AKTA Configurations Handbook

Optional Configurations
ÄKTAdesign
User Manual
18-1139-57
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____________________________________________________ Contents
Contents
1 Introduction
1.1 General ...............................................................................................................7
1.2 Scope of manual .................................................................................................9
1.3 Summary of optional configurations ................................................................10
1.4 Safety ...............................................................................................................23
2 ÄKTAexplorer
2.1 General .............................................................................................................25
2.2 Unpacking ........................................................................................................25
2.3 Pre-requisites ....................................................................................................25
2.4 General guidelines ............................................................................................26
2.5 Fraction collector Frac-901 ..............................................................................32
2.6 Fraction collector Frac-950 ..............................................................................36
2.7 Autosampler A-900 ..........................................................................................42
2.8 Flow direction valve .........................................................................................60
2.9 General function valve ......................................................................................63
2.10 pH flow cell and pH electrode mounting ..........................................................66
2.11 Sample pump P-950 and Sample valve V5 ........................................................69
2.12 Air sensors ........................................................................................................93
2.13 Auxiliary equipment controlled via Pump P-901/903 REMOTE connector ....102
2.14 Superloop .......................................................................................................105
2.15 Configuration dependent delay volume in ÄKTAexplorer ..............................107
2.16 Strategy and template overview in ÄKTAexplorer ..........................................109
3 ÄKTApurifier
3.1 General ...........................................................................................................111
3.2 Unpacking ......................................................................................................111
3.3 Pre-requisites ..................................................................................................111
3.4 General guidelines ..........................................................................................112
3.5 Fraction collector Frac-901 ............................................................................118
3.6 Fraction collector Frac-950 ............................................................................122
3.7 Autosampler A-900 ........................................................................................128
3.8 Column selection valves .................................................................................146
3.9 Buffer selection valves ....................................................................................149
3.10 Flow direction valve .......................................................................................152
3.11 General function valve ....................................................................................156
3.12 pH flow cell and pH electrode mounting ........................................................159
3.13 Sample pump P-950 and Sample valve V5 ......................................................161
3.14 Air sensors ......................................................................................................183
3.15 Auxiliary equipment controlled via Pump P-901/903 REMOTE connector ....192
3.16 Superloop .......................................................................................................195
3.17 Configuration dependent delay volume in ÄKTApurifier ...............................197
3.18 Strategy and template overview in ÄKTApurifier ...........................................199
ÄKTAdesign Optional Configurations

User Manual 18-1139-57 Edition AB v
Contents ____________________________________________________
4 ÄKTAbasic
4.1 General .......................................................................................................... 201
4.2 Unpacking ..................................................................................................... 201
4.3 Pre-requisites ................................................................................................. 201
4.4 General guidelines ......................................................................................... 202
4.5 Fraction collector Frac-901 ............................................................................ 208
4.7 Autosampler A-900 ....................................................................................... 218
4.8 Outlet valve ................................................................................................... 236
4.9 Column selection valves ................................................................................. 242
4.10 Buffer selection valves .................................................................................... 245
4.11 Flow direction valve ...................................................................................... 248
4.12 General function valve ................................................................................... 252
4.13 Sample pump P-950 and Sample valve V5 ..................................................... 255
4.14 Air sensors ..................................................................................................... 277
4.15 Auxiliary equipment controlled via Pump P-901/903 REMOTE connector ... 286
4.16 Superloop ...................................................................................................... 289
4.17 Configuration dependent delay volume in ÄKTAbasic .................................. 291
4.18 Strategy and template overview in ÄKTAbasic .............................................. 293
5 ÄKTAFPLC
5.1 General .......................................................................................................... 295
5.2 Unpacking ..................................................................................................... 295
5.3 Pre-requisites ................................................................................................. 295
5.4 General guidelines ......................................................................................... 296
5.7 Column selection valves ................................................................................. 312
5.8 Outlet valve ................................................................................................... 315
5.9 Buffer selection valves .................................................................................... 321
5.10 General function valve ................................................................................... 325
5.11 pH flow cell and pH electrode mounting ....................................................... 329
5.12 Sample pump ................................................................................................. 331
5.13 Air sensors ..................................................................................................... 351
5.14 Auxiliary equipment controlled via Pump P-920 REMOTE connector .......... 360
5.15 Superloop ...................................................................................................... 363
5.16 Configuration dependent delay volume in ÄKTAFPLC ................................... 365
5.17 Strategy and template overview in ÄKTAFPLC ............................................... 366
6 Accessories and consumables

6.1 Connections, chromatography – standard selection ....................................... 367
6.2 Optional components and consumables ........................................................ 368
Short instructions on back page

vi User Manual 18-1139-57 Edition AB
______________________________________________ Introduction 1
1 Introduction
1.1 General
ÄKTA™ design standard system configurations can be changed to
optional configurations. This flexibility in the ÄKTAdesign system
strategies allows the user to enhance already used purification methods
and also to develop new, more complex methods.
P-950
IV/PV-908 INV-907
When optional equipment is connected to a standard chromatographic

system, new sets of instructions to control the optional equipment
become accessible for the user.
Optional configurations are selected, installed and put into operation

by the user. An optional configuration consists of both hardware
components and software instructions.
Optional configurations are monitored and controlled via methods run

by the UNICORN™ control system in the same way as the ÄKTAdesign
standard system configurations.
Optional configurations are shown in the flow scheme displayed in the

System Control in UNICORN. Components used in an optional
configuration are highlighted in the flow scheme, whereas components
not installed are hidden. The status of the components is also shown,
e.g., valve position.
Note: The actual location of, for example, an optional valve is not
shown in its proper flow path position. This must be taken into account
by the user. However, a new flow path picture can be ordered with the
optional configuration parts included, and installed together with the
additional software to reflect the optional system configuration.

User Manual 18-1139-57 Edition AB 7
1 Introduction ______________________________________________
Generally speaking, there are two types of optional configurations.

Completely user-defined configurations and dedicated configurations
partially pre-defined and prepared at the factory, requiring user actions
in their final implementation.
Components used in completely user-defined configurations are

designated with general alpha-numeric position tags. For example,
Valve8 refers to the valve corresponding to the component identity code
8 in UNICORN.
Components used in dedicated configurations are referred to in plain

language, e.g. OutletValve, which is the same designation as the actual
instruction name used in UNICORN.

8 User Manual 18-1139-57 Edition AB
______________________________________________ Introduction 1
1.2 Scope of manual
To support the process of implementing optional configurations,
general guidelines regarding installation and operation are given in this
manual.
For specific information regarding function, maintenance, trouble-

shooting and spare parts, refer to the User Manual and/or Instruction
for the respective component, as well as the System Manual for your
ÄKTAdesign system.
This manual is divided into four main sections reflecting the

ÄKTAdesign families:
• ÄKTAexplorer (Main Section 2 ÄKTAexplorer)
• ÄKTApurifier (Main Section 3 ÄKTApurifier)
• ÄKTAbasic (Main Section 4 ÄKTAbasic
• ÄKTAFPLC ™ (Main Section 5 ÄKTAfplc)
Select the main section describing your ÄKTAdesign system and find
the sub-section describing the installation and operation of the optional
configuration you have decided to use.
On the back page, there is a short instruction giving guidelines for the
general steps in adding and removing optional configurations to your
ÄKTAdesign system.

1 Introduction ______________________________________________
1.3 Summary of optional configurations

An overview of all optional configurations available for each
ÄKTAdesign system is summarized below.
ÄKTAexplorer
Optional configurations supported by ÄKTAexplorer are:
• Connecting Fraction collector Frac-901.
• Connecting Autosampler A-900.
• Connecting a flow direction valve.
• Connecting a sample selection valve.
• Connecting up to 2 motorized multi-port optional function valves.

The actual use is completely user-defined.
• Connecting on-line pH measurement.
• Sample pump P-950.
• Connecting up to three air sensors.
• Connecting external equipment using digital input/output signals

through the system pump P-900 REMOTE connector.
• Connecting a Superloop™.
Fraction collector Frac-901

Fraction collector Frac-901 is an automated fraction collector for use
in ÄKTAexplorer chromatography systems under UNICORN control.
The functionality and performance is aimed for mid-range usage.

in ÄKTAexplorer chromatography systems under UNICORN control.
The functionality and performance is aimed for high-end usage.

______________________________________________ Introduction 1
Autosampler A-900
Autosampler A-900 used in ÄKTAexplorer enables:
• Automated multiple sample injections from a few microlitres to

one millilitre of sample solution.
• Quantification of amount, concentration and molecular size
• External standard quantification .
• Internal standard quantification.
• Standard addition.
• Recovery calculations.
• Molecule size calculations.
Quantification uses peak data from standards to produce

calibration curves which can then be used to evaluate the amount
and concentration of components in a sample. The molecular size
function determines the molecular size of components in a sample
using a molecular size curve prepared from one or more standards.
Note: Quantification requires additional software not included in

the standard package of Autosampler A-900.
Valves
• One INV-907 valve can be connected as a flow direction valve to
select flow direction through the column.
• One PV-908 valve can be connected as a sample selection valve.

This is commonly used in combination with an optional sample
pump P-950.
• Two optional valves, freely selected from INV-907, a seven-port,

three-way valve that can be used, for example, as a second injection
valve, and IV/PV-908 valves, which are eight-port rotary valves
with different pressure limits and internal flow path diameters.
On-line pH measurement
pH measurement is accomplished in ÄKTAexplorer by a pH flow cell
mounted in the main system flow path after the UV and conductivity
flow cells. The pH flow cell is fitted with a pH electrode for on-line
measurement. The pH electrode can be replaced by a dummy electrode
when pH measurement is not required.
Sample pump
A P-950 sample pump can be connected to accomplish automatic
sample application when using the same sample several times. P-950 is
a single-channel membrane laboratory pump that can be used to apply
samples directly on the column or for filling sample loops (including
Superloop).

1 Introduction ______________________________________________
Its flow rate is controlled and the pump pressure is monitored from
UNICORN.
The sample pump P-950 is commonly used in combination with a

multiport motorized valve PV-908 connected to the injection valve inlet
to enable automatic sample application of several different samples.
Air sensors
The use and location of the air sensors is user-defined. For example:
• One air cell can be used to detect air when automatic sample
application using P-950 is performed.
• One air cell can be used to detect air when applying large amounts
of sample with the system pump.
There are two types of air cells available. They are designated A-912
and A-925 and differ in inner diameter. This makes them suitable to use
with different sizes of capillary.
A-925 has a larger inner diameter and is suitable to use in the inlet flow
path to the system pump P-900.
A-912 has a smaller inner diameter and is suitable to use in the P-950
sample flow path.
Up to three detection points can be used in the process flow path. The
detectors are designated AS1, AS2 and AS3 in the flow scheme.
Note: In ÄKTAexplorer 100 Air systems, use of the AS2 cell detectors
is supported by method templates included in the standard
configuration.
Auxiliary equipment via Pump P-900 REMOTE connection

The system pump P-901/903 is provided with a digital input/output
connection comprising four output and four input channels. The
channels are isolated and accept standard TTL-level signals.
Examples of use include:
• External alarm input/output.
• Start/stop of external equipment.
Superloop
Superloop permits the introduction of larger volumes of sample
(1–150 ml) onto the column.

______________________________________________ Introduction 1
ÄKTApurifier
Optional configurations supported by ÄKTApurifier are:
• Connecting Fraction Collector Frac-901.
• Connecting column selection valves.
• Connecting a buffer selection valve.
• Connecting a flow direction valve to control flow direction through

the column.

• Connecting Sample pump P-950.

• Connecting a Superloop.

in ÄKTApurifier chromatography systems under UNICORN control.
The functionality and performance is aimed for mid-range usage.

in ÄKTApurifier chromatography systems under UNICORN control.
The functionality and performance is aimed for high-end usage.

1 Introduction ______________________________________________
Autosampler A-900
Autosampler A-900 used in ÄKTApurifier enables:

• Quantification amount, concentration and molecular size:
• External standard quantification.


Valves
• Two PV-908 rotary valves used in conjuction to switch between
multiple columns. The valves are controlled synchronously by one
common instruction.
• One IV-908 rotary valve for buffer selection. Used to switch

between different buffer solutions.


pump P-950.

pH measurement is accomplished in ÄKTApurifier by a pH flow cell
measurement. The pH electrode can be replaced by a dummy electrode

______________________________________________ Introduction 1
Sample pump with sample selection valve
Superloop). Its flow rate is controlled and the pump pressure is
monitored from UNICORN.
The Sample pump P-950 is commonly used in combination with a

multiport valve PV-908 connected to the injection valve inlet to enable
automatic sample application of several different samples.
Air sensors
sample flow path.

The system pump P-900 is provided with a digital input/output
Superloop

1 Introduction ______________________________________________
ÄKTAbasic
Optional configurations supported by ÄKTAbasic are:
• Connecting column selection valves.
• Connecting a buffer selection valve.
• Connecting an outlet valve.
• Connecting a flow direction valve to control flow direction through

the column.

• Connecting Sample pump P-950.


in ÄKTAbasic chromatography systems under UNICORN control. The
functionality and performance is aimed for mid-range usage.

in ÄKTAbasic chromatography systems under UNICORN control. The
functionality and performance is aimed for high-end usage.

______________________________________________ Introduction 1
Autosampler A-900
Autosampler A-900 used in ÄKTAbasic enables:

• Quantification amount, concentration and molecular size:
• External standard quantification.


Valves
multiple columns. The valves are controlled synchronously by one
common instruction.
• One PV-908 rotary valve used to divert between waste and seven
positions, to collect flow-through, or to collect several large
fractions.
• One IV-908 rotary valve for buffer selection. Used to switch

between different buffer solutions.


pump P-950.


1 Introduction ______________________________________________
Sample pump with sample selection valve

The sample pump P-950 is commonly used in combination with a

multiport valve PV-908 connected to the injection valve inlet to enable
automatic sample application of several different samples.
Air sensors
sample flow path.

Superloop

______________________________________________ Introduction 1
ÄKTAFPLC
Optional configurations supported by ÄKTAFPLC are:
• Connecting up to 8 motorized multi-port valves. These valves can

be used to accomplish the following functions:
• Column selection.
• Outlet diversion.
• Buffer selection.
• General function. The actual use is completely user-defined. For

example, they can be used as a reversed flow valve or a sample
selection valve.
• Connecting a Sample pump P-950.
• Connecting external equipment, such as a pump P-950 for large

sample application using digital input/output signals through the
system pump P-920 REMOTE connector.
• An extra rack panel is available as an accessory to accommodate

optional equipment.

in ÄKTAFPLC chromatography systems under UNICORN control. The
functionality and performance is aimed for mid-range usage.

in ÄKTAFPLC chromatography systems under UNICORN control. The
functionality and performance is aimed for high-end usage.

1 Introduction ______________________________________________
Valves
• Five PV/IV-908 valves can be connected as dedicated function
valves. These valves are used as follows:

multiple columns. The valves are controlled synchronously by
one common instruction.
• One PV-908 rotary valve used to divert between waste and seven
positions, to collect flow-through, or to collect several large frac-
tions.
• One or two IV-908 rotary valves for buffer selection. Used indi-
vidually to switch between different buffer solutions.
• Three optional valves, freely selected from INV-907, a seven-port,

three way valve that can be used, for example, as a reversed flow
valve or a second injection valve, and IV/PV-908 valves, which are
eight-port rotary valves with different pressure limits and internal
flow path diameters.
pH measurement is accomplished in ÄKTAFPLC by a pH flow cell
measurement. The pH electrode is replaced by a dummy electrode
Sample pump
The Sample pump P-950 is commonly used in combination with a

multiport motorized valve PV-908 connected to the injection valve
inlet to enable automatic sample application of several different
samples.

______________________________________________ Introduction 1
Air sensors
sample flow path.

Superloop

1 Introduction ______________________________________________
Extra rack panel
The extra rack panel is positioned on the vertical rods in the free space
between the lower and upper part of the system rack when the upper
part is raised to a higher position
Locking screws
Extra rack panel

for mounting
optional
equipment

______________________________________________ Introduction 1
1.4 Safety
• Components are designed for indoor use only.
• Do not use in a dusty atmosphere or close to spraying water.
• Operate in accordance with local safety instructions.
WARNING! When using hazardous chemicals, all suitable

protective measures, such as protective glasses, must be taken.
WARNING! Ensure that the entire chromatographic system has

been flushed thoroughly with distilled water before removing any
capillaries or components.
WARNING! Always disconnect the power supply before attempting

to replace any item on the equipment during maintenance.
WARNING! If there is a risk that large volumes of spilt liquid may

penetrate the casing of the equipment and come into contact with
the electrical components, immediately switch off the
chromatographic system and contact an authorized service
technician.
WARNING! Superloop 10 ml and Superloop 50 ml must not be

used at pressures above 4 MPa (40 bar, 580 psi). Superloop 150 ml
must not be used above 2 MPa (20 bar, 290 psi). At higher
pressures, the glass tube may shatter.
CAUTION! Make sure ÄKTAdesign is switched off before installing

the optional components. The mains power to ÄKTAdesign must be
switched OFF before disconnecting or connecting the UniNet-1 and
UniNet-2 cables.

_____________________________________________ ÄKTAexplorer 2
2 ÄKTAexplorer
2.1 General
To utilize optional equipment in a chromatographic run, suitable
instructions must be inserted in the method to be used. This is made
either by modifying existing method templates, or by creating a new
method. The general procedures for creating and editing method
templates are described in the UNICORN User Manual.
Note: The system configuration delay volume must be recalculated and

reset when the standard configuration after the UV flow cell is changed
to optional configuration, or vice versa. Details are given in section
2.15 Configuration dependent delay volume in ÄKTAexplorer.
The descriptions below deal with instructions used in specific functions

as indicated by the headings.
2.2 Unpacking
Unpack the optional components and check against the supplied
packing list. Inspect the items for obvious damage that may have
occurred during transportation.
CAUTION! Read the following information carefully, as well as all

the additional instructions supplied with the components, to ensure
that the ÄKTAexplorer optional equipment is installed correctly.
2.3 Pre-requisites
• UNICORN version 3.2 or higher must be installed in the computer
connected to your ÄKTAexplorer chromatography system.
• ÄKTAexplorer must be installed and fully tested before the
optional components are installed. See the Installation Guide for
your ÄKTAexplorer chromatography system.
WARNING! Ensure that the entire system has been flushed

thoroughly with distilled water before removing any capillaries or
components.
CAUTION! Make sure ÄKTAexplorer is switched off before

installing the optional components. The mains power to
ÄKTAexplorer must be switched OFF before disconnecting or
connecting the UniNet-1 and UniNet-2 cables.
ÄKTAdesign Optional configurations

2 ÄKTAexplorer ____________________________________________
2.4 General guidelines
Principle installation of components

Some optional components can be Mounting rails for optional
mounted in the mounting rails on the side components
of the modules. Both sides of the modules
are provided with rails for quick mounting
of components.
When selecting mounting location for the Monitor P-910
optional equipment it is important to

choose a location which minimize the
length of the capillaries used to connect
the optional components to the rest of the
Monito
r P-910
0, 6
system.
Many components that are attached to the

Mon
itor P-91
0
mounting rails uses a snap-in bracket. The

bracket is supplied separately with the
component and needs to be fitted as
shown below before the component can be attached.
2. Fit the attached

mounting bracket with
the two screws
Inserting components
1. Loosen the
two screws
Removing components
Capillaries are connected using unions as specified in section

6.1 Connections, chromatography – standard selection.

_____________________________________________ ÄKTAexplorer 2
All valves have a unique ID code which identifies
them to the UNICORN control system at system
start-up. This ID code should be checked before
installation.
The ID codes for the different function valves and the
corresponding instructions used in UNICORN to
control them are as follows:
Valve function ID code Valve type Instruction in UNICORN

Sample injection 1 INV-907 InjectionValve
Column selection 2 PV-908 ColumnPosition (combined
Column selection 3 PV-908 instruction controlling both
valves)
Outlet fractionation 4 PV-908 OutletValve, OutletFractions
(two instructions used for con-
trolling different type of frac-
tionation functions)
Sample selection 5 PV-908 SampleValve
Buffer selection 6 IV-908 BufferValveA1
Flow direction valve 7 INV-907 FlowDirectionValve
Optional function 8 INV-907, IV/PV-908 Valve8
Components are connected to the UNICORN control system using

either UniNet-1 or UniNet-2 cables*. Both the UniNet-1 and the
UniNet-2 data communication chain in standard configuration is
routed from the rear of P-900 via their respective components to the
last component in the chain where it is terminated with a plug.
Both the UniNet-1 and the UniNet-2 chain can be interrupted

anywhere between P-900 and the termination plug to interconnect the
optional components in the chain. The termination plug can be moved
to the last component (furthest away from P-900), if motivated by cable
routing considerations.
Te r m i n a t i o n
Plug
Fr a c t i o n
collector
(optional)
To UniNet 2
Computer Chain
UniNet 1
Chain
*
Frac-901, Frac-950 and A-900 are connected via the UniNet-1 chain. All
other optional components are connected via the UniNet-2 chain.

