Instrumental operation: ICS 3000
Standard Operational Procedure
Jagger
Instrumental operation: ICS 3000
Scope
The scope of this document is to present an overview of the Dionex Ion Chromatography System (ICS)
3000 and to provide the reader with guidelines for standard operations, routine maintenance and
simple trouble shooting. Detailed instrument description can be found in Dionex Reference Library (CD-
ROM) which contains all relevant and specific manuals.
Instrument overview
The Dionex ICS 3000 system is designed for ion chromatography (IC). It is compatible with high salt and
alkaline solvents and therefore fittings, tubings and interior of the system is made of PEEK (polyether
ether ketone) to prevent corrosion. It consists of a dual pump system with four channels on each pump.
It is capable of operating two columns with two different detectors at the same time and is equipped
with a pulsed amperometry detector (PAD) and a conductivity detector. In practice, the common use
currently will be one column at a time and primarily on Pump 1 and the PAD detector which will be
described here. The PAD is based on electrochemical detection and is in general superior for detection
of carbohydrates that are traditionally difficult to detect. The system also includes an eluent generator,
which can be used when ultra pure eluents are needed. In practice, the eluent generator has limitations
that make it difficult to use for many applications and it will therefore be bypassed for most
applications. The principle of the conductivity detector is that changes in conductivity is measured and
interpreted as analytes. This requires the installation of an ion suppressor prior to the conductivity
detector, which will suppress the inherently high conductivity coming from the eluents. Figure 1 shows
an instrument overview. As opposed to many other HPLC systems, the system should be operated with
more or less constant flow to avoid detrimental salt precipitations in the pump, column and detector. If
the flow will be stopped for a prolonged time period make sure to replace salt containing eluents in all
lines with e.g. 50% methanol (also to avoid microbial growth).
HPLC systems are delicate equipment and should be handled accordingly. Regularly check the status of
the instrument, preferably on a daily basis and always be careful when operating. Eluents should
ALWAYS be prepared from fresh top quality water (Milli-Q/18.2 MΩ·cm) and preferably sample
preparation should also be made based on this water quality.
The chromatographic system is controlled by a computer installed with Chromeleon™ 7 software
(hereafter CM) version 7.1.1.1127 for instrument control and data handling. All commands and
instrument handlings goes through this software.
Instrumental operation: ICS 3000
Standard Operational Procedure
Jagger
The system consists of 5 compartments/modules:
1. Autosampler
2. Dual pump
3. Eluent generator
4. Column compartment (lower part)
5. Detector compartment (upper part)
Figure 1: General system appearance of the ICS-3000
Instrumental operation: ICS 3000
Standard Operational Procedure
Jagger
Figure 2: Instrument Controller. Software unit that controls communication between computer and HPLC. Must be running
at all times when system is operated.
Communication between the computer (CM) and the HPLC is controlled by the Instrument Controller
Software found under the Start Menu/Programs/Chromeleon/Service Manager. Alternatively, double
click the Chromeleon Icon in the Windows task bar to open the Instrument Controller (figure 2)or right
click and select start Chromeleon Instrument Controller. When the HPLC is to be controlled/operated
the Instrument Controller must be running. Starting and stopping is done by pressing the command
“Start/Stop Instrument Controller” (figure 2). The window can be closed afterwards. The Instrument
Controller does not start automatically after a computer restart and it should therefore be checked that
it is running, before starting operation.
In the operating software (CM) each module has its own operational screen, which may be found as tabs
in the top of the window (figure 3).
Window for starting and stopping
Instrument Controller. Can be closed
after use.
Icon informing about Instrument
Controller status. Green: Controller
running. White with red cross:
Controller stopped.
Instrumental operation: ICS 3000
Standard Operational Procedure
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Figure 3: Module and system overview from controlling software Chromeleon™ 7. Unmarked tabs: “DC”; Detector
compartment. “EGC1”; Eluent generator. “Audit”; Audit trail logging every event in the system. “Startup”; Programmed
system startup (not recommended to use unless you know what you’re doing).
