched the model cell connection from BATH to CELL the
trace on the oscilloscope should be a square wave with large transients at the
frequency of the clamping signal (Figure 7). There are two main transients seen, a
fast transient due to the capacitance of the amplifier and holder, and a slow
transient due to the electrode access resistance and cell capacitance. The fast
transient can be cancelled out using CAPACITY COMPENSATION knobs
FAST 1 and FAST 2. Use both the FAST 1: GAIN and FAST 1: LAG to
minimize the transient. You will need to alternate between GAIN and LAG until
the trace looks similar to Figure 8. While working with a real cell, you may need
to use FAST2 if there is more than one time constant to the transient.
Whole Cell Compensation
Figure 7. Whole cell recording before
capacity compensation
0.m 2.m 4.m 6.m 8.m 10.m
-10.m
0.m
10.m
V(VIB)
T
V(10Vc)
Figure 8. Whole cell recording with
fast transients cancelled
0.m 2.m 4.m 6.m 8.m 10.m
-10.m
0.m
10.m
V(VIB)
T
V(10Vc)
Figure 9. Whole cell recording with
all transients cancelled
0.m 2.m 4.m 6.m 8.m 10.m
-10.m
0.m
10.m
V(VIB)*10
T
V(10Vc)
Make certain you
ground yourself before
handling the probe
CAUTION

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The slow transients often seen in whole cell recording can be minimized using
WHOLE CELL COMPENSATION. Whole cell compensation is only active
when the PROBE GAIN is LOW. Confirm that Raccess and Cmembrane
are fully counterclockwise. Turn on WHOLE CELL COMPENSATION.
Alternately adjust Raccess and Cmembrane in order to reduce the slow
transient. It will help as the transients get smaller to increase the voltage
sensitivity on your oscilloscope (by decreasing the volts/division) to
0.5V/division. Also, if the transients are hard to eliminate, you may want to turn
on the 3kHz filter in the CLAMP SIGNAL section of the Model 2400.
Following appropriate compensation, your trace should look like Figure 9. The
values of series resistance and capacitance should match the model cell’s electrode
resistance and membrane capacitance respectively. Note that full scale on the
Raccess 10-turn potentiometer is equal to 100Meg during voltage clamping.
Series Resistance Compensation
This section will give a brief description of how to provide series resistance
compensation. A more detailed description and listing of the differences between
correction and prediction is given in the instrument operations chapter (Chapter
3) and circuit description chapter (Chapter 6).
If the whole cell compensation is properly tuned, you should be able to turn
RsPRE and RsCOMP to 90% without producing large transients. Alternately
adjusting the LAG control, FAST1, FAST2, and Cmembrane should
produce a trace similar to Figure 10. A suggested step-by-step procedure follows:
·Turn FAST1 GAIN counterclockwise a little bit.
·Turn LAGfine fully clockwise and LAGcoarse fully counterclockwise.
·Turn RsPRE and RsCOMP fully counterclockwise.
·Turn on SERIES RESISTANCE (there should be no change in the
waveform on Channel 1 of the oscilloscope)
·Slowly adjust RsPRE clockwise to 90%. Observe that no oscillations
occur at the transients. If oscillations occur, adjust RsPRE
counterclockwise.
·Adjust FAST1-GAIN, FAST1-LAG, Raccess, and Cmembrane to
minimize transients.
·Slowly adjust RsCOMP to 90%. If necessary adjust FAST1, FAST2,
and Cmembrane to eliminate transients.
·If transients do not minimize try turning on the 3kHz CLAMP
SIGNAL filter.
Figure 10. Whole cell recording with
series resistance controls turned on to
90%
0.m 2.m 4.m 6.m 8.m 10.m
-10.m
0.m
10.m
V(VIB)*10
T
V(m10Vc)*-1