2 ÄKTAexplorer ____________________________________________
Components instruction overview

To enable optional components to be used, UNICORN must be set up
to include the optional component instructions in system control. This
is made in the System Settings:Component... list.
To control the optional components in a run, UNICORN control

instructions must either be inserted into a method by the user, or the
instructions are pre-defined in a method template supplied with the
optional component. For an overview of available method templates,
see 2.16 Strategy and template overview in ÄKTAexplorer.
In the following, a list of the most commonly used instructions for

optional components is given.
Function/
component Type UNICORN module Instruction Parameter
Fraction Frac-901 System:Settings:Special FracParameters_900 DelayVol (0-10 ml)

collectors TubeChange (Tube, DropSync,
WasteBetweenTubes)
FracNumberingMode_900 Mode (Reset, Continue)
Peak_FracParameters_900 Start_Slope, EndSlope, Min-
Width
Method/Manual:Frac Fractionation_900 FracSize (0-50 ml)

FractionationStop_900 -
FeedTube_900 -
ResetFracNumber_900 -
Peak_Fractionation_900 PeakSize (0-50 ml)
Peak_FracStop_900 -
Frac-950 System:Settings:Special FracParameters DelayVol (0-10 ml)
TubeChange (Accumulator,
DropSync, Tube)
Peak_FracParameters Start_Slope, EndSlope, Min-
Width
Method/Manual:Frac Fractionation Tube type (name from rack defi-

nition)
FracSize (0-99999.999 base)
Start at (First tube, Next tube,
Next line, Next group, Tube
number)
Base (Volume, Time)
FractionationStop -
FeedTube -
Peak_Fractionation Tube type (name from rack defi-
nition)
PeakSize (0-99999.999 base)
number)
Base (Volume, Time)
Peak_FracStop -
AccumulatorWash Strokes (1–10 (5),OFF)

_____________________________________________ ÄKTAexplorer 2
Function/ Type UNICORN module Instruction Parameter
component
Frac-950 Manual:Frac Man_Fractionation Rack (name from rack defini-
(contin- tion)
ued) Frac_order (Serpentine_row,
Row-by-row, Serpentine-col-
umn, Column-by-column)
Tube type (name from rack defi-
nition)
number)
Base (Volume, Time)
Man_Peak_Fractionation Rack (name from rack defini-
tion)
Frac_order (Serpentine_row,
nition)
number)
Base (Volume, Time)
Rack (name from rack defini-
tion)
nition)
Ignore_LastTube -
Reset_Frac_Position -
Flow direc- INV-907 Manual:Flowpath FlowDirection Direction (DownFlow-UpFlow)
tion valve
Sample PV-908 Manual:Flowpath SampleValve Position (1-8)
selection
valve
Optional valve INV-907, Manual:Flowpath Valve8 Position (INV-907: 1-3)
8 IV/PV-908 Position (IV/PV-908: 1-8)
Sample pump P-950 System:Settings:Monitors WatchPar_SamplePressure Delta_Peak (0-1 MPa)
Delta_Base (0-1 MPa)
WatchPar_SampleFlow Delta_Peak (0-50 ml/min)
Delta_Base (0-50 ml/min)
System:Settings:Alarms Alarm_SamplePressure Mode (Disabled, Enabled)

HighAlarm (0-1 MPa)
LowAlarm (0-1 MPa)
Method/Manual:Pump SampleFlow FlowRate (0-50 ml/min)

DirectLoad SampleFlow (0-50 ml/min)
Volume (0-1000 ml)
ReducedFlow (No, Yes)
MinFlow (0.1-49.9 ml/min)

2 ÄKTAexplorer ____________________________________________

component
Auto sampler A-900 System:Settings:Special AutosamplerParameters LoopVol (10-1000 µl) [30 µl]
SyrVol (100, 250, 500, 1000 µl)
FlushVol (30-100 µl) [40 µl]
WashVol (300-999 µl)
TrayType (StandardVial A,
MicroVial D)
HeadPress (Yes, No)
NeedleHght (0-40 mm) [2 mm)
AutosamplerSyringeSpeed Speed (Low, Normal, High)
Scaling (0.1 - 1.0)
Manual:Flowpath AutosamplerControl Command (InitialWash, Rota-

teTray, SyringeHome, Syrin-
geEnd, NeedleHome,
NeedleFront)
InjectionPartial VialNo (1-160)
InjVolume (1-500 µl) [10 µl]
WashExt (Yes, No)
AirSegment (Air, NoAir)
InjectionFlushed VialNo (1-160)
WashExt (Yes, No)
InjectionPickup VialNo (1-160)
TrpVialNo (1-160)
WashExt (Yes, No)
QuantitationData QuantitationData (Sample,
Level_1_Standard,
Level_2_Standard....
Level_20_Standard)
Air sensor1 A-9001 System:Settings:Monitors AirSensor1-3 Mode (High_Sensitivity,
Air-912 Medium_Sensitivity,
Air-925 Low_Sensitivity)
Flow from (SystemPump,
SamplePump)
Manual:Alarms&Monitors , Alarm_AirSensor1-3 Mode (Enabled, Disabled)1

or System:Settings:Alarms
Test Equal (Value 0, 1)
View:Windows:Instruction Watch_AirSensor1-3
Action (<Block Name>/
box:Watch
READY/PAUSE/
HOLD/END_BLOCK/
END_METHOD/
CONTINUE)
Auxiliary P-900 System:Settings:Special AuxOut1 Signal (0, 1)
equipment REMOTE or Manual:Pump AuxOut2 Signal (0, 1)
I/O AuxOut3 Signal (0, 1)
AuxOut4 Signal (0, 1)
View:Windows:Instruction Watch_AuxIn1-4 Test Equal (0, 1)

box:Watch Action (<Block Name>, READY,
PAUSE, HOLD, END_BLOCK,
END_METHOD, CONTINUE)

_____________________________________________ ÄKTAexplorer 2
Preparation for use
To start up your ÄKTAexplorer optional configuration system:
1 Switch on ÄKTAexplorer with the mains switch located at the

front left on the system base.
2 Check that the computer and printer are switched on.
3 Log in (see chapter 2 in Making your first run).
4 Make sure you have the correct strategy installed in your system,
see 2.16 Strategy and template overview in ÄKTAexplorer. A
new strategy is installed according to the software installation
instructions given in the UNICORN User Manual.
5 In the Main Menu, select Administration:System Setup.
6 Select System and then Edit. Click Component...
7 From the Component list, select the optional component(s) you

have installed and click Add>>. Click OK twice.
The instructions associated to the new component appears when
creating a new method or editing an existing method.
8 The correct tubing dimension for the column you intend to use
must be installed. For most columns the standard tubing can be
used. In complex configurations, using high flow rates, it may be
necessary to replace the standard tubing with a larger i.d. PEEK
tubing.
Note: If tubing with too large inner diameter is used, the peaks will
become broader than necessary. If tubing with too small inner diameter
is used, the back-pressure from the tubing might become higher than
the maximum pressure for the column and the run will stop
immediately after it is started.

2 ÄKTAexplorer ____________________________________________
2.5 Fraction collector Frac-901
General
Frac-901 can be used for both small scale and preparative scale
purifications with ÄKTAexplorer. It collects up to 175 fractions in
12 mm diameter tubes, up to 95 fractions in 10–18 mm diameter tubes,
and up to 40 fractions in 30 mm diameter tubes.
Frac-901 allows fixed volume fractionation or automatic peak

fractionation. The latter function is based on peak detection using slope
sensing. Fraction marks and fraction numbers make it easy to identify
fractions and peaks. Fast tube change minimises spills between tubes,
eliminating it entirely below flow rates of 5 ml/min.
Drop synchronisation eliminates sample loss during tube change.
Installation
1 Unpack and assemble Frac-901 according to the instructions given
in the Fraction Collector Frac-900/901 User Manual.
2 Place Frac-901 to the right of the system.
Shaded area covers the

components located
inside the valve door
On-linefilter
Mixer Column
Selection
Valve
Injection (V2)
Pump Pump Valve (V1)
17 18
A B 2 6 2 7
34 5 3 6
4 5 COND UV
pH
Flow restrictor
G11
A2 B1 B2
W2 W1
Fraction
collector
Frac-901
Buffer G10
Valve (V6) Outlet
Valve (V4)
18 18 1 8
2 7 2 7 2 7
3 6 3 6 3 6
45 4 5 45
Column
A11 A12 A13 A14 A15 A16 A17 A18 Selection W3 F3
Valve (V3)
3 Connect port 2 of the outlet valve to the tube holder on the delivery
arm on Frac-901. Follow the instructions given in the Frac-900/901
User Manual.

_____________________________________________ ÄKTAexplorer 2
4 Extend the drain tubing from Frac-901 (34) to a suitable waste
bottle.
5 Modify the UniNet-1 data communication chain by:

34
1 2
36 35
• Connecting a UniNet-1 cable (35) between Pump P-900 and

Frac-901.
• Connecting a UniNet-1 cable (36) between Frac-901 and the

computer.
6 Connect a mains cable supplied (38) between Frac-901 and a mains

socket at the rear of ÄKTAH[SORUHU.
1 2
38
To Frac-901
To mains socket

2 ÄKTAexplorer ____________________________________________
Operation
Preparing for operation
1 Start your ÄKTAexplorer optional configuration system as
described in Preparation for use.
2 In UNICORN, go to the Instruction box in Method Editor and select

Frac.
3 Check out the instructions available for Frac-901 to familiarize

yourself with the new functions available. Refer also to the Fraction
collector Frac-900/901 User Manual to learn more about using
Frac-901.
Fractionation principles and definitions

Fraction collector Frac-901 is used for collecting fixed fractions during
elution and/or peak fractions.
The instruction Fractionation_900 starts eluate fractionation. The

instruction Peak_Fractionation_900 starts peak fractionation.
Fractionation and peak fractionation can have different fractionation

sizes. If both are activated simultaneously, the size set for peak
fractionation is valid for collecting fractions during peaks and the size
set for eluate fractionation is valid for collecting fractions during
elution.
Eluate fractionation is terminated by issuing the command

Fractionation_Stop_900 or by setting the fraction size to zero using the
parameter FracSize. This will not terminate peak fractionation if the
two are used simultaneously.
Peak fractionation is terminated by issuing the command

Peak_FracStop_900 or by setting the peak fraction size to zero using the
parameter PeakSize. This will not terminate eluate fractionation if the
two are used simultaneously, even if the command is issued during a
peak.
Tube change should occur (after the set delay volume) if a new
fractionation command is issued during eluate or peak fractionation.
The new eluate or peak fraction size is used instantly.
If a new peak fractionation command is issued during eluate

fractionation, peak fractionation is valid during peaks and eluate
fractionation before, between and after peaks.
If a new eluate fractionation command is issued during peak

fractionation before, between and after peaks (this means that the new
command becomes effective when the peak is ended).

_____________________________________________ ÄKTAexplorer 2
Using Frac-901 in a run
With Fraction collector Frac-901, fixed volume fractionation and/or
peak fractionation can be performed.
Note: When using Frac-901 in ÄKTAexplorer 10 systems, a maximum

flow rate of 3.5 ml/min is recommended when the 0.25 mm tubing kit
is used.
Select a method template with ”FP” in the middle of the template name.
and fill in the fraction parameters as indicated on the variables page.
• Flowthrough_FracSize
One fixed volume is collected before the eluate fractionation (fixed
volume fractionation)
• Eluate_Frac_Size
One fixed volume size during eluate between detected peaks, where
the elution size window is set by parameters
Start_Frac_at and End_Frac_at.
• Peak_Frac_Size
Peak fractionation volume within the peak size window set by
parameters Start_Peak_Frac_at and End_Peak_Frac_at.
• Peak_Start_Slope, Peak_End_Slope and Minimum_Peak_Width

Variables for controlling peak slope characteristics.

2 ÄKTAexplorer ____________________________________________
General
Frac-950 can be used for both small
scale and preparative scale
purifications with ÄKTAexplorer. Up
to 392 fractions can be collected in
time or volume base, as fixed volume
and/or peak fractionations.
The fractionation tube order can be

selected as serpentine-row, row-by-
row, serpentine-column or column-by- FRAC 950
column.
Four standard sizes of tube racks for

tube diameters of 12, 18 and 30 mm
tubes, and 96-well micro plates are
available.
Sample loss during tube change can be eliminated by using drop

synchronisation, or by using a built-in accumulator for intermediate
liquid storage between tube change.
Installation
in the Fraction Collector Frac-950 User Manual.
• If you are upgrading from Frac-901 to Frac-950,

go to step 2.
• If you are installing Frac-950 for the first time, start from step 6.
2 Disconnect the mains cable from Frac-901.
3 Disconnect the UniNet-1 cables from Frac-901.
4 Disconnect the capillary from the tube holder on Frac-901.
5 Remove Frac-901 from the workbench.
6 Put Frac-950 on the workbench to the right of ÄKTAexplorer.
Note: Frac-950 requires slightly more space than Frac-901.

_____________________________________________ ÄKTAexplorer 2

components located
On-linefilter
Mixer Column
Selection
Valve
Injection (V2)
17 18
A B 2 6 2 7
34 5 3 6
4 5 COND UV
pH
Flow restrictor
G11
A2 B1 B2
W2 W1
Fraction
collector
Frac-950
Buffer G10
Valve (V6) Outlet
Valve (V4)
18 18 1 8
2 7 2 7 2 7
3 6 3 6 3 6
45 4 5 45
Column
Valve (V3)

7 Locate the capillary kit included with Frac-950 and select if you
want to include the accumulator in Frac-950 or not. Connect
port 2 on the outlet valve in ÄKTAexplorer to Frac-950 as
described in the Frac-950 User Manual.
8 Connect the included waste tubing to the waste outlet on Frac-950

and route it to a suitable waste container.
9 Connect the UniNet-1 cables to Frac-950.
10 Connect the mains cable to Frac-950.
11 Check that all capillary and electrical connections are carried out
correctly.

2 ÄKTAexplorer ____________________________________________
Operation
2 In UNICORN, go to the System Control:Manual and select Frac.

yourself with the new functions available. Refer also to the
Fraction collector Frac-950 User Manual to learn more about using
Frac-950.

The instruction Fractionation starts eluate fractionation. The instruction

PeakFractionation starts peak fractionation.

sizes. If both are activated simultaneuously, the size set for peak
elution.

FractionationStop or by setting the fraction size to zero using the

Peak_FracStop or by setting the peak fraction size to zero using the
peak.



_____________________________________________ ÄKTAexplorer 2
If the parameter Start_at is set to Next line or Next group for peak
fractionation and eluate fractionation, only the first time a peak is
detected or during the first eluate the command is executed.
If the parameter Start_at is set to Next tube, the fractionation starts in

the tube position after the previous fractionation. This is also valid
when starting a new method, i.e. the last fractionation position in the
previous run is stored and used in the next run, even if using another
method. However, if the instruction Reset_position is ordered, or if the
parameter Start_at is set to First tube, the fractionation will take place
in the first position. The position is reset when changing racks as well.
If Last Tube is defined and the fraction collector run out of tubes, an
alarm is generated and the fraction collector goes to waste. If:
• Continue is ordered, the fraction collector remains in the waste

position.
• Ignore last tube and Continue are ordered, the fraction collector
continues in next tube.
• a new fractionation command and Continue is ordered, the

fraction collector continues with Start at tube number.

and fill in the fraction parameters as indicated on the variables page:

2 ÄKTAexplorer ____________________________________________
Fixed volume fractionation means that fixed volumes are collected

during the complete eluate:
• Flowthrough_FracSize starting with tube number

Start_Flowthrough_at
or that two fixed volume sizes can be collected, one before and one after
a set %B window:
• Flowthrough_FracSize starting in tube number

Start_Flowthrough_at, and Eluate_Frac_Size starting with tube
number Start_Eluate_Frac_at
where the %B window set by parameters:
• Start_Frac_at and End_Frac_at
Peak fractionation means that three volume sizes, two fixed and one
peak size dependent, can be controlled. One fixed volume size before the
eluate:

One fixed volume size during eluate between detected peaks:
• Eluate_Frac_Size starting with tube number

Start_Eluate_Frac_at
Finally, the peak fractionation volume:
• Peak_Frac_Size starting with tube number Start_Peak_Frac_at

within the peak size window set by Start_Peak_Frac_at and
End_Peak_Frac_at
The peak slope characteristics is controlled with the parameters:
• Peak_Start_Slope, Peak_End_Slope and Minimum_Peak_Width.
Note: When using Frac-950 in ÄKTAexplorer 10 systems, a maximum

is used.

_____________________________________________ ÄKTAexplorer 2
Continue with setting up the rack characteristiscs. Go to variable page
Frac-950.
Tube positions indicated by coloured spots

30 mm tubes
18 mm tubes
30 mm tubes
Select Rack type, Fractionation order, Last tube and mark the tube
positions in which you intend to put tubes by clicking on the
corresponding coloured spots in the figure.

2 ÄKTAexplorer ____________________________________________
2.7 Autosampler A-900
General
Autosampler A-900 is intended for automated multiple sample
injections. A-900 is only used in ÄKTAexplorer 10 systems.
A sequence of valve switching and syringe dispenser controlled sample

withdrawal enables the A-900 to inject sample volumes in a range from
a few microliters to 1 ml in a simple and reproducible manner.
Cover in open position
Buffer tubing
High pressure valve
Tubing guide
Needle arm
Tray segment
Syringe waste
Syringe dispenser
Wash solvent Drain wash

bottle position
Wash position Condensed water
and leakage drain
Three different injection methods can be selected:
• Flushed loop
The sample loop is completely (quantitatively) filled with sample
resulting in extremely good reproducibility (better than 0.3%).
• Partial loopfill
The sample loop is partially filled with sample giving low sample
loss and allowing programmable injection volumes.
• µl pick-up
After aspiration from the vial, the sample volume is transported
into the loop with transport liquid (mobile phase) from another
vial. This eliminates sample loss.

_____________________________________________ ÄKTAexplorer 2
The A-900 uses a syringe to aspirate the sample from a vial into the
sample loop. To prevent contamination of the syringe, the A-900 is
equipped with a buffer tubing between the syringe and the injection
valve.
An integrated washing mechanism removes the sample from the buffer

tubing, and sample needle, and also rinses both components.
Installation
Preparations
A–900 needs approximately 28 cm of bench space and a mains
connection of 220-240 V or 100-120 V, 50/60 Hz, preferably taken
from the ÄKTAexplorer base platform.
The recommended position is to the left of the separation equipment of

ÄKTAexplorer, with approximately 3 cm spacing.
Locking screw
explorer
ÄKTA
Red cap
A-900 has two transportation safety devices which must be removed:
• Remove the locking screw holding the front cover. It is located at

the right side of the front cover. Save the locking screw.
• A red plastic cap is attached over the needle washing mechanism

below the front cover. Remove the red plastic cap and save it.
See also the installation instructions given in the Autosampler A-900

8VHU0DQXDO&KDSWHU

2 ÄKTAexplorer ____________________________________________
Electrical connections
WARNING! A-900 must be connected to a grounded mains socket.
CAUTION! The mains power to ÄKTAexplorer must be

switched OFF before disconnecting or connecting the UniNet-1
cables.
WARNING! For continued protection against risk of fire, replace

only with fuses of specified type and rating. See Technical
specifications in the A-900 User Manual for fuse data.
1 Make sure that the ÄKTAexplorer chromatography system is

switched off.
2 Disconnect the UniNet-1 cable running between P-900 and the

computer, in the computer end.
Note: If fraction collector Frac-901 or Frac-950 is used, disconnect the

8QL1HWFDEOHUXQQLQJEHWZHHQ)UDF)UDFDQGWKHFRPSXWHU
Plug
Fr a c t i o n
collector
(optional)
Disconnect at the
computer end, and
route to the A-900
UniNet 1 connector To UniNet 2
Computer Chain
UniNet 1
Chain

_____________________________________________ ÄKTAexplorer 2
3 Connect this UniNet-1 cable to any of the two UniNet-1 connectors
in A-900.
Ter m i nat i on
P l ug
Frac t i on !
c ol l ec t or
115 V
1
0
To c om put er
Uni Net 1
Chai n
4 Connect a new UniNet-1 cable (supplied with the A-900) from the
other UniNet-1 connector on the A-900 to the computer. The
UniNet-1 connection to the computer must be made to the board
with four green LEDs.
Four green LEDs
UniNet-1 connection
5 Check that the voltage setting of the A-900 matches the local mains
supply voltage and main fuses. If the indicated voltage is not
correct, select the proper voltage by removing, inverting, and then
re-inserting the voltage selector cartridge. Check that the correct
fuses are installed. If not, replace them with fuses as stated below:
• For 110-120 VAC, use two 5 AT-fuses (slow).
• For 220-240 VAC, use two 2.5 AT-fuses (slow).
CAUTION! DO NOT switch on any of the equipment yet!
6 When the voltage selection and fuses are correct for the local mains
power source, plug in the supplied power cord in a free outlet in the
base platform of the ÄKTAexplorer separation unit.

2 ÄKTAexplorer ____________________________________________
Capillary connections
The capillaries to be connected between A-900 and the ÄKTAexplorer
separation unit are included in ÄKTAexplorer 10 XT Tubing kit,
code no. 18-1122-92. The flow diagram below shows the location of A-
900 in the ÄKTAexplorer flow path.
Autosampler
Injection
21 valve
3 6
4 5
Sample
loop
Shaded area covers the G-A1 G-A2

components located (H-A2)
On-line filter
Mixer
Injection
Pump Pump v alve (V1)
17 18
A B 2 6 2 7
34 5 3 6
4 5 COND UV
pH
Flow restrictor
Column
selection
v alve (V2)
A2 B1 B2
W2 W1
Fraction
collector
(optional)
Buffer Column
valve (V6) selection Outlet
v alve (V3) valve (V4)
18 18 1 8
2 7 2 7 2 7
3 6 3 6 3 6
45 4 5 45
A11 A12 A13 A14 A15 A16 A17 A18 W3 F3
1 Remove the capillary marked G5, connecting the Injection valve V1

port 1 with the Column selection valve V2 centre port.
2 Select the capillaries matching the tubing kit to be used in

ÄKTAexplorer after the column. Use the capillaries marked G-A1
and G-A2 if you intend to use 0.25 mm tubings kit, and the
capillaries marked G-A1 and H-A2 if you intend to use 0.50 mm
tubings.
Note: See section 2.1 in the ÄKTAexplorer System Manual for gen-
eral recommendations on selecting column and tubing kit.
3 Both capillaries are routed in the space between the UV monitor

UV-900 and the pH/conductivity monitor pH/C-900. The
capillaries are supported in the mounting bracket of Buffer valve V6
and are thread through the holes in the valve door below the valves
they connect to. Start threading the capillaries from the valves,
through the holes in the valve door, and further via the support and
the space between the monitors to A-900.

_____________________________________________ ÄKTAexplorer 2
4 Connect the capillary marked G-A1 (l = 1.3 m) from the tubing kit
between the Injection valve V1 port 1 and the A-900 Injection valve
port 1 (marked “pump”). The end that is pre-bent is connected to
the ÄKTAexplorer Injection valve V1.
5 Connect the capillary marked G-A2/H-A2 (l = 1.15 m) from the

tubing kit between the A-900 Injection valve port 6 (marked
“column”), and the Column selection valve V2 centre port.
Note: This connection (the dotted line) is only temporary. When

the installation test is successfully performed, G-A2 (H-A2) is con-
nected to the top inlet of the column, and the capillary connecting
the column outlet and the UV flow cell inlet is refitted.
6 Check additional internal and waste tubing connections on A-900

according to the A-900 User Manual instructions.
Preparing for installation test

2 Press the Control System button, and select

System:Settings:Special.
3 Check that the default sample loop volume (100 µl), syringe
volume (250 µl) and needle height (2 mm) are selected.
4 Manually, run the SyringeEnd instruction under System

Control:Manual:Flowpath:AutosamplerControl.
5 Remove the syringe from the syringe valve. Make sure that the Luer
connector in the valve remains in place.
6 Fill the syringe with 20% isopropanol in water from a vessel.
Note: Methanol can be used instead of isopropanol.
7 Replace the filled syringe. Fully tighten the syringe so that a gap
between the plunger tip and the syringe end is obtained after having
executed the SyringeHome instruction under
System Control:Manual:Flowpath:AutosamplerControl.
Note: If the syringe holder is not fully tightened, i.e. no gap is ob-
tained, the plunger tip moves forcefully into the syringe end which
FDQGDPDJHWKHV\ULQJH.
8 Fill the wash solvent bottle with 20% isopropanol in water. If the
sample contains large amounts of salt, or if the sample is very
sensitive to isopropanol, use water as wash solvent instead (degas
and change the water every day!).
9 Screw the bottle into the wash solvent holder and place the holder
on the A-900.