Operating the pump
The internals of the dual pump system is depicted in figure 4. The lower half of the compartment is
Pump 1 and the upper half is Pump 2. The pumps are operated at flow rates between 0 and 10ml/min
and max pressure is 5000 psi. Each pump is connected to four eluent channels (A, B, C and D). A Rear
Seal Wash system is installed to periodically flush the backside of the pump heads in order to avoid salt
precipitations and corrosion. Wash setting may be “Interval”, “Automatic” or “Off”. Correct setting is
“Interval”, but the rear seal wash solutions needs to be changed on a weekly basis and consists of 20%
methanol/ethanol (figure 4). Note dates of change in the equipment logbook.
Switching between
Instrument and Data panel
Overview Autosampler PAD detector Queue Pump
System 1 – PAD
electrochemical
detection
Instrumental operation: ICS 3000
Standard Operational Procedure
Jagger
Figure 4: Internals of Dual Pump. Pump 2 is placed in the upper half of the compartment, Pump 1 in the lower half. Note the
screw valve at position 3 is the purge valve on Pump 2. Equivalent purge valve on Pump 1 found in the same position.
Position 11 is the rear seal wash reservoir which should be changed on a weekly basis (20% methanol or ethanol).
Preparing eluents
The eluents for ion chromatography are crucial for good performance and the chromatography (column)
is extremely sensitive to contaminations in the eluents. It is therefore of outmost importance that extra
care is taken when eluents are prepared. Remember that the lifetime of eluents for ion chromatography
is also very short compared to other types of chromatography. The biggest risk of contamination is
dissolution of carbonate in the eluents and this occurs when eluents are exposed to atmospheric air.
This carbonate accumulates on the columns stationary phase and reduces separation capacity and
carbonate has a higher push effect as mobile phase then the others regularly applied. It is impossible to
completely avoid carbonate contamination but with the proper precautions it can be minimized and
allow a certain lifetime of the eluents (max. 48 hours for top performance). This includes that
Instrumental operation: ICS 3000
Standard Operational Procedure
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headspace in the eluent bottles must be saturated with nitrogen to minimize the amount of carbon
dioxide. Nitrogen is delivered by a nitrogen generator via a restriction valve and distributed between
three of the four channels (channel D does not get nitrogen supply). Three different eluents are made to
accommodate the range of analytical methods and all three are changed every time new eluents are
prepared. The eluents are always prepared in the same way and always placed on the following
channels:
Channel A: 0.1 M NaOH
Channel B: 1 M NaOAc in 0.1 M NaOH
Channel C: Milli-Q water
Eluent A and B are most labile for carbonate contamination and should be handled accordingly.The
following is a description for generation of each eluent but should be performed alongside so that all
three eluents are done approximately at the same time. All eluents are sensitive to atmospheric air.
Therefore – don’t leave any of the newly prepared eluents standing without N2 over pressure in the
headspace for more than 15 minutes.
Before any eluents are made, turn off the flow on the system and release the nitrogen pressure on the
restriction valve positioned on the backside of the autosampler. Empty all eluent bottles and rinse them
thoroughly in Milli-Q water. Special pressure stable eluent bottles are used since strong corrosive
eluents are pressurized during running. Therefore, don’t send the bottles off to washing, but rinse them
properly before reuse.
Eluent A: 0.1 M NaOH (2L)
1. Measure out 2 L of Milli-Q water in a measuring flask. Make sure to use clean glassware.
2. Poor the two liter of water into the eluent bottle named Eluent A (0.1 M NaOH).
3. Loosely place a lid on the bottle, but don’t tighten it.
4. Place the eluent bottle in the sonication bath and adjust the water volume in the bath to the
level indicated as working level. The sonication bath will fit two eluent bottles at a time.
5. Sonication for 20 min. DON’T put your hands into the sonicator while it is operating and close
the door to the room when you leave it.
6. When the sonication is done, close the lid securely and move the eluent bottle to the HPLC.
7. Connect channel A to the eluent bottle and start the nitrogen flow. Exchange the air in the head
space by repeatedly opening and closing the lid on the bottle to relieve pressure. A total of three
reliefs are sufficient.
8. Use a sterile plastic pipette with a total volume of 10 ml and a manual “pipette bulb” for
pipetting a concentrated 50% NaOH solution (Fluka, 71868-1L). The solution is stored in the
exhausted waste cupboard underneath the HPLC and is extremely concentrated. ALWAYS wear
Instrumental operation: ICS 3000
Standard Operational Procedure
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proper personal protection equipment (glasses, gloves and lab coat/acid rubber coat) when
handling this solution. I am not kidding – getting that stuff in your eyes will ruin your eyesight!