2 ÄKTAexplorer ____________________________________________
10 Put the wash solvent tubing in the wash solvent bottle.
11 Fill the tubing using the SyringeEnd and SyringeHome instructions.
SyringeEnd draws a syringe volume of wash solvent from the wash

solvent bottle and fills the wash solvent tubing.
SyringeHome dispenses the syringe contents to the syringe waste.
Repeat this action a number of times until the wash solvent tubing
and the syringe are completely filled.
12 After the wash solvent tubing and the syringe are filled, use the
InitialWash instruction to perform a standard wash. All tubing
connected to the syringe valve will be filled and flushed with wash
solvent.
Purging the syringe in A-900

1 Manually, run the SyringeEnd and SyringeHome instructions
alternately.
2 Repeat until no more air is left in the syringe (more than 5 repeats
can be necessary).
3 Finish by performing the InitialWash instruction.
Checking the needle height in A-900

1 Remove the tray segment next to (on the left side) the one directly
below the needle arm.
2 Put a vial with a flat bottom, filled with e.g. water, in the tray
segment directly below the needle arm. The bottom of the vial must
be clearly visible.
3 Perform a manual injection by issuing the

System Control:Manual:Flowpath:InjectionFlushed instruction.
4 Check visually that the needle stops approximately 2 mm above the

vial bottom.
5 If not, go to System:Settings:Special to adjust the needle height.

The default setting is 2 mm.
Note: Check the needle height with 2 mm as default setting whenever

the needle is replaced. The 2 mm setting is only a relative measure to
ensure that the needle stops above the vial bottom regardless of which
type of vial used. Adjust the final setting in small steps until a needle
height suitable for the actual use is obtained (type of vial, sample volume
available, etc.).

_____________________________________________ ÄKTAexplorer 2
Installation test
General
The installation test performs a total check of the liquid processing
parts of the ÄKTAexplorer separation unit. This is to establish that the
integration of the optional components in ÄKTAexplorer has not
disturbed the total system function.
Preparation
1 Verify that the Pump P-900 is primed and purged according to the
instructions in the ÄKTAexplorer Installation Guide.
2 Prepare a flask containing 500 ml of 0.4% acetone in water

solution.
3 Fill two standard vials with this solution and seal them properly
with the correct type of cap or septum.
4 Put one vial in position 1, and the other vial in position 2 in a

standard tray.
5 Immerse the inlet tubing B1 into the rest of the 0.4% acetone
solution.
6 Immerse the inlet tubing A11 into a flask containing distilled water.
7 Click on the Main menu button in the Taskbar.

Main menu button
8 Select File:Printer setup... from the menu bar. Select the appropriate
printer from the list and select Landscape. Click OK to
acknowledge the printer chosen.
9 Return to the Main menu screen, and click on the Instant Run
button .
10 Select Installation test in the templates list and make sure that Any
is selected for both Technique: and For column:.
11 Click on Run.
12 When the Method window is shown, click on Next, and then Next
again.
13 Click on START to begin the installation test.

2 ÄKTAexplorer ____________________________________________
14 The curves can now be monitored on the screen as the test

progresses. The Installation test method run time is approximately
30 minutes.
100%
95%
Ny bild från Lotten 70%

Autosampler
peaks 1 and 2
30%
Gradient 5%
result Step response result 0%
15 When the test run is finished, the printer automatically prints out
the chromatogram and the test result.
Evaluating the installation test result

Automatic evaluation
If your chromatography system includes no other optional components

than the Autosampler, the automatic evaluation will give a reliable
result. The system automatically prints out the test result when the test
is finished. The print-out consists of a chromatogram and an evaluation
of the test result.
• If the gradient test result is OK, the print-out says

“Gradient linearity accepted”.
• If the step response test result is OK, the print-out says
“Step response accepted”.
• If the UV response test result is OK, the print-out says
“UV response accepted”.
• If the Autosampler peaks test result is OK, the print-out says
“Autosampler peaks accepted”.
If any of the evaluated values falls outside the specified range, refer to
Correcting faulty evaluation results.

_____________________________________________ ÄKTAexplorer 2
Manual evaluation
If your chromatography system includes other optional components

than the Autosampler, the automatic evaluation will not give a reliable
result. If so, a manual evaluation must be done.
2 Click on in the results panel and then double-click on

the Installation Test01 icon to open the result file.
3 Maximise the chromatogram window by clicking in the upper

right corner.
4 Click in the chromatogram window with the right mouse button,

and select Properties.
5 Click on the Curves tab and select the following curves:
• Installation Test01:1_UV1_265nm@01,SMTH
6 Click on Peak table. Check that Peak Table A is selected and select
“Percent of total peak area” in the Peak Table Column. Press OK.
7 Double-click on in the upper left corner of the chromatogram

window and read the absorbance (in mAU) for the steps
corresponding to instte01:1_UV1_265nm@01,SMTH by dragging
the vertical bar to the constant section of each plateau. Enter the
absorbance values (in mAU) in column 2 in the Step response table
of the Test record, leaving out the decimals.

2 ÄKTAexplorer ____________________________________________
8 Read the absorbance for the plateaus corresponding to 0% and

100% B for the curves:
and enter the values in column 2 in the UV response table of the Test
record.
9 Click on Print under File to print out the chromatogram.
Evaluating the gradient

1 Place a ruler along the gradient part of curve
Installation Test01:1_UV1_265nm@01,SMTH in the printed report.
2 The curve should be linear between 10% B and 90% B and void of
discontinuities.
3 Fill in the Test record.
Evaluating the step response

1 Calculate the relative adsorption plateau heights for curve
Installation Test01:1_UV1_265nm@01,SMTH in the following way:
Subtract the base line value (0% B) from each of the values in
column 2 in the Step response table of the Test record. Enter the
results in column 3.
Divide each value in column 3 by the base line corrected value

corresponding to 100% B. Multiply by 100 and enter the results in
column 4.
2 The values of column 4 should all fall within the intervals given in
column 5.
Evaluating the UV response

1 Calculate the UV response ratios in the following way:
Subtract the base line values (0% B) corresponding to each UV

curve from the values in column 2 of the UV response table of the
Test record. Enter the results in column 3.
Calculate the absorbance ratios 265 nm/254 nm and 265 nm/280

nm using the values from column 3 and enter the results in
column 4.
2 The ratios obtained should all fall within the intervals given in
column 5.

_____________________________________________ ÄKTAexplorer 2
Evaluating the Autosampler peaks
1 In the printed report, read the Area and the Area/Peak area %
values for peaks No. 1 and 2.
3 Check that the values fall within the given intervals.
The installation procedure is completed when all results fall inside the
limits specified in the Test record.
Correcting faulty evaluation results

• Faulty gradient
• The gradient is linear but the interval is to small. The mixer

chamber is too large, or the mixer is faulty.
• Disturbances. These may arise from air in the pump. Purge again
and repeat the test run. If persistent, faulty check valves or bad
piston sealings in the pump may be the cause. Refer to the
Pump P-900 User Manual.
• Faulty step response
• All values are faulty. This may arise from air in the pump or a
faulty pump. Purge again and repeat the test run. If persistent, a
faulty pump may be suspected. Refer to the Pump P-900 User
Manual.
• 5% and 95% faulty. This may be due to bad sealings in the pump
(5% faulty=pump module B, 95% faulty=pump module A).
Refer to the Pump P-900 User Manual.
• Faulty UV response
• 0%-value too far from 0 AU (±10 mAU). Auto zero not properly
performed. Repeat the test and re-evaluate. Refer to the
• Faulty Autosampler peak values
• If values are faulty, make sure that the A-900 is properly purged
and that all tubing fittings are properly tightened.

2 ÄKTAexplorer ____________________________________________
Test record
Gradient test result
Gradient linear from ....... % B to ....... % B. (10 – 90 %)
Step response test result

Step response table:
1 2 3 4 5
Programmed Value Base line Normalised Allowed
Conc. %B read corrected value interval
(mAU) value
100
95 94 - 96
70 69 - 71
30 29 - 31
5 4-6
0
UV response test result

UV response table:
1 2 3 4 5
Wavelength Value read Base line Absorbance Allowed
(nm) 100% B 0% B corrected ratio interval
value
254
265/254 1.11 - 1.26
265
265/280 1.26 - 1.53
280
Autosampler peak test result

Autosampler peak table:
1 2 3 4
Peak no. Area Area/Peak area Allowed
(mAU x min) (%) interval
(%)
1 66.1 - 67.2
2 32.8 - 33.9

_____________________________________________ ÄKTAexplorer 2
Operation
General
Autosampler A-900 is controlled from a PC running UNICORN
version 3.2 or higher. It cannot be used as a stand-alone module. A-900
can be controlled, either automatically from a pre-made method
template, or manually via commands.
To control A-900 automatically from a method template, select any of

the templates starting with Autosampler Injection... (refer to section
2.16 Strategy and template overview in ÄKTAexplorer).
The A-900 can be used for automatic sample injection from a few micro
litres to one millilitre of sample solution. To achieve optimal
performance, selection of sample loop volume, syringe size and speed,
plus the washing method between injections must be considered.
The sample injection techniques, which can be programmed in the

method editor and then utilised in automatic runs, are as follows:
• InjectionFlushed
resulting in extremely good reproducibility.
• InjectionPartial
• InjectionPickup
vial resulting in virtually no sample loss.
These sample injection techniques can also be ordered manually from

System Control:Manual:Flowpath.
Manual functions used for setting up and maintaining the A-900 are
found in System Control:Manual:Flowpath:AutosamplerControl.
The available commands are as follows:
• InitialWash
• RotateTray
• SyringeHome/End
• NeedleHome/Front
All of these functions are described in the following sections. A table

showing all the hardware related parameters for A-900 is given below.

2 ÄKTAexplorer ____________________________________________
These parameters are found under

System:Settings:Special:AutosamplerParameters
Parameter Value Comment

LoopVol 10-1000 µl Sample loop volume
(100 µl is default)
SyringeVol 100, 250, 500 or 1000 µl Syringe volume
FlushVol 30-100 µl Flush volume. The amount of sam-
(40 µl is default) ple, or transport liquid, taken from a
vial before the loop is filled with
sample.
WashVol 300-9999 µl Wash volume. Sets the volume for

(300 µl is default) the external wash (WashExt) pro-
grammable in the different injection
modes.
TrayType StandardVial_A A=Standard vial O.D. 12 mm.

MicroVial_D D=Micro vial O.D. 7 mm
HeadPress No, Yes Head-space pressure. Applied via
the outer air needle. Assures that
no air or vapour bubbles are formed
during sample aspiration.
NeedleHgt 0-40 mm Needle height. A relative measure
(2 mm is default) used toadjust the space between
the needle and the bottom of the
vial used.
AutosamplerSy- Low Syringe speed setting.
ringeSpeed Normal
High
(Normal is default)
Scaling 0.1-1.0 Syringe speed scaling factor.
(1.0 is default)
InjectionFlushed
The InjectionFlushed instruction selects the flushed loop injection mode
for A-900.
The syringe dispenser aspirates a flush volume of sample into the sample
line. The A-900 injection valve is turned to LOAD position, and the
sample loop is filled with 1.5–3 times the sample loop volume
(depending in the loop volume). The injection volume equals the loop
volume. When the sample loop size is changed, the loop volume setting
must be changed in System:Settings (100 µl is the default loop volume).

_____________________________________________ ÄKTAexplorer 2
After each injection, the needle is rinsed:
• If WashExt=NO is selected in the method, the needle is only washed

internally.
• If WashExt=YES is selected in the method, the needle is also washed

on the outside.
In both cases, the buffer tubing is also washed.
An air segment (Air segment=Air) is recommended to reduce the

amount of the flush volume. The air segment is not injected.
More details of flushed loop injection are found in the Autosampler

A-900 User Manual, Reference information A.
InjectionPartial
The InjectionPartial instruction selects the partial loopfill injection
mode for A-900.
The syringe dispenser aspirates a flush volume of sample into the

sample line. The A-900 injection valve is turned to LOAD position, and
the sample loop is filled by transporting the selected injection volume
into the sample loop, which is partially filled with sample.

internally.

on the outside.

More details of partial loopfill are found in the Autosampler A-900

User Manual, Reference information A.
InjectionPick-up
The InjectionPickup instruction selects the µl pick-up injection mode
for A-900.
The syringe dispenser aspirates transport liquid from the transport vial
into the sample line. The A-900 injection valve is turned to LOAD
position. The needle moves from the transport vial to a sample vial and
aspirates the selected injection volume. The needle moves back to the
transport vial and the sample is quantitatively transported into the
sample loop with transport liquid.

2 ÄKTAexplorer ____________________________________________

internally.

on the outside.
In both cases, the buffering tubing is also washed.
An air segment is not recommended for this injection mode

(Air segment=NoAir) because the air segment at the front of the
sample plug is injected.
More details of µl pick-up are found in the Autosampler A-900 User

Manual, Reference information A.
InitialWash
The InitialWash instruction performs a complete wash of the needle and
the syringe.
The syringe is first washed using one syringe volume.
Then the needle is moved to its home position and washed internally.
Finally, the needle moves to its front position and is washed both
internally and externally.
RotateTray
The RotateTray instruction is used to rotate the tray, e.g. when new vials
are loaded into the tray segments.
SyringeHome/End
The SyringeHome/End instructions are used when the syringe is
replaced. They are also used for purging the syringe, i.e. to remove air.
SyringeHome empties the syringe contents to the syringe waste

container.
SyringeEnd aspirates wash solvent into the syringe from the wash
bottle.
Note: When the performed action is completed, make sure the syringe
is run to the Home position before leaving the manual function.

_____________________________________________ ÄKTAexplorer 2
NeedleHome/Front
The NeedleHome/Front instructions are used when the sample needle
is replaced.
NeedleHome moves the needle to its normal position.
NeedleFront moves the needle closer to the front of A-900.
AutosamplerSyringeSpeed
The AutosamplerSyringeSpeed parameters are used to manually preset
the speed of the syringe. The syringe speed needs adjusting when
changing syringe size, using highly viscous samples, or using volatile
eluents.
Three different speed settings are available:
• Low
• Normal (default setting)
• High
The actual speed is dependent on the speed setting and a scaling factor,
and on the syringe volume. The scaling factor range of 0.1 - 1.0 is used
to reduce the basic speed setting.
Syringe volume Viscosity Speed setting Scaling factor

250 µl Normal Normal 1.0
250 µl High (5 - 10 cP) Low 0.2
500 µl Normal Low 1.0
Note: If the sample is viscous, the flush volume can be enlarged and the
syringe speed reduced to enhance performance.

2 ÄKTAexplorer ____________________________________________
2.8 Flow direction valve
Installation
components located G5
Flow
On-linefilter Direction
17 Valve (V7)
G6 2 6
34 5 G10
Mixer Column G9
Selection
Valve
Injection (V2)
17 18
A B 2 6 2 7
34 5 3 6
4 5 pH COND UV
G8
Restrictor
Fraction
collector
A2 B1 B2 (optional)
W2 W1
Buffer Outlet
Valve (V6) Valve (V4)
18 18 1 8
2 7 2 7 2 7
3 6 3 6 3 6
45 4 5 45
Column
A11 A12 A13 A14 A15 A16 A17 A18 Selection W3 F3 F4 F5 F6 F7 F8
Valve (V3)

connecting the UniNet-2 cables.
1 Mark the INV-907 valve with V7. Check that the ID code switch is
set to 7.
2 Attach the valve to the valve door in the recess next to Column
Valve V2. Use the mounting bracket and screws supplied with the
valve.
3 Cut and mount the required PEEK capillaries. Use a capillary with
the correct i.d. as required by your system. The principle flow path
through the valve is shown below:
Reversed flow configuration
DOWNWARD FLOW, position 1 UPWARD FLOW, position 2
1 1
2 7 Pump 7 Pump
2
To monitor To monitor 3
3 6 6
4 5 4 5
Stop plug Stop plug

_____________________________________________ ÄKTAexplorer 2
4 Use the attached UniNet-2 cables to connect the valve to the
UniNet-2 communication link. The valve can be connected
anywhere between the system pump P-900 and the termination
plug.
correctly.
Operation
2 The flow direction valve is automatically recognised by UNICORN

at system start-up. Check that the valve added is indicated in the
Flow scheme in UNICORN.
3 Verify that Flow Direction Valve V7 is functioning properly by

issuing manual commands from UNICORN as follows:
• In System Control, select Manual:FlowPath.
• Select instruction FlowDirection.
• Set FlowDirection to position DownFlow.
• Manually, run Pump P-900 with distilled water at a flow rate of

approximately 5 ml/min and check that water is flowing down-
wards the bypass capillary connected between ports 1 of the
column selection valves V2 and V3.
• Set FlowDirection to position UpFlow.

approximately 5 ml/min and check that water is flowing up-
wards the bypass capillary connected between ports 1 of the
column selection valves V2 and V3.
Using the flow direction function in a run

The flow direction valve instruction is designated FlowDirection. To use
the valve in a method, do as follows:
1 In the Main Menu, select File:New:Method to create a new method.

Select System:, Technique:, Template: and For column:. Click OK.

2 ÄKTAexplorer ____________________________________________
2 Select View:Text instructions to display the text instruction editor.

Double-click on the instruction block where you want to add the
flow direction valve instruction, e.g. Start_Conditions. Highlight the
instruction below which you want to insert the valve instruction.
Click here to define a

variable name for the
instruction parameter
3 Click the Flowpath radio button.
4 From the Instructions list, select FlowDirection. The valve

instruction parameter Position allows you to change position for the
flow direction valve using parameters DownFlow and UpFlow.
5 Define a variable name for the instruction parameter, e.g.

FlowDirection. This variable allows you to turn the position of the
valve to the direction you want.
6 Click OK. The defined variable is inserted in the method.
7 Return to the Variables page by selecting View:Run setup to set the

variable FlowDirection to the desired position.
8 Save the method.

_____________________________________________ ÄKTAexplorer 2
2.9 General function valve
Installation
To install any optional valve, proceed as follows:

1 Mark the optional function valve with V8 or V9 as required. Check

that the ID code switch is set to 8 or 9 respectively.
2 Mount the bracket and attach the valve to a rail on the system wall.
Choose the best location to facilitate the routing of the tubing that
connects the valve to the main system flowpath.
3 Cut and mount the necessary capillaries. Restrict the length of the
capillaries as much as practically possible to reduce band
broadening effects.
4 Use the attached UniNet-2 cable to connect the valve to the

anywhere between Pump P-900 and the termination plug.
correctly.
Operation
2 A connected valve is automatically recognised by UNICORN at

system start-up. Check that the added valve is indicated in the
flowscheme in UNICORN.
3 Verify that the valve is functioning properly by issuing manual

commands from UNICORN as follows:
• In System Control, select /Manual:FlowPath.
• Select instruction Valve X, where X=ID code for the valve.
• Set the valve to position 3 (Waste for INV-907).

approximately 5 ml/min and check that water is coming out
from port 3 on the valve (port 5 for INV-907).

2 ÄKTAexplorer ____________________________________________
Using the general function valve in a run

General function valve instructions are designated Valve8 and Valve9 in
the flow scheme and run data windows. To use a valve in a method, do
as follows:


general function valve instruction, e.g. Start_Conditions. Highlight
the instruction below which you want to insert the valve instruction.


_____________________________________________ ÄKTAexplorer 2
3 Click the Flowpath radio button
4 From the Instructions list, select the valve you intend to use (Valve8
or Valve9). The valve instruction parameters allow you to change
the position of the valves. For INV-907, only three positions can be
selected.

Valvex_pos, where x=8 or 9 depending on the ID code for the valve
used. This variable allows you to turn the position of the valve to
the port you want.

variable Valvex_pos to the desired position.
8 Save the method.

2 ÄKTAexplorer ____________________________________________
2.10 pH flow cell and pH electrode mounting
General
Using a pH electrode is optional in ÄKTAexplorer 10, 10 S and 10 XT,
and therefore, it is not mounted from the factory. The pH electrode is to
be mounted in a separate pH flow cell, housing the pH electrode itself,
as well as a dummy pH electrode. The dummy pH electrode replaces the
pH electrode whenever the pH electrode is to be removed from the
system, e.g. for storage.
The pH electrode contains a sealed Ag/AgCl reference which cannot be

refilled, an internal electrolyte bridge of 4 M KCl saturated with Ag/
AgCl, an outer electrolyte bridge of 1 M KNO3, an annular ceramic
reference junction and a low profile pH membrane. This is commonly
known as a sealed combination double junction type. The pH electrode
is delivered with a transparent cover.
Note: The flow cell should not be used with any other pH electrode.
Note: Always use the 0.50 tubing kit when the flow cell is mounted.
Note: Using the flow cell is not recommended when the peak volume is
less than 1 ml. The volume in the flow cell decreases the UHVROXWLRQRIWKH
SHDNV
Installation
WARNING! Never use 0.25 mm i.d. tubing (blue) when the pH flow
cell is mounted in the system. The electrode may rupture due to
high backpressure.
1 Install the pH flow cell holder (packed in Box-900) in the groove

directly below the Outlet Valve (V4) on the outside of the valve
door.
2 Push the slide clamp upwards to secure the pH flow cell holder.
3 If 0.25 mm i.d. tubing is installed, remove the capillaries G5 and

G7–G11. Connect Tubing kit 0.50 as described from step 4.
If 0.50 mm i.d. tubing is already installed, remove capillary G9 only.

Connect the new tubing according to steps 7 and 8.
4 Connect the centre port of the injection valve V1 to the centre port
of the column selection valve V2 using capillary H5.
5 Connect the centre port of the column selection valve V3 to the inlet
(on top) of the UV flow cell using capillary H7.

_____________________________________________ ÄKTAexplorer 2
6 Connect the outlet (at the bottom side) of the UV flow cell to the
inlet of the conductivity flow cell (opposite to the screw head end)
using capillary H8.

components located
On-linefilter
Mixer Column
Selection
Valve
(V2) H12 H9
Injection
17 18
A B 2 6 2 7
34 5 3 6
4 5 pH COND UV
Flow restrictor
A2 B1 B2 W2 W1
Fraction
collector
Buffer (optional)
Valve (V6) Outlet
Valve (V4)
18 18 1 8
2 7 2 7 2 7
3 6 3 6 3 6
45 4 5 45
Column
Valve (V3)
7 Connect the outlet of the conductivity flow cell (the screw head
end) to the inlet of the pH flow cell using capillary H9.
8 Connect the outlet of the pH flow cell to the FR-904 flow restrictor
(marked IN) using the H12 capillary.
9 Connect the outlet of the FR-904 flow restrictor to the centre port
of the V4 outlet valve using the H10 capillary.
10 Place the FR-904 flow restrictor (1) in the hole in the pH flow cell
holder.
1 11 Mount the pH electrode (2) for experiments where pH is to be

measured. Otherwise, mount the pH electrode dummy in the hole
2
in the pH flow cell.
Note: When using the pH electrode, the flow restrictor FR-904 must be
replaced with the supplied flow restrictor FR-902. Otherwise, the long
term stability and lifetime of the pH electrode will deteriorate.
Note: For calibration of the pH electrode, refer to the UNICORN

User Manual.

2 ÄKTAexplorer ____________________________________________
Operation
1 Start your ÄKTA optional configuration system as described in
Preparation for use.
2 If fraction collection should be used, remember to change the delay

volume. This setting is found in the System Control menu;
System:Settings:Special. See also 2.15 Configuration dependent
delay volume in ÄKTAexplorer.
3 Using pH measurement in a run is described in the ÄKTAexplorer

System Manual. Also refer to the Monitor UV-900 User Manual.

_____________________________________________ ÄKTAexplorer 2
2.11 Sample pump P-950 and Sample valve V5
General
Pump P–950 is a membrane type
laboratory pump for use as a
sample pump to apply samples P-950
directly on the column and/or to fill

sample loops, including Superloop.
Pumping action is provided by a
membrane driven by a stepper
motor. The pump is a low pulsation
pump and features a motor speed
independent of temperature and
load to give accurate and
reproducible flow rates up to
50 ml/min.
A Sample valve PV–908, with a pressure limit of

25 MPa, can be used in combination with the PV-908
Sample pump. It is a motorised 8-way valve used

for automatic sample application of up to 8
samples.
Pump P–950 and Sample valve PV–908 are

optional components in ÄKTAexplorer 10 and
ÄKTAexplorer 10 XT.
ÄKTA E10 S Upgrade kit, code no. 18-1147-02, contains all items that
are required for upgrading an ÄKTAexplorer 10 system with
Pump P–950 and Sample Valve V5.
The following items are included in ÄKTA E10 S Upgrade kit:
• Pump P–950
• Mounting kit P–950
• Sample Valve PV–908
• Sample holder parts
• UniNet cables
• ÄKTA E10 S Tubing kit
• Tefzel tubing, 4 m
• Finger-tight connectors

2 ÄKTAexplorer ____________________________________________
There are three ways of using Pump P–950 in the system:
• Loading a sample loop.
• Direct loading onto the column.
• Loading a Superloop.
Each way requires a specific installation configuration and different

procedures to remove air from the flow path inside the sample pump, as
described in the following.
Control of the sample pump can be achieved automatically from a

method, or manually via the functions available in UNICORN.
There are ready-made method templates available for automated

sample application in ÄKTAexplorer, refer to 2.16 Strategy and
template overview in ÄKTAexplorer.

_____________________________________________ ÄKTAexplorer 2
Installing the components
CAUTION! The mains power to ÄKTAexplorer must be switched

OFF before disconnecting or connecting the UniNet-2 cables.
This section describes how to install the components in the system. To

connect the capillaries, see the section describing that specific sample
application method—Loading a sample loop, Direct loading onto the
column or Loading a Superloop.
Installing Pump P-950

1 Make sure that the ÄKTAexplorer chromatography system is
switched off.
2 Open the valve door. Injection

Valve, V1
3 Install Pump P-950 in the (INV-907)
separation unit of
ÄKTAexplorer.
• Locate the fixing screws and

the clamp supplied with the 0, 6 P-950 Sample
pump. M-925
Valve, V5
(PV-908)
• Fit the pump as described in

the Mounting kit for
Pump P-950, Instructions Sample Pump P-950
supplied.
Installing the Sample valve

1 Install the Sample valve V5 (PV-908) above Pump P-950 (also refer
to IV/PV-908 Instructions).
• Set the rotary ID-switch at the bottom of

the V5 valve to position 5.
• Remove the black, circular plastic cap at-

tached in the mounting hole.
• Use the attached bracket and screws to fit

the valve in the same way as the V1 valve.
• Apply an identification number (5) from

the labels supplied. Place the number so
that it is easily readable from the front.
2 Close the valve door.

2 ÄKTAexplorer ____________________________________________
Installing Sample holder SH-900

1 Install the Sample Holder SH-900 on
the beige, plastic mounting plate below 1
the Sample valve V5
2 Install the Tubing holder (1), the Tube

holders (2) and the Sample tray (3) by
2
pushing them into the slits.
Installing UniNet-2 cables

3
1 From the Column selection valve V3
UniNet-2 connector, disconnect the
cable running between Injection valve V1 and Column selection
valve V3. If not installing a Sample valve, go directly to step 3.
2 Connect this cable between

Injection valve V1 and the
inner Sample valve V5
UniNet-2 connector. Loosen
the V5 attachment bracket if
it is difficult to plug in the
connector.
3 If not installing a Sample 0, 6

P-950
valve, connect this cable to a M-92

5
free Pump P-950 UniNet-2

connector.
From UniNet 2
connector at rear of
If the Sample valve is Pump P-901 T e r min a t io n p lu g
installed, connect a new

cable (supplied) between the outer Sample valve V5 UniNet-2
connector and the Sample pump P- 950 UniNet-2 connector.
4 Connect a new cable (supplied) between the other Sample pump

UniNet-2 connector and the free UniNet-2 connector in the Column
selection valve V3.

_____________________________________________ ÄKTAexplorer 2
This page is intentionally left blank.

2 ÄKTAexplorer ____________________________________________
Loading a sample loop

Automatic filling of sample loops can be useful when samples must be
applied repeatedly, for example, in scouting runs. By using sample loops
supplied by Amersham Pharmacia Biotech, volumes between
0.1–2.0 ml can be applied. The sample is drawn into the sample loop by
the sample pump.
Note: The flow rate should not exceed 0.5 ml/min when filling the
sample loop. Otherwise, cavitation may occur due to the resistance in
the flowpath.
Connecting the capillaries

1 Make sure that the Sample pump and the Sample valve (if included)
are properly installed according to section Installing the
components.
2 Connect all capillaries using connectors and ferrules as indicated in

the table and in the figures.
Use the Sample holder for storing sample 4

tubes and sample waste vessels (5), and for
routing the sample and waste capillaries (4).
5
3 Check that all capillary and electrical
connections have been carried out correctly.
Fingertight connector 1/16" Pump P-950
Outlet connection
Tubing connector for 1/16" o.d. tubing with
ferrule for 1/16" o.d. tubing
Union 5/16" female/ HPLC male

Inlet connection

_____________________________________________ ÄKTAexplorer 2
components located
On-linefilter
Mixer Column
Selection
Valve
Injection (V2)
Pump Pump Valve (V1) 3
17 18
A B 2 6 2 7
34 5 3 6
4 5 COND UV
Flow restrictor
4
Sample
A2 B1 B2
2 Pump Fraction
P-950 W2 W1
collector
(optional)
1
Buffer Sample Outlet
Valve (V6) Valve (V5) Valve (V4)
18 18 18 1 8
2 7 2 7 2 7 2 7
3
45
6 5 3
45
6 3
4 5
6 3
45
6
Column
A11 A12 A13 A14 A15 A16 A17 A18 S1 S2 S3 S4 S5 S6 S7 S8 Selection W3 F3
Valve (V3)
Capillary (max. length) Connector Connection points

1
PEEK i.d. 0.75 mm x 200 mm Fingertight connector 1/16", 18-1112-55 Injection valve, port 4
(Sample inlet tubing)
Fingertight connector 1/16", 18-1112-55* Sample selection valve, centre
port*
2
Tubing connector for 1/16" o.d. tubing, 18-1127-07 Sample pump, inlet connector
with ferrule for 1/16" o.d. tubing, 18-1127-06
3
Tefzel i.d. 0.75 mm x 1300 mm Fingertight connector 1/16", 18-1112-55 Injection valve, port 5
4
Teflon i.d. 1.6 mm x 1300 mm Union 5/16" female/ HPLC male, 18-1142-08. Sample pump, outlet
Tubing connector for 1/8" o.d. tubing, 18-1121-17 connector
with ferrule for 1/8" o.d. tubing, 18-1121-18.
5
Tefzel i.d. 0.75 mm x 320 mm* Fingertight connector 1/16", 18-1112-55 Sample selection valve,
port 1-8
*
Only if a sample selection valve is installed.
Note! Connect stop plugs to the non-used ports in the sample selection valve.

2 ÄKTAexplorer ____________________________________________

described in Preparation for use in section 2.4 General guidelines.
2 Check that the Sample pump and the Sample valve (if included) are
indicated in the flow scheme window.
3 Make sure the system is connected as previously described in section

Connecting the capillaries on page 74.
4 Verify that the sample pump is functioning properly by issuing

manual commands from UNICORN as follows:
• Fill a container with distilled water and submerge the sample

inlet capillary S1 (or S4 if the Sample valve is included) in the
container.
• Set the injection valve to position LOAD.
• Select System Control:Manual:Flowpath.
• If included, select the instruction SampleValve in the list and

select S4.
• Click on Execute.
• Select System Control:Manual:Pump.
• Select the instruction SampleFlow in the instruction list.
• Set Flowrate to 5 ml/min and click Execute. Allow the pump to

run for 3 minutes to prime the pump.
• Check that the sample pump is delivering flow through its outlet.
• Stop the pump by clicking End in the System Control toolbar.
Priming the sample inlet tubing

If no Sample valve is used, prime the tubing as follows:
1 Make sure that the system is prepared according to section

Preparing for operation.
2 Place the sample inlet tubing into the sample container.
3 Set the injection valve to position LOAD.

_____________________________________________ ÄKTAexplorer 2
4 Start the sample pump manually with a sample flow of 0.5 ml/min
and let the pump run to completely fill the inlet tubing to the
injection valve with sample.
Note: When filling the sample loop, the resistance in the flowpath
may cause cavitation if the flow rate exceeds 0.5 ml/min.
5 Stop the sample pump.
If using a Sample valve, prime the tubing in the following way:
1 Place the inlet tubing from port 8 of the Sample valve into a small
bottle of starting buffer, buffer A. Use this solution to rinse the
tubing between the Sample valve and the Injection valve to
minimize carry-over when changing from one sample to another.
2 Place the inlet tubing from the Sample valve ports 1–5 into test
tubes containing the samples.
3 Set the Injection valve to position LOAD (pos. 1).
4 Set the Sample valve in the position corresponding to the first

sample, position S1 (sample tube one).
5 Start the Sample pump manually from UNICORN. Set the flow
rate to 2 ml/min and let the pump run for approximately 10 s
(=0.35 ml) to completely fill the inlet tubing to the Sample valve
with sample.
6 Stop the Sample pump.
7 Set the Sample valve in the position for the next sample.
8 Repeat steps 6–9 until all inlet tubings to the Sample valve has been
filled with sample.
9 Set the Sample valve manually from UNICORN in position S8.

Start the Sample pump from UNICORN. Set the flow rate to
2 ml/min and let the pump run for approximately 1 minute (=2 ml)
to completely fill the tubing from the test tube to the Injection valve
with starting buffer.
11 Set the Injection valve to position INJECT, pos. 2.
12 Flush the sample loop by starting the Pump P-900. Let a volume of
5 times the volume of the sample loop pass through.
13 Stop the Pump P-900 and set the Injection valve to LOAD (pos. 1)
by clicking on End. The system is now ready to start a run.

2 ÄKTAexplorer ____________________________________________
Calibrating the sample pump

If the running conditions are changed, calibrate the sample pump after
priming the tubing. The calibration instruction is found in the Pump
P-950 User Manual.
Note: The injection valve must be in position LOAD when calibrating

the sample pump.
Preparing a method manually for filling a sample loop

If your optional configuration does not include a Sample valve, a
method must be prepared manually, as described in the following. If a
Sample valve is included, go directly to section Filling the sample loop.
The sample pump instructions are designated SampleFlow and

SamplePres in the flow scheme and run data windows. An example of
how to use the sample pump for automatic repetitive sample loading is
shown below.


Create a new block directly above the Sample injection block to
include the instructions necessary to control the sample pump. A
new block is created in the following way:
3 Select Block:New.
New block containing instructions

for controlling the sample pump.
Note that MethodBase is changed
* to SamplePump and then back to
*
* SystemPump, to achieve correct
* volume calculation.
*
*
Instructions marked with * must be
added manually.
4 Type a block name (e.g. Load_Sample_Loop) and click OK.
5 The new block is inserted at the end of the method. Click on the new
block with the left mouse button, drag it up and drop it on the
Sample_injection block. Double-click (left mouse button) on the
blue square to open the new block. Insert instructions according to
the figure below. The instructions marked with * must be added
manually.

_____________________________________________ ÄKTAexplorer 2
6 Select Base: Volume and Method base: SamplePump.
7 Insert the InjectionValve instruction and set the Position parameter

to LOAD.
8 Insert the SampleFlow instruction and select 0.5 ml/min.
9 Insert Method base: SystemPump.
10 Insert the SampleFlow instruction and select 0 ml/min to stop the

sample pump.
11 A breakpoint variable is created by clicking on Var in the

Breakpoint dialog window.
Click here to create a

breakpoint variable
Breakpoint variable.
Sets the sample volume to
be loaded into the sample
loop.
Note: For complete filling, an overfill of 2-5 times the loop volume
is needed for maximum reproducibility between the runs.
12 Save the method.

2 ÄKTAexplorer ____________________________________________
Filling the sample loop

If your optional configuration includes a Sample valve, a pre-made
method template is available for filling the sample loop. Alternatively,
use your manually made method.
Prepare the system as described previously and proceed as described

below.
1 Use any pre-made method template that begins with Sample Pump
Loop Injection.... Refer to 2.16 Strategy and template overview in
ÄKTAexplorer. In the method, enter 5 times the volume of the
sample loop for the variable Rinse_loop_with. This will rinse the
tubing between the Sample valve and the Injection valve as well as
the sample loop before the sample is loaded into the sample loop.
Note: When the same sample is used for repeated application, the
sample loop and the tubing between the Sample valve and the Injec-
tion valve do not need to be rinsed with buffer between runs. Enter
zero for the variable Rinse_loop_with.
2 Enter the sample volume in the variable Fill_loop_with. To

compensate for the volume between the Sample valve and the
Injection valve, 85 µl should be added to the sample volume entered
85 l in the variable in UNICORN. For complete filling, an overfill of 2-
5 times the loop volume is needed for maximum reproducibility
between the runs.
Note: When only one sample is used, it is only necessary to compen-

sate for the volume between the Sample valve and the Injection
valve the first time the sample is applied.
3 Selection of the different samples is specified in the Sample_inlet

variable. The samples will be applied automatically. In scouting
runs, you can enter different sample volumes with the variable
Fill_loop_with.
Emptying the sample loop

When emptying the sample loop, use a buffer volume of approximately
5 times the sample loop volume to flush the loop and ensure that all
sample is injected onto the column. In template methods, use the
variable Empty_loop_with to set the volume.

_____________________________________________ ÄKTAexplorer 2

2 ÄKTAexplorer ____________________________________________
Direct loading onto the column

Loading sample directly onto the column is useful when large amounts
of sample is to be loaded. The sample is pushed onto the column.

components.

Use the Sample holder for storing

4
sample tubes and sample waste vessels
(5), and for routing the sample and
waste capillaries (4).
5

connections have been carried out
correctly.

components located
On-linefilter
Mixer Column
Selection
Valve
2 Injection (V2)
17 18
A B 2 6
3 2 7
34 5 3 6
4 5 COND UV
4
Restrictor
Sample
A2 B1 B2 Pump Fraction
P-950 W2 W1
collector
(optional)
5
1
18 18 18 1 8
2 7 2 7 2 7 2 7
3 6 3 6 3 6
45 6 45
3
4 5
6 45
Column
A11 A12 A13 A14 A15 A16 A17 A18 S1 S2 S3 S4 S5 S6 S7 S8 Selection W3 F3
Valve (V3)

_____________________________________________ ÄKTAexplorer 2
Pump P-950
Fingertight connector 1/16"
Outlet connection

Inlet connection

1
Teflon i.d. 1.6 mm x 500 mm Union 5/16" female/ HPLC male, 18-1142-08. Sample selection valve, centre
(Sample inlet tubing) Tubing connector for port*
o.d. tubing, 18-1121-17
with ferrule for 1/8" o.d. tubing, 18-1121-18*.
2
Fingertight connector 1/16", 18-1112-55 Sample pump, outlet
connector
3
Fingertight connector 1/16", 18-1112-55 Injection valve, port 6
4
5
Teflon i.d. 1.6 mm x 1300 mm Union 5/16" female/ HPLC male, 18-1142-08. Injection valve, port 4
Tubing connector for 1/8" o.d. tubing, 18-1121-17
6
Teflon i.d. 1.6 mm x 1300 mm* Union 5/16" female/ HPLC male, 18-1142-08. Sample selection valve,
Tubing connector for 1/8" o.d. tubing, 18-1121-17 ports 1–8.
* Only if you have a sample selection valve in your configuration.

Note! Connect stop plugs to non-used ports in the sample selection valve.
Note! If low sample flows are used (≤ 10 ml/min), sample inlet tubings connected to ports 1–6 can be
replaced with PEEK i.d. 0.75 mm tubing. The sample inlet tubing should be adjusted to a suitable
length.

2 ÄKTAexplorer ____________________________________________




container.
• Set the injection valve to position WASTE.

select S4.

• Check that the sample pump is delivering flow through port 4

(W2) in the Injection valve.

_____________________________________________ ÄKTAexplorer 2
Removing air from the sample inlet tubing
If you have a Sample valve installed, a pre-made method template can
be used for purging the sample flowpath as described below.
Note: Never use any inlet filters on the sample inlet tubing in the
distilled water or the ethanol containers.
1 Prepare a container with degassed 96% ethanol and submerge the

sample inlet capillary connected to Sample valve port 7.
2 Prepare a container with degassed distilled water and submerge the

sample inlet capillary connected to port 6.
3 Prepare the sample containers with sample and submerge

the sample inlet capillaries connected to ports 1–5.
4 Submerge the sample inlet capillary connected to port 8 into a

sample path rinsing solution.
5 Manually, prime the sample inlet tubings with the actual sample.
6 In System Control, select File:Instant Run.
7 Select method template Purge Sample Pump and run it.
If no Sample valve is included, purge the tubing manually as follows:
1 Prepare a container with degassed distilled water and a container

with degassed 96% ethanol.
2 Submerge the sample inlet tubing in the distilled water.
3 Set the injection valve to position WASTE.
4 Manually, run the sample pump at 10 ml/min for 1 minute and then
stop the sample pump.
5 Move the sample inlet tubing carefully to the ethanol container.
6 Manually, run the sample pump at 50 ml/min for 2 minutes and then
7 Move the sample inlet tubing carefully back to the distilled water.
8 Manually, run the sample pump at 1 ml/min for 5 minutes.
Note: Be careful not to spill any liquid when moving the tubing.
Note: Remember to stop the sample flow before manually moving

the sample inlet tubing between containers.

2 ÄKTAexplorer ____________________________________________

priming the tubing. The calibration instruction is given in the Pump
P-950 User Manual.
Note: The injection valve must be in position WASTE when calibrating

the sample pump.
Preparing a method manually for direct loading onto column

If your optional configuration does not include a Sample valve, a
method must be prepared manually, as described in the following.
If a Sample valve is included, go directly to section Loading onto the

column.

SamplePres in the flow scheme and run data windows. Prepare a
method as follows:


Blue square
Set Base Volume
and
MethodBase SamplePump
3 Select Block:New.
4 Type a block name (e.g. Load_Column) and click OK.
5 The new block is inserted at the end of the method. Click on the new
block with the left mouse button, drag it up and drop it on the
blue square to open the new block to insert instructions.

_____________________________________________ ÄKTAexplorer 2
7 Insert the instruction DirectLoad.
8 Insert the instruction Method base: SystemPump directly after the

instruction DirectLoad.
9 Remove the Sample_injection block by clicking on it and then

clicking on the Delete button.
10 Create a variable for the parameter SampleFlow by clicking the

VAR... button. Select a name, e.g. SampleFlow. Create also a
variable for the parameter Volume.
Note: To avoid too high pressure in the system, adapt the sample
flow rate thoroughly to the viscosity of the sample.

instruction parameters
11 Insert the instruction Alarm_SamplePressure before the

DirectLoad instruction, and create a variable for the parameter
Hi-Alarm by clicking the VAR... button. Select a name, e.g.
Alarm_SamplePressure. This will protect the column from high
pressure generated by the sample pump when injecting sample onto
the column.
Loading onto the column

To load sample onto the column:
1 Make sure the sample pump is properly connected and prepared.
2 Select any of the templates starting with Direct Injection...(refer to

2.16 Strategy and template overview in ÄKTAexplorer) if a Sample
valve is installed. Otherwise, use your manually prepared method.

2 ÄKTAexplorer ____________________________________________
Loading a Superloop
The sample pump can be used to fill a Superloop. The Superloop allows
introduction of larger sample volumes into a pressurized fluid system.
The sample is pushed into the Superloop by the sample pump.
Superloop consists of a movable seal in a glass tube. The seal divides the
tube into two separate chambers. Depending on the flow direction, the
ml 10 seal moves towards either end piece of the glass tube.
5
Superloop is available in three sizes (10, 50, 150 ml) allowing
0
application of 1–10, 1–50 and 1–150 ml samples respectively.
When loading a Superloop, a Sample valve is usually not used.

1 Make sure that the Sample pump is properly installed according to
section Installing the components.

Use the Sample holder for storing

4
sample tubes and sample waste vessels
(5), and for routing the sample and
waste capillaries (4).
5

connections have been carried out
correctly.
Pump P-950
Outlet connection

Inlet connection

_____________________________________________ ÄKTAexplorer 2
components located
On-linefilter
Mixer Column
Selection
Valve
Injection (V2)
A B 2
17
6 2 2
18
7
34 5 3 6
4 5 COND UV
Restrictor
3
Buffer
A2 B1 B2 Fraction
W2 W1
collector
Sample
Super (optional)
1 Loop
Sample
Buffer Pump (P-950)
Outlet
2
3
18
7
6
4 2
18
7 2
3
1 8
7
6
3 6
45 4 5 45
Column
A11 A12 A13 A14 A15 A16 A17 A18 S1 Selection W3 F3
Valve (V3)

1
connector
2
3
4
Teflon i.d. 1.6 mm x 1300 mm Tubing connector for 1/8" o.d. tubing, 18-1121-17 Sample pump, inlet
with ferrule for 1/8" o.d. tubing, 18-1121-18. connector

2 ÄKTAexplorer ____________________________________________

2 Check that the Sample pump is indicated in the flow scheme

window.


• Fill a container with distilled water and submerge the sample inlet
capillary in the container.
• Set the injection valve to position INJECT.
• Set Flowrate to 5 ml/min and click on Execute. Allow the pump

to run for 3 minutes to prime the pump.
• Check that the sample pump is delivering flow.