9. Check the level of NaOH solution in the bottle – if the content is less than half of the original
replace with new to prevent risk of carbonate contamination in the NaOH solution. Place the
remainder of the solution in the toxic cupboard and mark it with “COMMON USE” for other
purposes than HPLC.
10. Pipette 10.4 mL to the 2 L bottle of eluent. Be careful when pipetting, as the solution is very
viscous. Don’t dip the entire pipette into the solution – only the tip underneath the surface.
Close the lid to the NaOH solution immediately after use and only use the same pipette ONCE in
the solution. Don’t double dip – take a new pipette.
11. Dispense the NaOH solution to the water in the eluent bottle underneath the water surface and
be patient as the liquid will flow very slowly (viscous). Slowly rinse the inside of the pipette by
gently pipetting up and down 3 times.
12. Close the lid to the eluent bottle and exchange the headspace air with nitrogen as done in step
7. Swirl the eluent bottle carefully – DON’T shake it, to distribute NaOH evenly.
Eluent B: 1 M NaOAce in 0.1 M NaOH (1L)
1. Weigh 82.04 g anhydrous sodium acetate (Sigma-Aldrich, 71183-1KG,or Fluka). It is important
that the correct quality of sodium acetate is used.
2. Poor this into a 1 L measuring flask (use clean glass ware) and add about 0.8L Milli-Q water.
3. Add a magnetic stir bar and place the solution on a magnetic stirrer till you have a complete
dissolution.
4. Remove the magnetic stir bar (without dipping the magic stick into the solution!) and fill the
remaining volume to 1 L (with freshly tapped Milli-Q water).
5. Mount a 0.22 µm sterile filter cup on top of the cleaned eluent bottle (f.ex. 0.22 µm stericup GP
Millipore Express PLUS membrane, 250 ml funnel, 45 mm neck size). Apply vacuum to filter the
eluent.
6. Loosely place a lid on top of the eluent bottle. Don’t tighten it.
7. Place the eluent bottle in the sonication bath, make sure water surface in the bath is at the
working level.
8. Sonicatefor 20 min. DON’T put your hands into the sonicator while it is operating and close the
door to the room when you leave it.
9. When the sonication is done, close the lid securely and move the eluent bottle to the HPLC.
10. Connect channel B to the eluent bottle and start the nitrogen flow. Exchange the air in the head
space by repeatedly opening and closing the lid on the bottle to relieve pressure. A total of three
reliefs are sufficient.
Instrumental operation: ICS 3000
Standard Operational Procedure
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11. Follow the procedures for NaOH addition as described for Eluent A step 8-12. EXCEPT! Only add
5.2 mL NaOH to the 1 L NaOAc-solution (ie. there should be the same NaOH concentration in
both channel A and B).
Eluent C: Milli-Q water
1. Fill the rinsed eluent bottle with Milli-Q water freshly tapped from the system.
2. Loosely place a lid on top of the eluent bottle. Don’t tighten it.
3. Place the eluent bottle in the sonication bath and adjust the water volume in the bath to the
level indicated as working level.
4. Sonicatefor 20 min . DON’T put your hands into the sonicator while it is operating and close the
door to the room when you leave it.
5. When the sonication is done, close the lid securely and move the eluent bottle to the HPLC.
6. Connect channel C to the eluent bottle and start the nitrogen flow. Exchange the air in the head
space by repeatedly opening and closing the lid on the bottle to relieve pressure. A total of three
reliefs are sufficient.
End the preparation procedure by purging all three channels as described below and note in the logbook
date of eluent change.
Purging eluents
When eluents are changed the pump needs to be purged to remove air bubble formation and old eluent
from the tubes. The system is sensitive to air (as all HPLCs) and purging is therefore important. Purging
the eluent channels is done be applying high flow through the specified channels and will create high
back pressure on the system, if the purge valve has not been open (Open the purge valve (figure 4) by
approx. half a turn). This will direct the flow path to a waste tube and relieve the pressure on the system
(and the column in particular).