Follow the procedure below to remove air from the sample flowpath.
Note: Remember to stop the sample flow before manually moving the
sample inlet tubing between containers.

3 Set the injection valve to position INJECT.
5 Move the sample inlet tubing carefully and submerge it in the

ethanol container.

_____________________________________________ ÄKTAexplorer 2
distilled water.
Note: Be careful not to spill any liquid when manually moving the
10 Calibrate the sample pump according to the instructions given in

the Pump P-950 User Manual.
Note: The injection valve must be in position INJECT when

calibrating the sample pump.
To fill the sample inlet tubing with sample:
1 Prepare a container with sample. If required, add an inlet filter to

the sample inlet tubing. Move it carefully and submerge it in the
sample container.
2 Prime the sample inlet tubing with the actual sample.
Preparing a method for loading a Superloop

method as follows:

3 Create a new block directly above the Sample injection block to

4 Select Block:New.
5 Type a block name (e.g. Load_Superloop) and click OK.
6 The new block is inserted at the end of the method. Click on the
new block with the left mouse button, drag it up and drop it on the

2 ÄKTAexplorer ____________________________________________
Blue square
Set Base Volume
and MethodBase
SamplePump
Create a variable for the parameter
SampleFlow to enable updating
from the column list.

Click here to define a instruction parameter
breakpoint variable
8 Insert the Flow instruction and select 0 ml/min.

to LOAD.
10 Insert the SampleFlow instruction and select 5 ml/min.

sample pump.
12 Create a breakpoint variable for the SampleFlow instruction and

select a variable name, e.g. Fill_Loop_With. A breakpoint variable
is created by clicking on Var in the Breakpointdialog window. This
variable is used to set the filling volume for the Superloop.
14 Insert the Flow instruction and set a suitable flow rate to start the
system pump.
15 Create a variable for the Flow instruction and select a variable name,
e.g. Flow. A variable is created by clicking on Var in the Parameters
dialog window. This variable is used for automatically adjusting the
flow rate to the selected column.
16 Save the method.

_____________________________________________ ÄKTAexplorer 2
2.12 Air sensors
General
Air sensors (AS1, AS2 and AS3) can be used in the ÄKTAexplorer flow
path. The examples below show two possible locations.
components located
Flow
17 Valve (V7)
2 6
34 5
Mixer Column
Selection
Valve
Injection (V2)
17 18
A B 2 6 2 7
34 5 3 6
4 5 pH COND UV
Restrictor
Sample Fraction
A2 B1 B2 Pump
W2 W1 collector
(optional)
Air cell
AS1

18 18 18 1 8
2 7 2 7 2 7 2 7
3 6 3 6 3 6 3 6
45 45 4 5 45
Column
A11 A12 A13 A14 A15 A16 A17 A18 S1 S2 S3 S4 S5 S6 S7 S8 Selection W3 F3 F4 F5 F6 F7 F8
Valve (V3)
One air cell located in the inlet flow of Pump P-900 for checking the buffer supply.

components located
Flow
17 Valve (V7)
2 6
34 5
Mixer Column
Selection
Valve
Injection (V2)
17 18
A B 2 6 2 7
34 5 3 6
4 5 pH COND UV
Restrictor
Sample
W2 W1
Collector
Air cell
AS2
18 18 18 1 8
2 7 2 7 2 7 2 7
3 6 3 6 3 6 3 6
45 45 4 5 45
Column
A11 A12 A13 A14 A15 A16 A17 A18 S1 S2 S3 S4 S5 S6 S7 S8 Selection W3 F3 F4 F5 F6 F7 F8
Valve (V3)
One air cell located in the sample inlet flow when applying sample directly onto the
column with a sample pump.
Air cells can be fitted in other locations in the ÄKTAexplorer main flow
path. The installation procedure is similar to the ones described below.

2 ÄKTAexplorer ____________________________________________
Installation

Fitting the Air-900 control box

1 Make sure that the ÄKTAFPLC chromatography system is switched
off.
2 Check that the ID switches on the connector panel of the Air-900

control box are set to code 0 (both switches in ON position).
LED ID switch (black) Code 0
ON
1 2 3 12
AIR CELL ID UniNet 2 1 2
3 Attach the Air-900 control box vertically to a suitable mounting

slot. Lock in position by pushing up the snap lock.
Release
Fasten
Connecting to the UniNet-2 communication network

Use the attached UniNet-2 cable to connect the Air-900 control box to
the UniNet-2 communication link. It can be connected anywhere
between the system pump P-900 and the termination plug.

_____________________________________________ ÄKTAexplorer 2
Using air cell Air-925 (AS1) in the system pump inlet flowpath
To system pump inlet 1 Remove the capillary from the Switch valve inlet A1.
2 Cut new Teflon capillaries as shown in the figure and attach the
purge valve and the air cell using the connectors and ferrules
supplied.
Purge
L1=920 mm Note: The length of L1 is dependent on which flow, sensitivity and
valve
type of instruction (alarm or watch) is used in UNICORN. The
optimal length of L1 must be tested in the user configuration.
Note: When running gradients, the air sensor detects smaller air
A-925 bubbles than expected. During a gradient, the flow rate through the
Air cell
air sensor is lower than the total flow rate (pump module A + pump
module B), but the air detection level is always based on the total
L2=300 mm flow rate.
3 Route the signal cable from the air cell to the Air cell 1 connector
Inlet filter on the Air-900 control box and plug it in.
Using air cell Air-925 (AS2) in the sample inlet flowpath

To sample pump inlet 1 Remove the sample inlet capillary from the Sample pump P-950
sample pump inlet connector.
Outlet connection
2 Cut the sample inlet capillary and attach the air
cell using the fingertight connectors supplied.
Purge
valve Note: The length of L1 is dependent on which
flow, sensitivity and type of instruction (alarm
or watch) is used in UNICORN. The optimal
length of L1 must be tested in the user
configuration.
A-925
Air cell Inlet connection
Note: Make sure that no air bubbles are
trapped in the sample pump flow path before
using the pump (refer to section 2.11 Sample pump P-950 and
Sample valve V5).
Inlet filter or
Sample valve
on the Air-900 control box and plug it in.

2 ÄKTAexplorer ____________________________________________
Operation
2 Go to System Control.
3 Select System:Settings.
4 Click the Monitors radio button. In the Instruction panel, select, e.g.,
AirSensor1. In the Parameters panel, select High_Sensitivity for the
air cell(s) to be used. Select the flow source for the air sensor, i.e.
SystemPump if the air cell is located in the main system flow path,
or SamplePump if the air cell is located in the sample injection flow
path. Scroll down to access air cell 3 if necessary.
Note: The sensitivity of the air cells can be selected at three levels;
high, medium or low sensitivity. We recommend you use the high
sensitivity setting as standard.
5 Check that the connected air cell(s) are indicated in the flow scheme
in UNICORN.
6 Verify that the air cells are functioning properly by issuing manual
commands from UNICORN as follows.

_____________________________________________ ÄKTAexplorer 2
Testing the AS1 air cell located in the main system flow path
1 Confirm that air sensor AS1 is shown in the flow scheme.
2 Fill a sample container with distilled water and submerge the inlet
filter of the A1 tubing. Purge the air cell using a syringe and the
purge valve mounted on the inlet capillary. The purge valve is
opened to the syringe by turning the syringe connector a 1/2 turn
counter clockwise.
3 Select System Control:Manual:Pump.
4 Select the instruction Flow in the instruction list.
5 Set Flowrate to 5 ml/min and click Execute.
6 While the system pump is running, check that the AS1 symbol in
the flow diagram is indicated with white colour.
7 Lift the inlet filter out of the flask and check that the white
indication changes to red when air enters the AS1 flow sensor.
8 Submerge the inlet filter and check that the red indication changes
back to white again. Stop the system pump by clicking End in the
System Control toolbar.
9 Set up instructions as described below to monitor the AS1 air cell

signal to suit your needs.
Testing the AS2 air cell located in the sample injection flow path
1 Make sure that the sample pump is functioning properly.
3 Fill a sample container with distilled water and submerge the

sample tubing.
4 Select System Control:Manual:Pump in UNICORN.
5 Select the instruction SampleFlow in the instruction list.
7 While the sample pump is running, check that the AS2 symbol in
8 Lift the sample tubing inlet out of the flask and check that the white
9 Submerge the sample tubing inlet and check that the red indication
changes back to white again. Stop the pump by clicking End in the

2 ÄKTAexplorer ____________________________________________
Testing an optionally located air cell

1 Depending on which connector in the control box you have
connected the air cell to, either AS1, AS2 or AS3 should be
connected.
2 Prepare ÄKTAexplorer to enable distilled water to be pumped

through the optionally located air cell.
4 Select the instruction Flow or SampleFlow in the instruction list,

depending on which location in the flow path you have selected for
the air cell.
5 Set Flow/Flowrate to 5.0 ml/min and click Execute.
6 While pumping is in progress, check that the air cell symbol in the
flow diagram is indicated with white colour.
7 Allow air to enter into the air cell and check that the white
indication changes to red when air enters the air cell.
back to white again. Stop the pump by clicking End in the System
Control control bar.
9 Set up instructions as described below to monitor the air cell signal

to suit your needs.
Setting up instructions for the air sensor function

The presence of air in the flow path is detected by the Air-900 control
box, which reports its status to UNICORN. The desired control action,
based on the status report, is governed by instructions in the method
used. In Instructions for handling the AS1, AS2 and AS3 air cells must
be inserted manually in a method.
Note: In ÄKTAexplorer 100 Air systems, the AS2 air sensor is

supported by pre-made method templates containing the word Air at
the end of the template name.
Alarm_AirSensor1-3
Enables/disables alarm generation for the air cells. An active alarm sets
the chromatography system to Pause mode. Alarms can be activated/
deactivated from System:Settings and inserted in a method.

_____________________________________________ ÄKTAexplorer 2
Watch_AirSensor1-3
Monitors the air sensor status signals from the air cells. Performs a user-
defined action if the test condition is fulfilled. Monitoring is cancelled
automatically once the condition has been fulfilled. The user-defined
action can be, for example, PAUSE or a call to a block.
Preparing a method for using air sensors

To use air cells in a method you should insert instructions to modify an
existing method or create a new method, see the following examples.
Example 1
In example 1, air sensor AS1 is used for checking the supply of buffer A.


Double-click on block Start_Conditions. Highlight the instruction
below which you want to insert the alarm air sensor instruction.
1. Double-click on block
Start_Conditions.
2. Highlight the instruction

below which you want to
insert the alarm air sensor
instruction.
5. Click on the Enable

radio button.
3. Click on the Alarm&Mon

radio button
4. Select Alarm_AirSensor1

2 ÄKTAexplorer ____________________________________________
3 Click the Alarm&Mon radio button.
4 From the Instructions list, select Alarm_AirSensor1.

If the air sensor detects air, e.g. if the buffer vessel is empty, the
system will be paused and the system pump stops. After filling the
buffer vessel the run can be resumed.
5 In Parameters, click the Enable radio button under Mode.
6 Click on Insert.
7 Save the method.
Example 2
In example 2, air sensor AS2 is used together with sample pump P-950
in a configuration designed to accomplish direct sample loading onto
the column and monitoring the sample stock.


Create a new block directly above the Sample_Injection block to
control the sample pump and to set a watch instruction for the air
sensor.
New block containing

instructions for controlling
the sample pump flow,
introducing the air sensor
watch condition and setting
up high alarm supervision for
the sample pump pressure.
*)
The variable High_alarm is
used for automatically
adjusting the flow rate to the
selected column. The
highlighted DirectLoad
instruction sets the system to
HOLD. When the sample
volume is injected and air is
detected, the system is set to Click here to create a
CONTINUE. variable
Note that MethodBase is

changed to SamplePump to
achieve correct volume
calculation.

_____________________________________________ ÄKTAexplorer 2
A new block is created in the following way:
3 Select Block:New.
4 Type a block name, e.g. Direct_Sample_Injection (see the

illustration) and click OK.
5 Insert the instruction as described in the figure.
6 The parameter High_alarm in the instruction

Alarm_SamplePressure must be a variable to automatically update
its value according to the selected column.
Sample_Injection block. Double-click (left mouse button) on the
8 Remove the Sample_Injection block by clicking on it and then

clicking on Delete.
9 At the dialogue window, click on Yes to remove the block

permanently, or click on No to be able to re-use the block.
10 Save the method.

2 ÄKTAexplorer ____________________________________________
2.13 Auxiliary equipment controlled via Pump P-901/903

REMOTE connector
Installation

installing the optional components.
Valve 1
Auxiliary equipment using digital input/output signals is connected to
the 9-pole D-SUB female REMOTE connector on the rear panel of Pump
P-900. Pins 1-4 handle digital input signals, and pins 6–9 handle digital
Valve A
Valve B
Pressure,
output signals. Pin 5 is signal ground.

analog
out 0 - 1V UniNet 2 Remote UniNet 1 Mains
The inputs are TTL compatible with pull-up resistors. Maximum input
REMOTE connector
voltage is 5.5 V d.c. The outputs are open collector transistors, each
on rear of P-901/903
capable of sinking 300 mA at maximum 30 V d.c.
Pin Signal Function

5 4 3 2 1 1-4 AuxIn 1-4 Active state can be selected to 0 or 1 in UNICORN.
9 8 7 6
Min. duration=50 ms. Max. frequency=1 Hz.
5 0V Signal ground
6-9 AuxOut 1-4 Open collectors. Active state can be selected to 0 or 1 in
UNICORN.
Use a shielded cable with a 9-pin D-SUB male connector for the P-900
REMOTE connector. The other end of the cable connects to the auxiliary
equipment and must be adapted to its relevant connector.
The connection example below shows the principle for connecting

input/output signals.
P-900
Digital signal interface
Pin 1 Pin 2 Pin 3 Pin 4 Pin 5 Pin 6 Pin 7 Pin 8 Pin 9 REMOTE
Connector
AuxIn 1 AuxIn 2 AuxIn 3 AuxIn 4 AuxOut 1 AuxOut 2 AuxOut 3 AuxOut 4
From external equipment outputs To external equipment inputs
Signal ground in external equipment

_____________________________________________ ÄKTAexplorer 2
Operation
Four digital input and four digital output signals can be handled via the
P-900 REMOTE connector. The input signals are designated AuxIn1-4
and the output signals are designated AuxOut1-4.
Instructions for signal handling

To set up signal handling in a method, the following parameters and
instructions must be inserted manually in a method.
AuxOut1-4
To set up and use the output signals, do the following:
3 Click on the Specials radio button.
4 In the Instruction panel, select the signal you want to use, e.g.,
AuxOut1.
5 Check in the external equipment User Manual which logical value,

0 or 1, that corresponds to inactive mode.
6 For AuxOut1 Parameters/Signal, set this value (0/1) as default. This

ensures that the external equipment is reset to inactive mode when
the method is completed.
7 Repeat this for all the output signals you want to use.

2 ÄKTAexplorer ____________________________________________
Watch_AuxIn1-4
These instructions set up the initial status of the input signals. The status
of the signals (1/0) can be shown in Run data in UNICORN.
The instructions also monitor the signal status for the input signals and
perform user-defined actions if test conditions are fulfilled. Monitoring
is cancelled automatically once a condition has been fulfilled. Pre-
selected action can be, for example, call to a block.
Curve representation of input/output signals

The status for signals AuxIn1 and AuxOut1 can be stored as curves in the
chromatogram. This is selected in System:Settings by setting
Curves:Store ON.
The parameter Time between samples is used to set the scan cycle for
curve updating. The shortest possible interval is 1 s.

_____________________________________________ ÄKTAexplorer 2
2.14 Superloop


(1–150 ml) onto the column. Superloop must not be used with columns
generating a column pressure higher than that stated in the table below.
If the column pressure is higher, the flow must be reduced in order not
to exceed the pressure limit. Set the pressure limit in the method with
ml 10 the variable Pressure_limit.
5
Superloop is an accessory available in three sizes:
0 Volume Max. allowed column pressure Code no.

10 ml 4 MPa 18-1113-81
50 ml 4 MPa 18-1113-82
150 ml 2 MPa 18-1023-85
All the sample is applied, which gives good reproducibility and high
recovery. The sample is not diluted as the buffer pushing the movable
seal is kept separate. The loaded sample can be injected all at once or
in several smaller volumes, down to 1 ml portions, permitting
automated repetition of sample injection.
Normally, Superloop is filled manually with a syringe. Use method

template Manual Injection_FP_Type of gradient.
Superloop can also be filled automatically, refer to section Loading a

Superloop on page 88.
Preparation
Prepare the injection valve and connect Superloop as follows:
1 Connect the supplied Pos. 1 LOAD

Union Luer female/1/16” 1
Column
male to port 3 of the 7
2
injection valve. Pump
Sample 3 6
2 Ensure that tubing for the syringe
waste is connected to port 4 4
5
Super
of the injection valve. loop
3 Ensure that Superloop is Waste Waste

filled with liquid (see
separate Superloop
Instruction).

2 ÄKTAexplorer ____________________________________________
4 Mount Superloop in a column holder as close to the injection valve

as possible.
5 Connect the bottom tubing to injection valve port 2.
6 Connect the top tubing to injection valve port 6.
7 Ensure all connections are fingertight.
Filling Superloop
Filling Superloop is achieved as follows:
2 Start Pump P-900 and let it run until the

movable seal has reached
the bottom of Superloop.
3 Stop Pump P-900 and set the injection valve

to position LOAD. ml 40
35
4 Load a large volume syringe with sample. 30
25
20
5 Gently load the syringe contents into 15
Superloop through port 3. 10
6 Leave the syringe in position. The loaded

sample can be injected all at once or in
several smaller volumes, down to 1 ml
portions. The volume to inject is set in the
method template by variable
Empty_loop_with.
7 The sample is applied to the column when the injection valve is set
to position INJECT. When the required volume has been injected,
set the valve to LOAD. When using method templates, this is
performed automatically.

_____________________________________________ ÄKTAexplorer 2
2.15 Configuration dependent delay volume in ÄKTAexplorer
The system configuration delay volume must be recalculated and
altered when standard configuration after the UV cell is changed to
optional configuration, or vice versa. This is set in parameter
System:Settings:Specials:FracParameters:DelayVol.
The standard configuration of an ÄKTAexplorer 10 system has the

following delay volume values:
Delay volume
Tubing kit incl. Frac-901 incl. Frac-950
0.25 mm i.d. tubing kit 105 µl 256 µl
The standard configuration of an ÄKTAexplorer 100 system (includes

the pH flow cell) has the following delay volume values:
Delay volume
The volumes calculated for Frac-950 include the accumulator with

tubing.
Calculate the contribution from each new component and capillary,

according to the table below, and add this additional volume to the
default value.
The example below shows a calculation for including the pH flow cell
in an ÄKTAexplorer 10 system.
Capillary volume + cell volume = 110/100 x 19.6 + 100 µl = 122 µl.
(The length of the additional i.d. 0.5 capillary is 110 mm, the volume
of 100 mm tubing is 19.6 µl, and the volume of the pH flow cell is
100µl.)
The new delay volume value is:
301 + 122= 423 µl (including Frac-901), and

451 + 122 = 573 µl (including Frac-950)

2 ÄKTAexplorer ____________________________________________
Component and capillary volumes

The volumes are theoretical values.
Component/capillary Volume (µl)

Valve PV-908 7
Valve IV-908 27
Valve INV-907 9
Valve FV-903 113
Flow cell UV-900, 2 mm 1
Conductivity flow cell 24
pH flow cell 100
Accumulator (Frac-950) 40
Finger-tight connector male/male 7
Flow restrictor FR-902 5
Capillary i.d. 0.25 x 100 mm 4.9

_____________________________________________ ÄKTAexplorer 2
2.16 Strategy and template overview in ÄKTAexplorer
In the table below the optional ÄKTAexplorer system configurations
are listed, and the additional components that are required.
System designation Optional components installed

Air Air sensors with control box
S Sample pump and sample selection valve
XT Autosampler
Templates overview
Type of ÄKTAexplorer system

Template name* Exp 100
Exp 100 Air Exp 10 Exp 10S Exp 10XT
Autosampler Injection FP BufferPrep X

Autosampler Injection FP Isocratic X
Autosampler Injection FP Segmented X
Autosampler Injection FP Step X
Chromatofocusing X X X X X
CIP X X X X X
Direct Injection FP BufferPrep X
Direct Injection FP BufferPrep Air X
Direct Injection FP Isocratic X
Direct Injection FP Isocratic Air X
Direct Injection FP Segmented X
Direct Injection FP Segmented Air X
Direct Injection FP Step X
Direct Injection FP Step Air X
Direct Injection V BufferPrep X
Direct Injection V BufferPrep Air X
Direct Injection V Isocratic X
Direct Injection V Isocratic Air X
Direct Injection V Segmented X
Direct Injection V Segmented Air X
Direct Injection V Step X
Direct Injection V Step Air X
Installation Test X X X X X
Manual Injection FP BufferPrep X X X X X
Manual Injection FP Isocratic X X X X X
Manual Injection FP Segmented X X X X X
Manual Injection FP Step X X X X X
Manual Injection V BufferPrep X X
Manual Injection V Isocratic X X
Manual Injection V Segmented X X

2 ÄKTAexplorer ____________________________________________
Type of ÄKTAexplorer system

Template name* Exp 100
Exp 100 Air Exp 10 Exp 10S Exp 10XT
Manual Injection V Step X X

Purge Sample Pump X X X
Sample Pump Loop Injection FP BufferPrep X X X
Sample Pump Loop Injection FP Isocratic X X X
Sample Pump Loop Injection FP Segmented X X X
Sample Pump Loop Injection FP Step X X X
Sample Pump Loop Injection V BufferPrep X X
Sample Pump Loop Injection V Isocratic X X
Sample Pump Loop Injection V Segmented X X
Sample Pump Loop Injection V Step X X
Oligo Nucleotide Purification X X
*
These template versions belong to the strategy versions listed below.
Strategies overview
ÄKTAexplorer system Strategy name*
Explorer 100 with Frac-901 E100_200
Explorer 100 with Frac-950 E100F200
Explorer 100 Air with Frac-901 ECA200
Explorer 100 Air with Frac-950 ECAF200
Explorer 10 with Frac-901 E10_200
Explorer 10 with Frac-950 E10F200
Explorer 10 S with Frac-901 E10S200
Explorer 10 S with Frac-950 E10SF200
Explorer 10 XT with Frac-901 E10X200
Explorer 10 XT with Frac-950 E10XF200
* The last three digits in the strategy name are the version
number.

______________________________________________ ÄKTApurifier 3
3 ÄKTApurifier
3.1 General
To utilise optional equipment in a chromatographic run, suitable

3.17 Configuration dependent delay volume in ÄKTApurifier.
The description below deals with instructions used in specific functions

3.2 Unpacking

that the ÄKTApurifier optional equipment is installed correctly.
3.3 Pre-requisites
connected to your ÄKTApurifer chromatography system.
• ÄKTApurifer must be installed and fully tested before the optional
components are installed. See the Installation Guide for your
ÄKTApurifer chromatography system.

components.
CAUTION! Make sure ÄKTApurifer is switched off before installing

the optional components. The mains power to ÄKTApurifer must be
UniNet-2 cables.

3 ÄKTApurifier _____________________________________________

mounted in the mounting rails on the side components (at opposite
of the modules. Both sides of the modules side too)
are provided with rails for quick
mounting of components.
Note: Make sure there is enough room on

the side of the system for installing the Monitor P-910
desired components.
When selecting mounting location for the

optional equipment it is important to Monitor
P-910
choose a location which minimize the

length of the capillaries used to connect
the optional components to the rest of the
system. Mon
itor P-91
0
Many components that are attached to

the mounting rails uses a snap-in bracket. The bracket is supplied
separately with the component and needs to be fitted as shown below
before the component can be attached.
2. Fit the attached

the two screws
1. Loosen the
two screws
Removing components

______________________________________________ ÄKTApurifier 3
installation.

valves)


Plug
Fr a c t i o n
collector
(optional)
To
Computer UniNet 2
Chain
UniNet 1
Chain
*

3 ÄKTApurifier _____________________________________________

To enable optional components to be used, UNICORN must be setup

optional component. For a overview of available method templates, see
3.18 Strategy and template overview in ÄKTApurifier

Function/

WasteBetweenTubes)
Peak_FracParameters_900 Start_Slope, End_Slope, Min-
Width

FeedTube_900 -
Peak_FracStop_900 -
DropSync, Tube)
Method/Manual:Frac Peak_FracParameters Start_Slope, End_Slope, Min-

Width
Fractionation Tube type (name from rack def-
inition)
number)
Base (Volume, Time)
FractionationStop -
FeedTube -
Peak_Fractionation Tube type (name from rack def-
inition)
number)
Base (Volume, Time)
Peak_FracStop -
AccumulatorWash Strokes (1–10 (5), OFF)

______________________________________________ ÄKTApurifier 3
component
(contin- tion)
Tube type (name from rack def-
inition)
number)
Base (Volume, Time)
tion)
inition)
number)
Base (Volume, Time)
Ignore_LastTube -
tion valve
selection
valve

HighAlarm (0-1 MPa)
LowAlarm (0-1 MPa)
Volume (0-1000 ml)

3 ÄKTApurifier _____________________________________________

component
SyrVol (100, 250, 500, 1000 µl)
MicroVial D)
HeadPress (Yes, No)
Scaling (0.1 - 1.0)

geEnd, NeedleHome,
NeedleFront)
WashExt (Yes, No)
WashExt (Yes, No)
TrpVialNo (1-160)
WashExt (Yes, No)
Level_1_Standard,
Level_20_Standard)
Air sensor1 A-9001 System:Settings:Monitors AirSensor1-3 Mode ( High_Sensitivity,
SamplePump)
Manual:Alarms&Monitors Alarm_AirSensor1-3 Mode (Enabled, Disabled)1

View:Windows:Instruction Watch_AirSensor1-3 Test Equal (Value 0, 1)

PAUSE, HOLD/END_BLOCK,


______________________________________________ ÄKTApurifier 3
Preparation for use
To start up your ÄKTApurifier optional configuration system:
1 Switch on ÄKTApurifier with the mains switch located at the

front left on the system base.
3 Log in (see chapter 2 in Making your first run).
see 3.18 Strategy and template overview in ÄKTApurifier. A new
strategy is installed according to the software installation

creating a new method or opening an existing method.
tubing.
is used, the backpressure from the tubing might become higher than the
maximum pressure for the column and the run will stop immediately
after it is started.

3 ÄKTApurifier _____________________________________________
General
purifications with ÄKTApurifier. It collects up to 175 fractions in
12 mm diameter tubes, up to 95 fractions in 10–18 mm diameter tubes,
and up to 40 fractions in 30 mm diameter tubes.

Installation
On-line
filter Column
COND
Flow restrictor
Mixer
Injection
Valve (V1) UV
17
Pump A Pump B 2 6
34 5
Fraction
Sample loop
Collector
A2 A1 B2 B1 W2 W1 8
1 7 Frac-901
W3 2 6
345
Outlet
Valve (V4)
F3
User Manual.

bottle.
34

______________________________________________ ÄKTApurifier 3
• Connecting a UniNet-1 cable (35) between Pump P-900 and

Frac-901.
• Connecting a UniNet-1 cable (36) between Frac-901 and the

computer.
1 2
36 35
6 Connect a mains cable supplied (38) between Frac-901 and a mains

socket at the rear of ÄKTApurifier.
1 2
39
38
38
To Frac-901
To mains socket

3 ÄKTApurifier _____________________________________________
Operation
1 Start your ÄKTApurifier optional configuration system as

Frac.

Frac-901.



elution.

two are used simultaneuously.

two are used simultaneuously, even if the command is issued during a
peak.



______________________________________________ ÄKTApurifier 3
Note: When using Frac-901 in ÄKTApurifier 10 systems, a maximum

is used.
• Peak_Frac_Size


3 ÄKTApurifier _____________________________________________
General
purifications with ÄKTApurifier. Up
to 392 fractions can be collected in

row, serpentine-column or column- FRAC 950
by-column.

available.

Installation
CAUTION! Make sure ÄKTApurifier is switched off before

ÄKTApurifier must be switched OFF before disconnecting or
To install Frac-950:


go to step 2.

______________________________________________ ÄKTApurifier 3
6 Put Frac-950 on the workbench to the right of ÄKTApurifier.
want to include the accumulator in Frac-950 or not.
Connect port 2 on the outlet valve in ÄKTApurifier to Frac-950 as
On-line
filter Column
COND
Flow restrictor
Mixer
Injection
Valve (V1) UV
17
Pump A Pump B 2 6
34 5
Fraction
Sample loop
Collector
A2 A1 B2 B1 W2 W1 8
1 7 Frac-950
W3 2 6
345
Outlet
Valve (V4)
F3

correctly.

3 ÄKTApurifier _____________________________________________
Operation

Frac-950.

The instruction Fractionation starts eluate fractionation. The

instruction PeakFractionation starts peak fractionation.

elution.


peak.



______________________________________________ ÄKTApurifier 3

alarm is generated and the fraction collecter goes to waste. If:

position.



3 ÄKTApurifier _____________________________________________


a set %B window:

peak size dependent, can be controlled. One fixed volume size before
the eluate:



End_Peak_Frac_at
Note: When using Frac-950 in ÄKTApurifier 10 systems, a maximum

is used.

______________________________________________ ÄKTApurifier 3
Continue with setting up the rack characteristics. Go to variable page
Frac-950.

30 mm tubes
18 mm tubes
30 mm tubes

3 ÄKTApurifier _____________________________________________
General
Auto sampler A-900 is intended for automated multiple sample
injections. A-900 is only used in ÄKTApurifier 10 systems.

a few microlitres to 1 ml in a simple and reproducible manner.
Buffer tubing
High pressure valve
Tubing guide
Needle arm
Tray segment
Syringe waste
Syringe dispenser

bottle position
and leakage drain
• Flushed loop
• µl pick-up

______________________________________________ ÄKTApurifier 3
valve.

Installation
Preparations
from the ÄKTApurifier base platform.

ÄKTApurifier, with approximately 3 cm spacing.
Locking screw
pur i fier
ÄKTA
Red cap



8VHU0DQXDO&KDSWHU

3 ÄKTApurifier _____________________________________________
CAUTION! The mains power to ÄKTApurifier must be switched


1 Make sure that the ÄKTApurifier chromatography system is

switched off.

Note: If a fraction collector Frac-901/Frac-950 is used, disconnect the

Ter mi nator
P l ug
Frac ti on
C o l l ec tor
Disconnect at the
computer end, and
route to the A-900
UniNet 1 connector
To
C o mp u t e r Uni Net 2
Chai n
Uni Net 1
Chai n

______________________________________________ ÄKTApurifier 3
in A-900.
Te r mi n a t i o n
plug
Fr a c t i o n !
Co l l e c t o r
115 V
1
0
To
C om pu t e r UniNet 2
Chain
UniNet 1
Chain
Four green LEDs
UniNet-1 connection
base platform of the ÄKTApurifier separation unit.

3 ÄKTApurifier _____________________________________________
The capillaries to be connected between A-900 and the ÄKTApurifier
separation unit are included in ÄKTApurifier 10 XT Tubing kit, code
no. 18-1122-65. The flow diagram below shows the location of A-900
in the ÄKTApurifier flow path.
Autosampler A-900
21 Injection
3 6
45 valve
W4
Sample
loop
G-A1
G-A2
(H-A2)
On-line
filter Column
COND Flow restrictor
Mixer
Injection
valve (V1) UV
17
Pump A Pump B 2 6
34 5
Fraction
collector
Sample loop (optional)
8
1 7
2 6 Outlet valve (V4)
345
1 Remove the capillary connecting the Injection valve V1 §with the

Column top inlet.

ÄKTApurifier after the column. Use the capillaries marked G-A1
and G-A2 if you intend to use the 0.25 tubing kit, and the
capillaries marked G-A1 and H-A2 if you intend to use the 0.50
tubing kit.
Note: See section 2.1 in the ÄKTApurifier System Manual for gen-
eral recommendations on selecting column and tubing kit.
3 Both capillaries are routed in the space between the UV monitor

UV-900 and the pH/Conductivity monitor pH/C-900.
4 Connect the capillary marked G-A1 (l=750 mm) from the tubing kit
the ÄKTApurifier Injection valve V1.

______________________________________________ ÄKTApurifier 3
5 Connect the capillary marked G-A2/H-A2 (l=800 mm) from the
“column”), and the UV flow cell inlet.





Control:Manual:Flowpath:AutosamplerControl
on the A-900.

3 ÄKTApurifier _____________________________________________
SyringeEnd aspirates a syringe volume of wash solvent from the

wash solvent bottle and fills the wash solvent tubing.
solvent.

alternately.
can be necessary).

be clearly visible.
3 Carry out a manual injection by issuing the


vial bottom.

Note: Check the needle height with 2 mm as default setting when-

ever the needle is replaced. The 2 mm setting is only a relative meas-
ure to ensure that the needle stops above the vial bottom regardless
of which type of vial used. Adjust the final setting in small steps un-
til a needle height suitable for the actual use is obtained (type of vi-
al, sample volume available, etc.).

______________________________________________ ÄKTApurifier 3
Installation test
General
parts of the ÄKTApurifier separation unit. This is to establish that the
integration of the optional components in ÄKTApurifier has not
Preparation
instructions in the ÄKTApurifier Installation Guide.

solution.

standard tray.
5 Immerse the inlet tubing B1 into the rest of the 0.4% acetone
solution.
6 Immerse the inlet tubing A11 into a flask containing distilled water.

Main menu button
button
11 Click on Run.
again.

3 ÄKTApurifier _____________________________________________

30 minutes.
100%
95%

Autosampler
peaks 1 and 2
30%
Gradient 5%


of the test result.

If any of the evaluated values fall outside the specified range, refer to

______________________________________________ ÄKTApurifier 3
Manual evaluation



right corner.



3 ÄKTApurifier _____________________________________________

and enter the values in column 2 in the UV response table of the

Test record.

discontinuities.


column 4.
column 5.


Calculate the absorbance ratios 265 nm/254 nm and 265 nm/280

nm using the values from column 3 and enter the results in
column 4.
column 5.

______________________________________________ ÄKTApurifier 3

• Faulty gradient

piston sealings in the pump may be the cause. Refer to the Pump
P-900 User Manual.
Manual.
(5% faulty=pump module B, 95% faulty=pump module A). Re-
fer to the Pump P-900 User Manual.
performed. Repeat the test and re-evaluate. Refer to the Pump
P-900 User Manual.

3 ÄKTApurifier _____________________________________________
Test record

1 2 3 4 5
(mAU) value
100
95 94 - 96
70 69 - 71
30 29 - 31
5 4-6
0

UV response table:
1 2 3 4 5
value
254
265/254 1.11 - 1.26
265
265/280 1.26 - 1.53
280

1 2 3 4
(%)
1 66.1 - 67.2
2 32.8 - 33.9

______________________________________________ ÄKTApurifier 3
Operation
General
A-900 is controlled from a PC running UNICORN version 3.2 or
higher. It cannot be used as a stand-alone module. A-900 can be
controlled, either automatically from a pre-made method template, or
manually via commands.

3.18 Strategy and template overview in ÄKTApurifier).
The A-900 can be used for automatic sample injection from a few
microlitres to one millilitre of sample solution. To achieve optimal

• InjectionPickup

• InitialWash
• RotateTray
• SyringeHome/End


3 ÄKTApurifier _____________________________________________


sample.

modes.

vial used.
ringeSpeed Normal
High
(Normal is default)
(1.0 is default)
InjectionFlushed
The InjectionFlushed instruction selects the flushed loop injection
mode for A-900.

the sample loop is filled with 1.5 - 3 time the sample loop volume
(depending on the loop volume). The injection volume equals the loop
must be changed in System:Settings (100 µl is the default loop
volume).

______________________________________________ ÄKTApurifier 3

internally.

on the outside.


InjectionPartial
mode for A-900.


internally.

on the outside.


InjectionPick-up
for A-900.

3 ÄKTApurifier _____________________________________________

internally.

on the outside.


InitialWash
the syringe.
The syringe is first washed using one syringe volume.
Then the needle is moved to its home position and washed internally.
Finally, the needle moves to its front position and is washed both
internally and externally.
RotateTray
SyringeHome/End

container.
SyringeEnd draws wash solvent into the syringe from the wash bottle.
NeedleHome/Front
is replaced.

______________________________________________ ÄKTApurifier 3
eluents.
• Low
• High

250 µl High (5 - 10 cP) Low 0.2

3 ÄKTApurifier _____________________________________________
3.8 Column selection valves
Installation
On-line filter
Mixer
Injection
Pump Pump valve (V1)
17 18
A B 2 6 2 7
34 5 3 6
4 5 COND UV
pH
Flow restrictor
Column
selection
valve (V2)
A1 A2 B1 B2
W2 W1
Fraction
collector
Column Outlet
selection
valve (V3) valve (V4)
18 1 8
2 7 2 7
3 6 3 6
4 5 45
W3 F3

1 Mark the two PV-908 valves with Column valve 2 and Column
valve 3. Check that the ID code switch is set to 2 and 3 respectively.
2 Mount the brackets and attach the two valves to a system mounting
rail.
the correct i.d. as required by your system.
Mount one capillary between ports 1 on Column valve 2 and
Column valve 3. This is the column bypass position used as default
position for the valves in UNICORN.
4 Use the attached UniNet-2 cables to connect the valves to the

UniNet-2 communication link. The valves can be connected
plug connected to the injection valve.
correctly.

______________________________________________ ÄKTApurifier 3
Operation
2 The column selection valves are automatically recognised by

UNICORN at system start-up. Check that the valves added are
indicated in Flow scheme in UNICORN.
3 Verify that the ColumnPosition valves are functioning properly by

• Select instruction ColumnPosition.
• Set the valves to position 4.
• Manually, run the pump P-900 with distilled water at a flow rate
of approximately 2 ml/min and check that water is coming out
from port 4 of Column Valve 2.
• Connect a capillary between port 4 of Column Valve 2 and port

4 of Column Valve 3 and check that water is coming out from
the centre port of Column Valve 3.
Using the column selection function in a run

The column selection valves are designated ColumnValves in the flow
scheme and run data windows.
To use the column selection valves in a method, do as follows:

Select System:Technique:Template: and For column:. Click OK.

column selection instruction, e.g. Start_Conditions. Highlight the

3 ÄKTApurifier _____________________________________________
Click here to define a variable

name for the instruction parameter
4 From the Instructions list, select ColumnPosition. The valve

instruction parameter Position allows you to change all eight
positions of the column selection valves synchronously.

ColumnValves_Pos. This variable allows you to synchronously
control the position of the column valves to the position you want.

variable ColumnValves_Pos to the desired position.
8 Save the method.

______________________________________________ ÄKTApurifier 3
3.9 Buffer selection valves
Installation
On-line filter
Mixer Column
Injection
17
A B 2 6
34 5
COND UV
A
Restrictor
Fraction
A2 B1 collector
W2 W1
(optional)
Buffer Outlet
18 1 8
2 7 2 7
3 6 3 6
45 45
A11 A12 A13 A14 A15 A16 A17 A18 W3 F3 F4 F5 F6 F7 F8

1 Mark the IV-908 valve with Buffer Valve A. Check that the ID code
switch is set to 5.
2 Mount the brackets and attach the valve to the system rack.
3 Cut and mount the required teflon tubings (i.d. 1.6 mm in

ÄKTApurifier 10 and i.d. 2.9 mm in ÄKTApurifier 100).

plug connected to the outlet valve.
correctly.

3 ÄKTApurifier _____________________________________________
Operation
2 The buffer selection valve is automatically recognised by

UNICORN at system start-up. Check that the valve added are
indicated in the Flow scheme in UNICORN.
3 Verify that BufferValve A is functioning properly by issuing manual

• Select instruction BufferValve A.
• Set BufferValve A to position 4.
• Manually, run pump P-900 with distilled water at a flow rate of

approximately 2 ml/min and check that water is drawn through
port 4 in BufferValve A.
Using the buffer selection function in a run

The buffer valve is designated BufferValveA in the flow scheme and run
data windows. In the run data window, it is designated V5_BufferA.
To use the valve in a method, do as follows:


buffer valve instruction, e.g. Start_Conditions_zz. Highlight the
4 From the Instructions list, select BufferValveA. The valve

positions of the buffer valve individually.

______________________________________________ ÄKTApurifier 3

5 Define variable names for the buffer selection instruction

parameters, e.g. BufferValveA_Pos. These variables allow you to
control the position of the buffer valves individually to the position
you want.
6 Click OK. The defined variables are inserted in the method.

variable BufferValveA_Pos to the desired position.
8 Save the method.

3 ÄKTApurifier _____________________________________________
Installation
Flow Direction
Valve (V7)
17
2 6
34 5
COND UV
Flow restrictor
Column
On-line
filter
Mixer
Injection
Valve (V1) Fraction
17
Pump A Pump B 2 6 collector
34 5
(optional)
8
1 7
W3 2 6
345 Outlet Valve (V4)
NC NO NC NO Sample loop
A2 A1 B2 B1 W2 W1
F3

installing the optional components. The mains power ÄKTApurifier
must be switched OFF before disconnecting or connecting the
UniNet-2 cables.
1 Mark the INV-907 valve with Flow Direction Valve V7. Check that
the ID code switch is set to 7.
2 Attach the valve to a system mounting rail. Use the mounting

bracket and screws supplied with the valve.

1 1
2 7 Pump 7 Pump
2
To UV To UV
monitor 3
3 6 monitor 6
4 5 4 5
Stop plug Stop plug

______________________________________________ ÄKTApurifier 3
plug.
correctly.
Operation

3 Temporarily, replace the column with a transparent bypass

capillary.


wards the bypass capillary replacing the column.

• When completed, remove the bypass capillary.

3 ÄKTApurifier _____________________________________________



flow direction valve instruction, e.g. Start_Conditions. Highlight
the instruction below which you want to insert the valve
instruction.


instruction parameter Position allows you to change position for
the flow direction valve using parameters DownFlow and UpFlow.

______________________________________________ ÄKTApurifier 3

8 Save the method.

3 ÄKTApurifier _____________________________________________
Installation

1 Mark the optional function valve with Valve 8 or Valve 9 as

required. Check that the ID code switch is set to 8 or 9 respectively.
2 Mount the bracket and attach the valve to a mounting rail on the
system. Choose the best location to facilitate the routing of the
tubing that connects the valve to the main system flowpath.
broadening effects.

correctly.

______________________________________________ ÄKTApurifier 3
Operation

system start-up. Check that the added valve is indicated in the flow
scheme in UNICORN.

• Set the valve to position 3.


as follows:

Select View:Text instructions to display the text instruction editor.
instruction.
selected.

3 ÄKTApurifier _____________________________________________


the port you want.

7 Save the method.

______________________________________________ ÄKTApurifier 3
General
Using a pH electrode is optional in ÄKTApurifier, and therefore, it is
not mounted from the factory. The pH electrode is to be mounted in a
separate pH flow cell, housing the pH electrode itself, as well as a
dummy pH electrode. The dummy pH electrode replaces the pH
electrode whenever the pH electrode is to be removed from the system,
e.g. for storage.

refilled, an internal electrolyte bridge of 4M KCl saturated with
Ag/AgCl, an outer electrolyte bridge of 1M KNO 3, an annular ceramic
Note: Always use the 0.50 tubing kit when the flow cell is mounted.
Note: Using the flow cell is not recommended when the peak volume
is less than 1 ml. The volume in the flow cell decreases the UHVROXWLRQRI
WKHSHDNV
Installation
WARNING! Never use 0.25 mm i.d. tubing (blue) when the pH

flow cell is mounted in the system. The electrode may rupture due to
high back-pressure.
1 Install the pH flow cell holder (packed in Box-900) in the groove

directly below the Outlet Valve (V4).
New capillary of New capillary of
180 mm 130 mm
pH flow cell
On-line
filter Column
COND
Flow restrictor
Mixer
G6
Injection (H6)
Valve (V1) UV
17
Pump B Pump A 2 6
34 5
Fraction
Collector
NC NO NC NO Sample loop (optional)
A2 A1 B2 B1 W2 W1 8
1 7
2 6 Outlet Valve (V4)
345

3 ÄKTApurifier _____________________________________________
2 Push the slide clamp upwards to secure the pH flow cell holder.
3 Connect the new capillary of 180 mm to the inlet of the pH flow

cell.
4 Connect the outlet of the pH flow cell to the FR-904 flow restrictor
(marked IN) using the new capillary of 130 mm.
5 Connect the outlet of the FR-904 flow restrictor to the centre port
of the V4 outlet valve using the G8 capillary.
1
6 Place the FR-904 flow restrictor (1) in the hole in the pH flow cell
holder.
2
7 Mount the pH electrode (2) for experiments where pH is to be

measured. Otherwise, mount the pH electrode dummy in the hole
in the pH flow cell.
Note: When using the pH electrode, the flow restrictor FR-904 must be
replaced with the supplied flow restrictor FR-902. Otherwise, the long
term stability and lifetime of the pH electrode will deteriorate.
Note: For calibration of the pH electrode, refer to the UNICORN

User Manual.
Operation
2 If fraction collection should be used, remember to change the delay

volume. This setting is found in the System Control menu;
System:Settings:Special. See also 3.17 Configuration dependent
delay volume in ÄKTApurifier.
Using pH measurement in a run is described in the ÄKTApurifier

System Manual.