Once the purge valve has been opened choose the appropriate channels and flow distribution to purge
in CM and activate the purge/prime function (figure 5). It is possible to purge more than one channel at
a time by setting the percentage distribution between the channels as desired (equal distribution, 50:50
when two channels, 33% when three and 25% when all four channels are to be purged). The software is
preprogrammed to perform purging for 500 seconds at 5.7 mL/min. Repeat purging (if necessary) until
air bubbles are no longer visible in the eluent channels from the bottles to the pump.
IMPORTANT: Before purging the software will inform that this operation will generate a
high flow through the system and that the purge valve should be open before
executing. Press “Execute despite warning”, if the purge valve is open. If the system
senses any pressure increase (even minor) during purging, the command will be
terminated. Make sure that the valve has been sufficiently loosened and try again.
Instrumental operation: ICS 3000
Standard Operational Procedure
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Figure 5: Pump tab. Relevant functions marked for purging, flow settings ect.
Once the purging is done, close the purge valve again tightly to redirect the flow path to the column. If
the valve is not tightened enough, the pressure during operation will not be sufficiently high meaning
that the flow delivery to the column is insufficient. No warnings about low pressure will occur, unless
pressure is completely lost (less than 200 psi) so please do observe that correct pressure is obtained
during start up. Normal operating pressure for CarboPac PA1 incl. guard is at least 15-1700 psi (this
includes a pressure restrictor coil at 1000 psi).
Operating the autosampler
Samples are placed in the autosampler prior to operation. The front door of the autosampler is opened
(held in place by a relatively strong magnet – pull it hard!) and the vials placed in the sample tray (figure
6). On the outside of the autosampler in the lowest left corner is the main power switch – not a door
opener! Only press this button if you really want to cut the power to the autosampler. Sample position
in the tray is straight forward numbering from 1 to 100. Each position is marked on the tray. The tray
may be removed from the autosampler when samples are being loaded (be careful…). The sample tray is
placed in a cooled compartment and the autosampler will not operate if the front door is open. A sound
alarm will occur if the door is left open (turns off again when the door is closed). Don’t open the door if
the robot arm is moving as this will immediately interrupt the movements. The front panel will inform
Prime/purge
function
Pump motor
Flow settings
Pressure
indicator
Solvent
distribution
Module connection
Retention time count
Instrumental operation: ICS 3000
Standard Operational Procedure
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about the “state” of the autosampler: “Idle” means that the autosampler can be opened for vial
handling (this is also the case for idle status during a sequence run). Otherwise, the front panel is used
for manual handling and is not used during routine operation. All normal operational control is handled
via CM. Figure 8 shows the routine operations of the autosampler from CM. The sampler has a fixed
volume loop and if this needs to be changed, call a super user. Partial injection may be applied if
necessary, but will result in increased standard deviations, but please consult super user or manual. NB:
Be aware that the sampler consumes approx. 40 µL sample for a 2 µL injection.
On top of the autosampler (towards the back) is the syringe tower (figure 6). This is where sample,
buffer etc. is taken and measured before injection to the column. Make sure no air bubbles are trapped
in the syringe by priming sampler before you start your analysis. The syringe is connected to a wash
solution consisting of 20% methanol.
The sample is introduced via the injection port, via the diverter valve (position 1 directs sample to
system 1), via the loop (flow path is via the loop when the injection valve is in inject mode, meaning
always except during an injection cycle) to the column. When the needle leaves the inject port (figure 7)
the bed should be dry with no droplets/spillage. If not, something is obstructing the injection and
poor/no signals will be observed in the following chromatogram. The needle is washed before each
injection in the flush port (figure 7) next to the injection port. Here, wash solution will appear (and may
spill) which is normal.
Instrumental operation: ICS 3000
Standard Operational Procedure
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Figure 6: Autosampler with sample tray dismounted from the tray compartment
Figure 7: Injection and flush port in autosampler
Diverter valve
Instrumental operation: ICS 3000
Standard Operational Procedure
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Figure 8: Autosampler tab in CM.
Operating the column compartment
The column compartment is the module where the column is connected (figure 9). During operation the
front cover of the column compartment should preferably be closed and especially if temperature
control is applied (improved reproducibility is observed when temperature is set slightly above
ambient). As mentioned above the flow is directed from the autosampler via the left injection valve (for
pump 1) to the column. The loop is also mounted on the injection valve. Do not mess around with the
tube connections here! The column is fitted in an appropriate holder in the back of the compartment.