______________________________________________ ÄKTApurifier 3
Pump P-950 is a membrane type

50 ml/min.
A Sample valve PV-908, with a pressure limit of


samples.
The capillaries used between the Sample valve

inlets and the Sample vessels are cut in suitable
lengths from the tubing supplied, code no.
18-1119-74. Use the fingertight connectors
supplied.
There are three ways of using Pump P-950 in the system:
Each one requires a specific installation configuration and different

procedures to remove air from the flowpath inside the sample pump,
which is described in the following.


3 ÄKTApurifier _____________________________________________
CAUTION! The mains power to ÄKTApurifier must be switched


connect the capillaries, see the section that describes the specific sample

1 Make sure that the ÄKTApurifier
chromatography system is
switched off.
INV-908
2 Install the Sample pump P-950

vertically in the separation unit P-950
of ÄKTApurifier (also refer to

the Pump P-950 User Manual). PV-908

the clamp supplied with the
pump.
M-925
• Fit the pump using the clamp

and the screws.
Sample Pump P-950

1 If included, install the Sample valve V5 (PV-908) above
Pump P-950 (also refer to IV/PV-908 Instructions).





______________________________________________ ÄKTApurifier 3
1 From the Injection valve V1 UniNet-2 connector, disconnect the
termination plug.
2 If no Sample valve is to be Sample valve

installed, connect a new
cable (supplied) between
Injection valve V1 and a
Sample pump UniNet-2
connector (the dotted line). PV-908
INV-908
If the Sample valve is P-950
installed, do as follows:
• Connect a new cable (sup-

PV-908
plied) between the Injec-

tion valve and a Sample
valve V5 UniNet-2 con- M-925
nector. Loosen the V5 at-

tachment bracket if it is
difficult to plug in the con-
nector.
• Connect the second new Termination plug

the free Sample valve V5 UniNet-2 connector and the Sample
pump P-950 UniNet-2 connector.
3 Connect the termination plug to the free UniNet-2 connector on the

Sample pump.

3 ÄKTApurifier _____________________________________________

applied repeatedly, for example, in scouting runs. By using sample
loops supplied by Amersham Pharmacia Biotech, volumes between
0.1–2.0 ml can be applied. The sample is drawn into the sample loop
by the sample pump.
sample loop. Cavitation may occur due to the resistance in the
flowpath.

components.

3 Check that all capillary and electrical connections have been

carried out correctly.
Outlet connection

Inlet connection

______________________________________________ ÄKTApurifier 3
On-linefilter
Column
Mixer
Injection
17
A B 2 6
34 5
3
COND UV
Sample
Pump
Restrictor
P-950
2 4 Fraction
A2 B1 B2
Collector
W2 W1
1
18 18 1 8
2 7 2 7 2 7
3
45
6 5 3
45
6 3
45
6
A11 A12 A13 A14 A15 A16 A17 A18 S1 S2 S3 S4 S5 S6 S7 S8 W3 F3 F4 F5 F6 F7 F8

1
port*
2
3
4
5
port 1-8
*

3 ÄKTApurifier _____________________________________________

2 Check that the Sample pump and the Sample valve (if installed) are
3 Make sure the system is connected as previously described in

section Connecting the capillaries on page 164.


container.

select S4.




______________________________________________ ÄKTApurifier 3

with sample.
filled with sample.


3 ÄKTApurifier _____________________________________________

P-950 User Manual.

the sample pump.

shown below.


3 Select Block:New.

*
*
*
added manually.
manually.

to LOAD.

______________________________________________ ÄKTApurifier 3

sample pump.


breakpoint variable
loop.
12 Save the method.


3 ÄKTApurifier _____________________________________________


components.


On-linefilter
Mixer
Column
2 Injection
17
A B 2 6
3
34 5
COND UV
Restrictor
A 4
Sample
P-950 W2 W1
Collector
5
1
18 18 1 8
2 7 2 7 2 7
3
45
6 6 3
45
6 3
45
6
A11 A12 A13 A14 A15 A16 A17 A18 S1 S2 S3 S4 S5 S6 S7 S8 W3 F3 F4 F5 F6 F7 F8

______________________________________________ ÄKTApurifier 3
Pump P-950
Outlet connection

Inlet connection

1
o.d. tubing, 18-1121-17
2
connector
3
4
5
6

length.

3 ÄKTApurifier _____________________________________________




container.

select S4.



______________________________________________ ÄKTApurifier 3
If no Sample valve is included, purge the tubing as follows:


10 If the running conditions are changed, calibrate the sample pump

according to the instructions in the Pump P-950 User Manual.
Note: The injection valve must be in position WASTE when

To fill the sample tubing with sample:

sample container.

3 ÄKTApurifier _____________________________________________
If a sample selection valve is used (assumed to be connected as

SampleValve in this example), proceed as follows to remove air from
the sample flowpath:



5 In the Method Editor, select File:New.
6 Select No template and click on OK.
7 Select View:Text Instructions and prepare a method as shown

below.


9 Save the method and run it. It takes approx. 12 minutes to

complete.

______________________________________________ ÄKTApurifier 3
method as follows:


Blue square
Set Base Volume
and
3 Select Block:New.



3 ÄKTApurifier _____________________________________________

flow rate thoroughly to the viscosity of the sample.


the column.

______________________________________________ ÄKTApurifier 3

3 ÄKTApurifier _____________________________________________
Loading a Superloop
Superloop consists of a movable seal in a glass tube. The seal divides

the tube into two separate chambers. Depending on the flow direction,
ml 10 the seal moves towards either end piece of the glass tube.
5
0



Pump P-950
Outlet connection

Inlet connection

______________________________________________ ÄKTApurifier 3
On-linefilter
Mixer
Column
Injection
17
A B 2 6
34 5 2
COND UV
3
Buffer
Fraction
Restrictor
A2 B1 B2
W2 W1
Collector
Sample
Super
1 Loop
Sample
Pump (P-950)
Buffer Outlet
18 1 8
2
3
45
7
6 4 2
3
45
7
6
A11 A12 A13 A14 A15 A16 A17 A18 A18 W3 F3 F4 F5 F6 F7 F8

1
connector
2
3
4

3 ÄKTApurifier _____________________________________________


window.



inlet capillary in the container.




ethanol container.

______________________________________________ ÄKTApurifier 3
distilled water.



sample container.

method as follows:


4 Select Block:New.

3 ÄKTApurifier _____________________________________________
Blue square
Set Base Volume
and MethodBase
SamplePump

breakpoint variable

to LOAD.

sample pump.

system pump.
15 Create a variable for the Flow instruction and select a variable

name, e.g. Flow. A variable is created by clicking on Var in the
Parametersdialog window. This variable is used for automatically
adjusting the flow rate to the selected column.
16 Save the method.

______________________________________________ ÄKTApurifier 3
3.14 Air sensors
General
Air sensors (AS1, AS2 and AS3) can be used in the ÄKTApurifier flow
On-line filter
Mixer Column
Injection
17
A B 2 6
34 5
COND UV
Restrictor
Fraction
A2 B1 collector
W2 W1
(optional)
Air cell
AS1
Buffer
A1
1 8
2 7
3 6
45
W3 F3
On-line filter
Mixer Column
Injection
17
A B 2 6
34 5
COND UV
Restrictor
Sample Fraction
A1 A2 B1 B2 Pump
W2 W1 collector
(optional)
Purge valve
Air cell
AS2
Sample container
1 8
2 7
3 6
45
W3 F3
Air cells can be fitted in other locations in the ÄKTApurifier main flow

3 ÄKTApurifier _____________________________________________
Installation
CAUTION! Make sure ÄKTApurifier is switched off before installing

the optional components. The mains power to ÄKTApurifier must be
cables.

1 Make sure that the ÄKTAbasic chromatography system is switched
off.

ON
1 2 3 12

Release
Fasten


______________________________________________ ÄKTApurifier 3
supplied.
Purge
valve
Air cell

Outlet connection
Purge
configuration.
A-925
Sample valve V5).
Inlet filter or
Sample valve

3 ÄKTApurifier _____________________________________________
Operation
4 Click the Monitors radio button. In the Instruction panel, select,

e.g., AirSensor1. In the Parameters panel, select High_Sensitivity
for the air cell(s) to be used. Select the flow source for the air sensor,
i.e. SystemPump if the air cell is located in the main system flow
path, or SamplePump if the air cell is located in the sample injection
flow path. Scroll down to access air cell 3 if necessary.
in UNICORN.

______________________________________________ ÄKTApurifier 3
filter of the A1 tubing. Purge the air cell using a syringe and the
purge valve mounted on the inlet capillary. The purge valve is
opened to the syringe by turning the syringe connector a 1/2 turn
counter clockwise.


sample tubing.

3 ÄKTApurifier _____________________________________________

connected.
2 Prepare ÄKTApurifier to enable distilled water to be pumped

through the optionally located air cell.

the air cell.

to suit your needs.

Alarm_AirSensor1-3

______________________________________________ ÄKTApurifier 3
Watch_AirSensor1-3

Example 1


Start_Conditions.

instruction.

radio button.

radio button

3 ÄKTApurifier _____________________________________________

6 Click on Insert.
7 Save the method.
Example 2


sensor.

*)
CONTINUE. variable

calculation.

______________________________________________ ÄKTApurifier 3
3 Select Block:New.



clicking on Delete.

10 Save the method.

3 ÄKTApurifier _____________________________________________

REMOTE connector
Installation
CAUTION! Make sure ÄKTApurifier is switched off before installing

the optional components.
Valve 1
Valve A
Valve B
Pressure,

analog
REMOTE connector
Pin Signal Function

9 8 7 6
5 0V Signal ground
6-9 AuxOut 1-4 Open collectors. Active state can be selected to 0 or 1
in UNICORN.

P-900
Connector

______________________________________________ ÄKTApurifier 3
Operation

AuxOut1-4
AuxOut1.



3 ÄKTApurifier _____________________________________________
Watch_AuxIn1-4
These instructions set up the initial status of the input signals. The
status of the signals (1/0) can be shown in Run data in UNICORN.

Curves:Store ON.

______________________________________________ ÄKTApurifier 3
3.16 Superloop


5

10 ml 4 MPa 18-1113-81
50 ml 4 MPa 18-1113-82
150 ml 2 MPa 18-1023-85


Preparation

Column
2
Sample 3 6
5
Super

separate Superloop
Instruction).

3 ÄKTApurifier _____________________________________________

as possible.
Filling Superloop

movable seal has reached the bottom of the
Superloop.

35
25
20

Empty_loop_with.

______________________________________________ ÄKTApurifier 3
3.17 Configuration dependent delay volume in ÄKTApurifier
The standard configuration of an ÄKTApurifier 10 system has the

Delay volume
The standard configuration of an ÄKTApurifier 100 system has the

Delay volume
The volumes calculated for Frac-950 include the accumulator with

tubing.

default value. The example below shows a calculation for including the
pH flow cell in an ÄKTApurifier 10 system.
Capillary volume + cell volume = 110/100 x 19.6 + 100 µl = 122 µl.
(The length of the additional i.d. 0.5 capillary is 110 mm, the volume
of 100 mm tubing is 19.6 µl, and the volume of the pH flow cell is
100µl.)
232 + 122= 354 µl (including Frac-901), and


3 ÄKTApurifier _____________________________________________


Valve PV-908 7
Valve IV-908 27
Valve INV-907 9
Valve FV-903 113
pH flow cell 100

______________________________________________ ÄKTApurifier 3
3.18 Strategy and template overview in ÄKTApurifier
The table below lists the additional component that is required for a
specific optional system configuration.
System designation Optional component installed

XT Autosampler
Templates overview
Type of ÄKTApurifier system
Template name*
Pur 100 Pur 10 Pur 10XT
Chromatofocusing X X X
CIP X X X
Installation Test X X X
Manual Injection FP BufferPrep X X X
Manual Injection FP Isocratic X X X
Manual Injection FP Segmented X X X
Manual Injection FP Step X X X
Manual Injection V BufferPrep X
Manual Injection V Isocratic X
Manual Injection V Segmented X
Manual Injection V Step X
* These template versions belong to the strategy versions listed

below.
Strategies overview
ÄKTApurifier system Strategy name*
Purifier 100 with Frac-901 P100_201
Purifier 100 with Frac-950 P100F201
Purifier 10 with Frac-901 P10_200
Purifier 10 with Frac-950 P10F201
Purifier 10 XT with Frac-901 P10X200
Purifier 10 XT with Frac-950 P10XF201
*
The last three digits in the strategy name are the version
number.

_______________________________________________ ÄKTAbasic 4
4 ÄKTAbasic
4.1 General
To utilise optional equipment in a chromatographic run, suitable

4.17 Configuration dependent delay volume in ÄKTAbasic.
The description below deals with instructions used in specific functions

4.2 Unpacking

that the ÄKTAbasic optional equipment is installed correctly.
4.3 Pre-requisites
connected to your ÄKTAbasic chromatography system.
• ÄKTAbasic must be installed and fully tested before the optional
ÄKTAbasic chromatography system.

components.
CAUTION! Make sure ÄKTAbasic is switched off before installing

the optional components. The mains power to ÄKTAbasic must be
UniNet-2 cables.

4 ÄKTAbasic _______________________________________________

mounted in the mounting rails on components (at opposite
the side of the modules. Both sides side too)
of the modules are provided with
rails for quick mounting of
components.
Note: Make sure there is enough

room on the side of the system for
installing the desired components.
When selecting mounting location

for the optional equipment it is
important to choose a location
which minimize the length of the
capillaries used to connect the
optional components to the rest of
the system.
Many components that are attached to the mounting rails uses a snap-
in bracket. The bracket is supplied separately with the component and
needs to be fitted as shown below before the component can be
attached.
2. Fit the attached

the two screws
1. Loosen the
two screws
Removing components

_______________________________________________ ÄKTAbasic 4
installation.

valves)


Plug
Fr a c t i o n
collector
(optional)
To
Computer UniNet 2
Chain
UniNet 1
Chain
*

4 ÄKTAbasic _______________________________________________
To enable optional components to be used, UNICORN must be set up

optional component. For a overview of available method templates, see
4.18 Strategy and template overview in ÄKTAbasic

Function/

WasteBetweenTubes)
Peak_FracParameters_900 Start_Slope, End_Slope, Min-
Width

FeedTube_900 -
Peak_FracStop_900 -
DropSync, Tube)
Method/Manual:Frac Peak_FracParameters Start_Slope, End_Slope, Min-

Width
Fractionation Tube type (name from rack def-
inition)
number)
Base (Volume, Time)
FractionationStop -
FeedTube -
Peak_Fractionation Tube type (name from rack def-
inition)
number)
Base (Volume, Time)
Peak_FracStop -

_______________________________________________ ÄKTAbasic 4
component
(contin- tion)
inition)
number)
Base (Volume, Time)
tion)
inition)
number)
Base (Volume, Time)
Ignore_LastTube -
tion valve
Outlet valve PV-908 Manual:Flowpath OutletValve Position (Feed, F1-F8)
Manual:Frac OutletFractions How Many? (0-7)
Volume (5-10000 ml)
Start at (F2-F8)
selection
valve

HighAlarm (0-1 MPa)
LowAlarm (0-1 MPa)
Volume (0-1000 ml)

4 ÄKTAbasic _______________________________________________
component
SyrVol (100, 250, 500, 1000 µl)
MicroVial D)
HeadPress (Yes, No)
Scaling (0.1 - 1.0)

geEnd, NeedleHome,
NeedleFront)
WashExt (Yes, No)
WashExt (Yes, No)
TrpVialNo (1-160)
WashExt (Yes, No)
Level_1_Standard,
Level_20_Standard)
Air sensor1 A-9001 System:Settings:Monitors AirSensor1-3 Mode ( High_Sensitivity,
SamplePump)
Manual:Alarms&Monitors Alarm_AirSensor1-3 Mode (Enabled, Disabled)1


PAUSE, HOLD/END_BLOCK,


_______________________________________________ ÄKTAbasic 4
Preparation for use
To start up your ÄKTAbasic optional configuration system:
1 Switch on ÄKTAbasic with the mains switch located at the rear

of the system.
3 Log in (see chapter 3 in ÄKTAbasic User Manual).
see 4.18 Strategy and template overview in ÄKTAbasic. A new
strategy is installed according to the software installation

creating a new method or opening an existing method.
tubing.
is used, the backpressure from the tubing might become higher than the
maximum pressure for the column and the run will stop immediately
after it is started.

4 ÄKTAbasic _______________________________________________
General
purifications with ÄKTAbasic. It collects up to 175 fractions in 12 mm
diameter tubes, up to 95 fractions in 10–18 mm diameter tubes, and up
to 40 fractions in 30 mm diameter tubes.

Note: When installing Frac-901 in an ÄKTAbasic system, an Outlet

valve must also be installed (refer to section 4.8 Outlet valve).
Installation
Column
Flow restrictor
Mixer UV
Injection
Valve (V1)
17
Pump A Pump B 2 6
34 5
A1 B1 Fraction
Sample loop
Collector
W2 W1 8
1 7 Frac-901
W3 2 6
345
Outlet
Valve (V4)
F3
User Manual.
34 4 Extend the drain tubing from Frac-901 (34) to a suitable waste
bottle.

_______________________________________________ ÄKTAbasic 4
• Connecting a UniNet-1 cable between Pump P-900 and

Frac-901.
• Connecting a UniNet-1 cable between Frac-901 and the

computer.
Termination
plug
Frac-901
Pump P-900
1 2
UniNet 2
UniNet 1
To computer
6 Connect a mains cable supplied between Frac-901 and a mains

socket at the rear of ÄKTAbasic.
Frac-901
To
computer
Mains
inlet
1 2

4 ÄKTAbasic _______________________________________________
Operation
1 Start your ÄKTAbasic optional configuration system as described
in Preparation for use.

Frac.

Frac-901.



elution.


peak.



_______________________________________________ ÄKTAbasic 4
Note: When using Frac-901 in ÄKTAbasic 10 systems, a maximum

is used.
• Peak_Frac_Size


4 ÄKTAbasic _______________________________________________
General
purifications with ÄKTAbasic. Up to
392 fractions can be collected in

by-column.

available.

Installation

UniNet-2 cables.
To install Frac-950:


go to step 2.

_______________________________________________ ÄKTAbasic 4
6 Put Frac-950 on the workbench to the right of ÄKTAbasic.
want to include the accumulator in Frac-950 or not.
Connect port 2 on the outlet valve in ÄKTAbasic to Frac-950 as
Column
Flow restrictor
Mixer
UV
Injection
Valve (V1)
17
Pump A Pump B 2 6
34 5
Fraction
Sample loop
Collector
A1 B1 W2 W1 8
1 7 Frac-950
W3 2 6
345
Outlet
Valve (V4)
F3

correctly.

4 ÄKTAbasic _______________________________________________
Operation

Frac-950.



elution.


peak.



_______________________________________________ ÄKTAbasic 4


position.



4 ÄKTAbasic _______________________________________________

a set %B window:

peak size dependent, can be controlled. One fixed volume size before
the eluate:



End_Peak_Frac_at
Note: When using Frac-950 in ÄKTAbasic 10 systems, a maximum

is used.

_______________________________________________ ÄKTAbasic 4
Continue with setting up the rack characteristics. Go to variable page
Frac-950.

30 mm tubes
18 mm tubes
30 mm tubes

4 ÄKTAbasic _______________________________________________
General
Autosampler A-900 is intended for automated multiple sample
injections. Note that A-900 is only used in ÄKTAbasic 10 systems.

a few microlitres to 1 ml in a simple and reproducible manner.
Buffer tubing
High pressure valve
Tubing guide
Needle arm
Tray segment
Syringe waste
Syringe dispenser

bottle position
and leakage drain
• Flushed loop
• µl pick-up

_______________________________________________ ÄKTAbasic 4
valve.

Installation
Preparations
from the rear of the ÄKTAbasic system.

ÄKTAbasic, with approximately 3 cm spacing.
Locking screw
Red cap



8VHU0DQXDO&KDSWHU

4 ÄKTAbasic _______________________________________________
CAUTION! The mains power to ÄKTAbasic must be switched OFF

before disconnecting or connecting the UniNet-1 cables.


off.

Note: If a fraction collector Frac-901/Frac-950 is used, disconnect the

Termination
plug
Frac-901
Pump P-900
1 2
UniNet 2
UniNet 1
To computer
Disconnect at the
computer end, and
route to the A-900
UniNet 1 connector

_______________________________________________ ÄKTAbasic 4
in A-900.
Termination
plug
Fraction
collector
UniNet 2 115 V
chain 1
0
UniNet 1
chain To computer
Four green LEDs
UniNet-1 connection
base platform of the ÄKTAbasic separation unit.

4 ÄKTAbasic _______________________________________________
The capillaries to be connected between A-900 and the ÄKTAbasic
separation unit are included in ÄKTApurifier 10 XT Tubing kit, code
no. 18-1122-65. The flow diagram below shows the location of A-900
in the ÄKTAbasic flow path.
Autosampler A-900
21 Injection
3 6
4 5 valve
W4
Sample
loop
G-A1
G-A2
(H-A2)
Column
Mixer
Injection
valve (V1) UV Flow restrictor
17
Pump A Pump B 2 6
34 5
Sample loop
1 Remove the capillary connecting the Injection valve V1 with the

Column top inlet.

ÄKTAbasic after the column. Use the capillaries marked G-A1 and
G-A2 if you intend to use the 0.25 tubing kit, and the capillaries
marked G-A1 and H-A2 if you intend to use the 0.50 tubing kit.
Note: See section 2.1 in the ÄKTAbasic User Manual for general
recommendations on selecting column and tubing kit.
3 Connect the capillary marked G-A1 (l=750 mm) from the tubing kit
the ÄKTAbasic Injection valve V1.

_______________________________________________ ÄKTAbasic 4
4 Connect the capillary marked G-A2/H-A2 (l=800 mm) from the
“column”), and the UV flow cell inlet.





Control:Manual:Flowpath:AutosamplerControl
on the A-900.

4 ÄKTAbasic _______________________________________________
SyringeEnd aspirates a syringe volume of wash solvent from the

wash solvent bottle and fills the wash solvent tubing.
solvent.

alternately.
can be necessary).

be clearly visible.
3 Carry out a manual injection by issuing the


vial bottom.

Note: Check the needle height with 2 mm as default setting when-

ever the needle is replaced. The 2 mm setting is only a relative meas-
ure to ensure that the needle stops above the vial bottom regardless
of which type of vial used. Adjust the final setting in small steps un-
til a needle height suitable for the actual use is obtained (type of vi-
al, sample volume available, etc.).