Prime syringe
Temperature control in sample tray
Autosampler status
Sample loop size
flush sample
Diverter valve/inject valve
Instrumental operation: ICS 3000
Standard Operational Procedure
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Figure 9: Internals of column and detector compartment. Blue arrow indicates flow direction on the column, which should fit
with column flow directions. Upper thrid of the compartment is not present in this particular instrument set up.
When a column is fitted and flow is applied please ensure that there are no leaks. Leaks at the column
inlet will result in pressure decrease, whereas leaks at the column outlet may not affect the pressure but
will affect the detector signal and potential flooding of compartment. If leaks are observed or registered
by the system, further tighten the connections either by hand or by a wrench key (be careful, PEEK
connections are weak).
Column temperature is specified in CM (figure 10) and the desired temperature is written in the field for
“Set”. Press Enter to execute the command. The field “Actual” will inform about the actual temperature
in the column compartment (which initially is not necessarily the same as the column temperature).
Analytical Column flow direction
Left injection valve for Pump Guard column – Note flow
direction: Left to right!
Analytical column – Note
flow direction: Right to left!
Instrumental operation: ICS 3000
Standard Operational Procedure
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Figure 10: Column compartment tab for controlling column temperature.
Operating the PAD detector
The detector is placed as can be seen in figure 9 (item 2) and a close-up photo in figure 11. The detector
consists of a gold working electrode and a pH reference electrode. The pH reference electrode (number
3 in figure 11) has very limited needs for maintenance during routine operations and should not be
removed from its socket. The working electrode on the other hand is disposable and needs to be
changed regularly. Routines for changing working electrode are described below. The detector can only
be turned on when flow is on the system and a pump link exists where the cell voltage on the electrode
is turned off, if the flow is stopped. However, cell voltage should only be turned on during sequence
running and other diagnostic runs, since prolonged voltage will decrease the electrodes life time. When
running, the electrode senses two signals, a total signal (a background signal that depends on the type
of eluents and state of the electrode) and a baseline/real sample signal which can be autozero’ed. The
detector responds to any substances that are either oxidizable or reducible and the electrical output
results from an electron flow caused by the chemical reaction that takes place at the surface of the
electrodes. The eluents are highly charged and this can be observed as the background (Total) signal.
Normal operating Total Signal should be between 15 and 35 nC. The reference electrode is capable of
Temperature control
Instrumental operation: ICS 3000
Standard Operational Procedure
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adjusting/compensating changes in background signal when gradient elution profiles are run and in that
sense maintain a stable baseline. If the reference electrode is not installed/out of order, the baseline will
rise considerably when gradient profiles are used. PAD is a particularly strong detection principle for
carbohydrates, because the highly alkaline eluents will charge the hydroxyl groups and therefore give
rise to signals in the detector. Furthermore, this detection principle is highly sensitive and therefore
makes it superior for carbohydrate analysis.
Figure 11: PAD electrode
From column
To waste
Instrumental operation: ICS 3000
Standard Operational Procedure
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Figure 12: PAD tab in CM.
Changing working electrode
When the Total Signal becomes too high (above 40-45 nC) or the signals during running behaves strange
(extremely high signal or seemingly no signal at all/increased background signals), changing the working
electrode may be needed (The electrode has a life time normally between 1 and 3 weeks continuous
operation). If you suspect the electrode may be worn out, check last date of change in the logbook to
verify. New users should consult a super user before changing the electrode the first time. Hereafter
do as follows:
1. Turn off the flow and thereby the cell voltage.
2. Disconnect the inlet and outlet of the detector (by the incoming and out coming of the red
tubing in figure 11). You may need a wrench key.
3. Unplug the blue and the yellow power plugs (figure 11).
4. Grab the electrode by two hands and pull it firmly towards you in one motion (yes, it will come
off).
5. While performing the following steps, try to keep the entire electrode in an upright position. The
pH reference electrode contains KCl which may leak if turned upside down.
6. Loosen the big knob (picture A, figure 13), press the two side-wings on the clips that holds the
spacer block in place and take off the entire clips.
Turning on/off cell voltage
Signal and Total Signal (background)
Autozero (of Signal)
Instrumental operation: ICS 3000
Standard Operational Procedure
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7. Remove the spacer block and now the individual parts of the electrode are visible (picture B,
figure 13). The working electrode is the disposable one but take EXTRA care of the gasket. They
are not supplied enough to be changed every time. Unless it is torn or otherwise damages it
should be reused.
8. Inspect the working electrode on the “gold”-side. Salt precipitations or loss of connection
between top and bottom is reason for loss of signal. Discard the electrode.
9. Inspect the cell body and carefully rinse it if visible salt precipitations are seen (picture B, figure
13).
10. Take a new working electrode from the package, but ONLY lift it in the snip of white tape that
goes outside the actual electrode which says “lift here”. Use a pair of tweezers if necessary.
11. Place the working electrode and the gasket back on the cell body. Note that the two positioning
holes in the gasket are different in shape, which compared to the spacer block indicates the
positioning. The same goes for the working electrode. Furthermore, the electrode should be
placed so that the golden parts face the cell body and so that the small circle fits over the slit in
the gasket. Do not bend the working electrode in any way! Position it precisely over the two
rods and slide it onto the cell body. See correct placement of the working electrode in picture C,
figure 13.
12. Place the spacer block and the clips back on the electrode and turn the big knob until you hear a
click. This is as tight as it gets.
13. Reposition the entire electrode in the detector compartment (it needs to say click), re-plug the
electrical cords (yellow in yellow, blue in black!), reconnect column and waste tubings, restart
flow on system (make sure there are no leaks!!) and set cell voltage on.
14. Monitor baseline for a while (approx. 20-30 min). The signal can be quite unstable in the
beginning (and very high), but will eventually stabilize. Total signal should preferably stabilize
between 15 and 30 nC.
Figure 13: Changing disposable
gold electrode
Working
electrode
Spacer block
Gasket
Cell body Spacer block
Spring- clips
A B C
Instrumental operation: ICS 3000
Standard Operational Procedure
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Waste collection
From the detector the flow stream is passed to a waste container standing in the exhaust cabin
underneath the system. This waste container needs to be checked regularly and emptied. The waste is
almost exclusively consisting alkaline salts and can therefore be poured in the drain. For the same
reason – spillage of waste will generate vast amounts of salt precipitations and should therefore be
avoided.
Starting up for operation
1. Make sure that eluent level is good and check the last date for eluent preparation. If the eluents
are more than 2 days old, consider if you should make new eluents before your run. If preparing
new eluents, remember to purge all channels in use (A,B and C) after eluent preparation
2. Check that the correct column is on.
3. Check that flow is on and pressure correct (column flow: 0.25 ml/min, pressure: 1500-1700 psi)
4. If column temperature control is applied make sure that the column compartment is closed
(lower door).
5. Turn on the detector cell (apply cell voltage).
6. Start “Baseline Monitoring” (see figure 14) and watch the detector signal stabilize.
7. If the signal does not stabilize or if the total signal is very high (above 45 nC) it may be time to
change the working electrode. Follow the procedure for doing so, if necessary.
8. Prime sampler, flush sample loop (make sure no air is entrapped in sample syringe)
9. Place the samples in the autosampler and prepare the sequence (see section below). If starting
up after eluent change, the first sample should ALWAYS be a blank sample running the
instrument method: “Start after eluent change”. This is to ensure regeneration of column after
possible carbonate contaminations. Afterwards condition the column with 1-2 blank runs with
the instrument method you intend to use before injection of the first real sample/standard.
These conditioning blanks should always be run no matter if eluent has been changed or not.
10. Stop baseline monitoring.
11. Start the run either by starting the sequence directly or by queuing the sequence (add to queue
in the “Queue” tab). Press “Ready Check” to get an estimate of eluent use during the entire
queue.
Instrumental operation: ICS 3000
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Figure 14: Queue tab. Here sequences can be queued for automatic start and general checking of system status (ready check)
is performed.
Setting up a sequence in CM
The data structure in CM is shown in figure 15, which is initially divided between different instruments.
The PAD detector is set up to system ED_1. Hereafter, a general data folder contains a folder for each
user, specified by name. Maintain this order of data structure and save your sequence under your own
name.
Monitoring baseline.
Monitor pump pressure Queue tab. For
queuing several
sequences
Adding a sequence
to the queue.
Ready check. Testing
if the system is
ready for operation.
Gives an estimate of
eluent use.
Successful ready check
Instrumental operation: ICS 3000
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Figure 15: Data structure in CM. Note the switch between Instrument and Data panel.
When making a sequence, choose “Create” from the top menu and choose “Sequence”. Hereafter a
wizard will start, that will guide you through setting up of a sequence:
1. Choose system: ED_1
2. Write a recognition name that it will be copied to for each sample (may be edited later), choose
number of samples, number of injections pr. sample (most often 1), position of 1st
vial and
injection volume (uL), normally the same as the sample loop size.
3. Choose Instrument Method (analytical run method) by accepting the lastly used method or by
browsing through folders. Choose Processing Method if applicable. Can also be added later. Set
default channel to ED_1.
4. Write a comment describing the particular samples (not obligatory).
5. Press finish and save the sequence to your folder under an appropriate name starting with the
following date format: YYMMDD_
Figure 16 shows and example of a prepared, finished sequence. Most fields in the sequence can be
edited before, during and after the run. This includes: Name, Type (blank, unknown, standard), Level
(only relevant for quantification), Position (vial position in the autosampler), Volume (injection
volume), Instrument Method (analytical run method, can only be edited before running) and
Switching between
Instrument and Data panel
Instrumental operation: ICS 3000
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Processing Method (qualititaive/quantitative method). Some fields have scroll down menus, others
are text fields. Always press “Save” after editing any fields. A shortcut called “Fill Down” is made for
fast assigning sample position in the autosampler. Mark the “Position” field from the position of the
first vial, press F9 and choose “Renumber”. Press enter. Alternatively, select the Fill Down menu
(figure 17), choose Renumber and write the name of the first sample position. Press OK. Note that
the fill down function may be used in all the other columns as well.
Figure 16: Prepared, finished sequence.
IMPORTANT: Changing any of these fields mentioned above will not affect the
results, but changing the field “Status” will erase data behind, if the samples have
been run; “Idle” indicates a sample not yet analysed, “Finished” indicates a sample
analysed and therefore a file containing data. “Interrupted” is a sample stopped
during analysis due to some sort of error message, may also contain data, but
sometimes not useful data.
Instrumental operation: ICS 3000
Standard Operational Procedure
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Figure 17: Fill down menu for fast sample position assignment. Press “Fill Down” or F9 to open window.
Alarms and warming
Pressure alarm
If the pressure falls below or reaches the upper limits of the settings (which are mainly set to protect the
column) a pressure sound alarm will be initiated. At the same time, all flow will be stopped. High
pressure may occur if the samples contain insoluble particulate material, if the solvent concentration
suddenly changes, temperature control fails or if the column is overloaded with sample due to many
injections without regeneration. Loss of pressure may occur if the eluent bottles are empty, air is
introduced in the system or if leaks happen on the “upstream”/pressure side of the column. If leakage is
the cause, the system will detect the compartment where this is occurring via leak detectors and the
audit trail will inform about this.
Identify the source of the pressure violations, take proper action and restart the system.
Leak/humidity alarm
Identify the reason for leakage (which compartment) and correct if possible. Dry up any spillage in the
compartments with soft tissue paper.
Fill Down menu
Marked cell for
1st
vial position
Instrumental operation: ICS 3000
Standard Operational Procedure
Jagger
No pressure?
Check that the purge valve is properly closed. Check that all fittings are securely tightened. If this is not
enough purge the channels as air may have been trapped in the pump.
No connection to modules?
Check that the Instrument Controller is running (figure 2). When this is positivly running, reconnect the
non-connected modules by pressing “Connect” in each module tab (see figure 5 or 8). If there is still no
connection restart the computer and start the Instrument Controller again. Still no connection – call
security.
Software error messages
Close all active windows and restart CM. Still software errors, restart computer and start the Instrument
Controller again. Still errors – call security.
No contact with Autosampler
The autosampler needs a restart once in a while. If no contact can be achieved through the software,
turn off the autosampler (power button lower left corner, front). Leave it off for 1-2 minutes and start
again. It will now perform a few self tests (this takes a couple of minutes. When this is done, reconnect
in CM. If there is still no connection, call security.
Call security
For more advanced guidance either contact Super users or consult the general equipment manuals
found on the CD-rom: ”Reference Library” provided by Dionex.
Super users:
Jane Wittrup Agger ([email protected]; 6496 5856)
Bjørge Westereng ([email protected]; 6496 5909)