_______________________________________________ ÄKTAbasic 4
Installation test
General
parts of the ÄKTAbasic separation unit. This is to establish that the
integration of the optional components in ÄKTAbasic has not
Preparation
instructions in the ÄKTAbasic Installation Guide.

solution.

standard tray.
5 Immerse inlet tubing B into the rest of the 0.4% acetone solution.
6 Immerse inlet tubing A into a flask containing distilled water.

Main menu button
button
11 Click on Run.
again.

4 ÄKTAbasic _______________________________________________
30 minutes.
100%
95%

Autosampler
peaks 1 and 2
30%
Gradient 5%


of the test result.

If any of the evaluated values fall outside the specified range, refer to

_______________________________________________ ÄKTAbasic 4
Manual evaluation



right corner.



4 ÄKTAbasic _______________________________________________
and enter the values in column 2 in the UV response table of the

Test record.

discontinuities.


column 4.
column 5.


Calculate the absorbance ratios 265 nm/254 nm and 265 nm/

280 nm using the values from column 3 and enter the results in
column 4.
column 5.

_______________________________________________ ÄKTAbasic 4

• Faulty gradient

piston sealings in the pump may be the cause. Refer to the Pump
P-900 User Manual.
Manual.
(5% faulty=pump module B, 95% faulty=pump module A).
Refer to the Pump P-900 User Manual.
performed. Repeat the test and re-evaluate. Refer to the Pump
P-900 User Manual.

4 ÄKTAbasic _______________________________________________
Test record

1 2 3 4 5
(mAU) value
100
95 94 - 96
70 69 - 71
30 29 - 31
5 4-6
0

UV response table:
1 2 3 4 5
value
254
265/254 1.11 - 1.26
265
265/280 1.26 - 1.53
280

1 2 3 4
(%)
1 66.1 - 67.2
2 32.8 - 33.9

_______________________________________________ ÄKTAbasic 4
Operation
General
Autosampler A-900 is controlled from a PC running UNICORN
version 3.2 or higher. It cannot be used as a stand-alone module. A-900
can be controlled, either automatically from a pre-made method
template, or manually via commands.

4.18 Strategy and template overview in ÄKTAbasic).
The A-900 can be used for automatic sample injection from a few
microlitres to one millilitre of sample solution. To achieve optimal

• InjectionPickup

• InitialWash
• RotateTray
• SyringeHome/End


4 ÄKTAbasic _______________________________________________



sample.

modes.



vial used.

ringeSpeed Normal
High
(Normal is default)

(1.0 is default)
InjectionFlushed
The InjectionFlushed instruction selects the flushed loop injection
mode for A-900.

the sample loop is filled with 1.5–3 times the sample loop volume
(depending on the loop volume). The injection volume equals the loop
must be changed in System:Settings (100 µl is the default loop
volume).

_______________________________________________ ÄKTAbasic 4

internally.

on the outside.


InjectionPartial
mode for A-900.


internally.

on the outside.


InjectionPickup
for A-900.

4 ÄKTAbasic _______________________________________________

internally.

on the outside.


InitialWash
the syringe.
The syringe is first washed using one syringe volume. Then the needle
is moved to its home position and washed internally. Finally, the needle
moves to its front position and is washed both internally and externally.
RotateTray
SyringeHome/End

container.
SyringeEnd draws wash solvent into the syringe from the wash bottle.
NeedleHome/Front
is replaced.

_______________________________________________ ÄKTAbasic 4
eluents.
• Low
• High

250 µl High (5 - 10 cP) Low 0.2

4 ÄKTAbasic _______________________________________________
4.8 Outlet valve
Installation
Column
Flow restrictor
Mixer UV
Injection
Valve (V1)
17
Pump A Pump B 2 6
34 5
Fraction
collector
A1 B1
Sample loop
1 8
W2 W1 2 7 Outlet
3 6
45 Valve (V4)
W3 F3 F4 F5 F6 F7 F8

UniNet-2 cables.
1 Mark the PV-908 valve with Outlet Valve 4. Check that the ID code
switch is set to 4.
2 Mount the bracket and attach the valve to a mounting rail.
3 Cut and mount the new capillaries. Use a capillary with the correct
i.d. as required by your system.



_______________________________________________ ÄKTAbasic 4
Operation
in section Preparation for use.
2 The outlet valve is automatically recognized by UNICORN at

system start-up. Check that the valve added is indicated in the flow
scheme in UNICORN.
3 Verify that the outlet valve is functioning properly by issuing

• Select instruction OutletValve.
• Set the outlet valve to position 4.

from port 4 of the Outlet valve.
Using the outlet valve for fractionation

Two different fractionation instructions are available to use with the
outlet valve:
• OutletValve
This instruction parameter turns the outlet valve to one of the eight
positions available. This makes the instruction suitable for
controlling flow-through fractionation before elution starts.
• OutletFractions
This instruction has three parameters, How_many?, Volume and
Start_at. This makes the instruction suitable for controlling large
volume fractionation (min. 5 ml) during elution. Both instructions
are independent of using Fraction collector Frac-901 or Frac-950.
Note: Installing an outlet valve in the flow path affects the system delay
volume. See section 4.17 Configuration dependent delay volume in
ÄKTAbasic for instructions on how to re-calculate and change the
delay volume.

4 ÄKTAbasic _______________________________________________
Collecting flow-through fractions with the outlet valve
This instruction is designated OutletValve in the flow scheme and
V4_Outlet in the run data window. To use the instruction in a method,
proceed as follows:


Outlet valve instruction, e.g. Flowthrough_Fractionation. Highlight
the instruction below which you want to insert the flowthrough
fractionation instruction.
Delete the
FractionCollector
instruction.

3 Select the FractionCollector instruction and press Delete.
5 From the Instructions list, select OutletValve. The valve instruction

parameter Position allows you to change all eight positions of the
outlet valve.

_______________________________________________ ÄKTAbasic 4
6 Define a variable name for the outlet valve instruction parameter,
e.g. OutletValve_Pos. This variable allows you to control the
position of the outlet valve to the position you want.
8 Insert an instruction to reset the valve to the desired position after

flowthrough fractionation is terminated.

variable OutletValve_Pos to the desired position.
10 Save the method.

4 ÄKTAbasic _______________________________________________
Collecting large fractions with the outlet valve
This instruction is designated OutletFractions. To use the instruction in
a method, proceed as follows:


fractionation instruction, e.g. Eluate_Fractionation. Highlight the
instruction below which you want to insert the fractionation
instruction.
Delete the old

FractionCollector
instruction and add the
new OutletFractions
instruction.
Click here to define

variable names for the
4 Click on the Frac button.
5 From the Instructions list, select OutletFractions. The instruction

parameters allow you to select manually how many fractions to
collect, the volume of each fraction and which position to start
from.

_______________________________________________ ÄKTAbasic 4
6 Click the upper Var button to define a variable name for how many
fractions to collect, e.g. No_of_Fracs. This variable allows you to
control the number of fractions to collect. When the number of
fractions have been collected, the outlet valve is set to start position.
Note: The instruction parameter How_many? can also be used to

terminate outlet fractionation by setting it to 0. The outlet valve is
then reset to start position.
8 Click the middle Var button to define a variable name for the size
(volume) of the fractions to collect, e.g. FracSize_Vol. This variable
allows you to control the volume of the fractions to collect.
10 Click the lower Var button to define a variable name for the port to
start from, e.g. Start_at_Pos. This variable allows you to control
the position of the outlet valve from which you start collecting
fractions from.

variables created above to the desired values.
13 Save the method.

4 ÄKTAbasic _______________________________________________
Installation
Mixer
UV
Injection
Flow restrictor
Pump Pump valve (V1)
17 18
A B 2 6 2 7
34 5 3 6
4 5
Column
selection
valve (V2)
A1 B1
W2 W1
W3
Column
selection
valve (V3)
18
2 7
3 6
4 5

UniNet-2 cables.
2 Mount the brackets and attach the two valves to a system mounting
rail.
the correct i.d. as required by your system.
Mount one capillary between ports 1 on Column valve 2 and
Column valve 3. This is the column bypass position used as default
position for the valves in UNICORN.

correctly.

_______________________________________________ ÄKTAbasic 4
Operation
2 The column selection valves are automatically recognised by


from port 4 of Column Valve 2.
• Connect a capillary between port 4 of Column Valve 2 and port

4 of Column Valve 3 and check that water is coming out from the
centre port of Column Valve 3.

scheme and run data windows.
To use the column selection valves in a method, do as follows:


column selection instruction, e.g. Start_Conditions. Highlight the

4 ÄKTAbasic _______________________________________________




8 Save the method.

_______________________________________________ ÄKTAbasic 4
Installation
Mixer Column
Injection
17
A B 2
34 5
6
Flow
restrictor
UV
B1
W2 W1
W3
Buffer
Valve (V5)
18
2 7
3 6
45
A11 A12 A13 A14 A15 A16 A17 A18

cables.
1 Mark the IV-908 valve with Buffer Valve A. Check that the ID code
switch is set to 5.
2 Mount the brackets and attach the valve to a mounting rail on the
system.
3 Cut and mount the required teflon tubings (i.d. 1.6 mm in

ÄKTAbasic 10 and i.d. 2.9 mm in ÄKTAbasic 100).

plug connected to the outlet valve.
correctly.

4 ÄKTAbasic _______________________________________________
Operation
2 The buffer selection valve is automatically recognised by

UNICORN at system start-up. Check that the valve added are
indicated in the Flow scheme in UNICORN.
3 Verify that BufferValve A is functioning properly by issuing manual

• Select instruction BufferValve A.
• Set BufferValve A to position 4.

approximately 2 ml/min and check that water is drawn through
port 4 in BufferValve A.

The buffer valve is designated BufferValveA in the flow scheme and run
data windows. In the run data window, it is designated V5_BufferA.
To use the valve in a method, do as follows:


4 From the Instructions list, select BufferValveA. The valve

positions of the buffer valve individually.

_______________________________________________ ÄKTAbasic 4


parameters, e.g. BufferValveA_Pos. These variables allow you to
control the position of the buffer valves individually to the position
you want.

variable BufferValveA_Pos to the desired position.
8 Save the method.

4 ÄKTAbasic _______________________________________________
Installation
Flow Direction
Valve (V7)
17
2 6
34 5
UV
Flow restrictor
Column
Mixer
Injection
Valve (V1)
17
Pump A Pump B 2 6
34 5
W3
Sample loop
A1 B1
W2 W1

the optional components. The mains power ÄKTAbasic must be
cables.
1 Mark the INV-907 valve with Flow Direction Valve V7. Check that
the ID code switch is set to 7.
2 Attach the valve to a system mounting rail. Use the mounting

bracket and screws supplied with the valve.

1 1
2 7 Pump 7 Pump
2
To UV To UV
monitor 3
3 6 monitor 6
4 5 4 5
Stop plug Stop plug

_______________________________________________ ÄKTAbasic 4
plug.
correctly.
Operation

3 Temporarily, replace the column with a transparent bypass

capillary.



• When completed, remove the bypass capillary.

4 ÄKTAbasic _______________________________________________


flow direction valve instruction, e.g. Start_Conditions. Highlight
instruction.


instruction parameter Position allows you to change position for
the flow direction valve using parameters DownFlow and UpFlow.

_______________________________________________ ÄKTAbasic 4

8 Save the method.

4 ÄKTAbasic _______________________________________________
Installation

cables.
1 Mark the optional function valve with Valve 8 or Valve 9 as

required. Check that the ID code switch is set to 8 or 9 respectively.
2 Mount the bracket and attach the valve to a mounting rail on the
system. Choose the best location to facilitate the routing of the
tubing that connects the valve to the main system flowpath.
broadening effects.

correctly.

_______________________________________________ ÄKTAbasic 4
Operation

system start-up. Check that the added valve is indicated in the flow
scheme in UNICORN.

• Set the valve to position 3.


as follows:

Select View:Text instructions to display the text instruction editor.
instruction.
selected.

4 ÄKTAbasic _______________________________________________


the port you want.

7 Save the method.

_______________________________________________ ÄKTAbasic 4

50 ml/min.
A Sample valve PV-908, with a pressure limit of


samples.
The capillaries used between the Sample valve

inlets and the Sample vessels are cut in suitable
lengths from the tubing supplied, code no.
18-1119-74. Use the fingertight connectors
supplied.
There are three ways of using Pump P-950 in the system:

procedures to remove air from the flowpath inside the sample pump,
which is described in the following.


4 ÄKTAbasic _______________________________________________
CAUTION! The mains power to ÄKTAbasic must be switched OFF

before disconnecting or connecting the UniNet-2 cables.

connect the capillaries, see the section that describes the specific sample

1 Make sure that the ÄKTAbasic
chromatography system is
switched off.
2 Install the Sample pump P-950

vertically in the separation unit
of ÄKTAbasic (also refer to the
P-950
Pump P-950 User Manual).

the clamp supplied with the
pump.
• Fit the pump using the clamp

and the screws.
Pump P-950

1 If included, install the Sample valve V5 (PV-908) at a suitable
location on the system (also refer to IV/PV-908 Instructions).





_______________________________________________ ÄKTAbasic 4
1 From the Injection valve V1 UniNet-2 connector, disconnect the
termination plug.
2 If no Sample valve is to be Sample valve

installed, connect a new (placed at a suitable
location)
Injection valve V1 and a
Sample pump UniNet-2
connector (the dotted line).
If the Sample valve is PV-908
installed, do as follows:
P-950
• Connect a new cable

(supplied) between the INV-907
Injection valve and a

Sample valve V5 UniNet-
2 connector. Loosen the
V5 attachment bracket if M-925
it is difficult to plug in
the connector.
• Connect the second new Termination plug

the free Sample valve V5 UniNet-2 connector and the Sample
pump P-950 UniNet-2 connector.
3 Connect the termination plug to the free UniNet-2 connector on the

Sample pump.

4 ÄKTAbasic _______________________________________________
by the sample pump.
sample loop. Cavitation may occur due to the resistance in the
flowpath.

components.


Outlet connection

Inlet connection

_______________________________________________ ÄKTAbasic 4
Column
Mixer
Injection
17
A B 2 6
34 5
3
Sample
Pump
P-950 UV
A1 B1 2 4
Restrictor
W2 W1
Sample
Valve (V5)
18
5 2
3
7
6
45 W3
S1 S2 S3 S4 S5 S6 S7 S8

1
port*
2
3
4
5
port 1-8
*

4 ÄKTAbasic _______________________________________________
in Preparation for use in section 4.4 General guidelines.
2 Check that the Sample pump and the Sample valve (if installed) are



container.

select S4.




_______________________________________________ ÄKTAbasic 4

with sample.
filled with sample.


4 ÄKTAbasic _______________________________________________
P-950 User Manual.

the sample pump.

shown below.


3 Select Block:New.

*
*
*
added manually.
manually.

to LOAD.

_______________________________________________ ÄKTAbasic 4

sample pump.


breakpoint variable
loop.
12 Save the method.


4 ÄKTAbasic _______________________________________________

components.


Mixer
Column
2 Injection
17
A B 2 6 3
34 5
4
Sample
A2 B2 Pump
W1
UV
P-950 W2
Restrictor
5
1
Sample
Valve (V5)
18
6 2
3
7
6
45
W1
S1 S2 S3 S4 S5 S6 S7 S8

_______________________________________________ ÄKTAbasic 4
Pump P-950
Outlet connection

Inlet connection

1
o.d. tubing, 18-1121-17
2
connector
3
4
5
6

length.

4 ÄKTAbasic _______________________________________________



container.

select S4.



_______________________________________________ ÄKTAbasic 4
If no Sample valve is included, purge the tubing as follows:





sample container.

4 ÄKTAbasic _______________________________________________
If a sample selection valve is used (assumed to be connected as
SampleValve in this example), proceed as follows to remove air from
the sample flowpath:




below.


9 Save the method and run it. It takes approx. 12 minutes to

complete.

_______________________________________________ ÄKTAbasic 4
method as follows:


Blue square
Set Base Volume
and
3 Select Block:New.



4 ÄKTAbasic _______________________________________________
Note: To avoid too high pressure in the system, adjust the sample
flow rate thoroughly according to the viscosity of the sample.


the column.

_______________________________________________ ÄKTAbasic 4

4 ÄKTAbasic _______________________________________________
Loading a Superloop

ml 10 the seal moves towards either end piece of the glass tube.
5
0



Pump P-950
Outlet connection

Inlet connection

_______________________________________________ ÄKTAbasic 4
Mixer
Column
Injection
17
A B 2 6
34 5 2
3
Buffer
A1 B1
W2 W1
Sample
UV
Restrictor
Super
1 Loop
Sample
Pump (P-950)
4
W3
A18

1
connector
2
3
4

4 ÄKTAbasic _______________________________________________

window.







ethanol container.

_______________________________________________ ÄKTAbasic 4
distilled water.



sample container.

method as follows:


4 Select Block:New.

4 ÄKTAbasic _______________________________________________
Blue square
Set Base Volume

and MethodBase
SamplePump

breakpoint variable

to LOAD.

sample pump.

system pump.

16 Save the method.

_______________________________________________ ÄKTAbasic 4
4.14 Air sensors
General
Air sensors (AS1, AS2 and AS3) can be used in the ÄKTAbasic flow
Mixer Column
Injection
17
A B 2 6
34 5
B1 UV
Restrictor
Air cell W2 W1
AS1
Buffer
A1
W3
Mixer Column
Injection
17
A B 2 6
34 5
Sample UV
A1 B1
Restrictor
Pump
W2 W1
Purge valve
Air cell
AS2
Sample container
W3
Air cells can be fitted in other locations in the ÄKTAbasic main flow

4 ÄKTAbasic _______________________________________________
Installation

cables.

off.

ON
1 2 3 12

Release
Fasten


_______________________________________________ ÄKTAbasic 4
supplied.
Purge
valve
Air cell

Outlet connection
Purge
configuration.
A-925
Sample valve V5).
Inlet filter or
Sample valve

4 ÄKTAbasic _______________________________________________
Operation

in UNICORN.

_______________________________________________ ÄKTAbasic 4
filter of the A tubing. Purge the air cell using a syringe and the purge
valve mounted on the inlet capillary. The purge valve is opened to
the syringe by turning the syringe connector a 1/2 turn counter
clockwise.


sample tubing.

4 ÄKTAbasic _______________________________________________
connected.
2 Prepare ÄKTAbasic to enable distilled water to be pumped through

the optionally located air cell.

the air cell.

to suit your needs.

Alarm_AirSensor1-3

_______________________________________________ ÄKTAbasic 4
Watch_AirSensor1-3

Example 1


Start_Conditions.

instruction.

radio button.

radio button

4 ÄKTAbasic _______________________________________________

6 Click on Insert.
7 Save the method.
Example 2


sensor.

*)
CONTINUE. variable

calculation.

_______________________________________________ ÄKTAbasic 4
3 Select Block:New.



clicking on Delete.

10 Save the method.

4 ÄKTAbasic _______________________________________________
REMOTE connector
Installation

the optional components.
Valve 1
Valve A
Valve B
Pressure,

analog
REMOTE connector
Pin Signal Function

9 8 7 6
5 0V Signal ground
6-9 AuxOut 1-4 Open collectors. Active state can be selected to 0 or 1
in UNICORN.

P-900
Connector

_______________________________________________ ÄKTAbasic 4
Operation

AuxOut1-4
AuxOut1.



4 ÄKTAbasic _______________________________________________
Watch_AuxIn1-4

Curves:Store ON.

_______________________________________________ ÄKTAbasic 4
4.16 Superloop


5

10 ml 4 MPa 18-1113-81
50 ml 4 MPa 18-1113-82
150 ml 2 MPa 18-1023-85

template Manual Injection FP Type of gradient.

Preparation

Column
2
Sample 3 6
5
Super

separate Superloop
Instruction).

4 ÄKTAbasic _______________________________________________
as possible.
Filling Superloop

movable seal has reached the bottom of the
Superloop.

35
25
20

Empty_loop_with.

_______________________________________________ ÄKTAbasic 4
4.17 Configuration dependent delay volume in ÄKTAbasic
The example below shows how to calculate the delay volume in an

ÄKTAbasic 10 system when using Frac-901 and an Outlet valve.
Volume [µl] Component

Delay volume = 4.0 + UV cell volume/2
7.5 + Union connector
5.0 + Flow restrictor FR-902
9.8 + FR-902 – Outlet valve, 200 mm
7.0 + Outlet valve
29.4 Outlet valve – Fraction collector, 600 mm
Total volume 62.6 µl
To calculate the volume of a tubing:
Volume of 200 mm of i.d. 0.25 mm tubing =
200/100 x 4.9 = 9.8 µl (4.9 µl = volume of 100 mm tubing, from

table on the next page)
If using Frac-950 in the example above, the delay volume is 196,3 µl.
This includes the accumulator with tubing but no Outlet valve.
The corresponding volumes in an ÄKTAbasic 100 system would be as

follows:
• 374.0 µl with Frac-901 installed
• 429.1 µl with Frac-950 installed

4 ÄKTAbasic _______________________________________________

Valve PV-908 7
Valve IV-908 27
Valve INV-907 9
Valve FV-903 113

_______________________________________________ ÄKTAbasic 4
4.18 Strategy and template overview in ÄKTAbasic
The table below lists the additional component that is required for a
specific optional system configuration.
System designation Optional component installed

XT Autosampler
Templates overview
Type of ÄKTAbasic system
Template name*
Bas 100 Bas 10 Bas 10XT
Chromatofocusing X X X
Installation Test X X X
Manual Injection FP Isocratic X X X
Manual Injection FP Segmented X X X
Manual Injection FP Step X X X
* These template versions belong to the strategy versions listed

below.
Strategies overview
ÄKTAbasic system Strategy name*
Basic 100 with Frac-901 B100_200
Basic 100 with Frac-950 B100F200
Basic 10 with Frac-901 B10_200
Basic 10 with Frac-950 B10F200
Basic 10 XT with Frac-901 B10X200
Basic 10 XT with Frac-950 B10XF200
*
The last three digits in the strategy name is the version
number.

________________________________________________ ÄKTAFPLC 5
5 ÄKTAFPLC
5.1 General
To utilize optional equipment in a chromatographic run, suitable

5.16 Configuration dependent delay volume in ÄKTAFPLC .
The descriptions below deal with instructions used in specific functions

5.2 Unpacking

that the ÄKTAFPLC optional equipment is installed correctly.
5.3 Pre-requisites
connected to your ÄKTAFPLC chromatography system.
• ÄKTAFPLC must be installed and fully tested before the optional

ÄKTAFPLC chromatography system.

components.
CAUTION! Make sure ÄKTAFPLC is switched off before installing

the optional components. The mains power to ÄKTAFPLC must be
UniNet-2 cables.

5 ÄKTA FPLC ________________________________________________
CAUTION! Make sure ÄKTAFPL C is switched off before installing

the optional components. The mains power to ÄKTAFPL C must be
UniNet-2 cables.
Optional equipment can be mounted on the system rack with the rack
door closed or open. Both the inside and the outside are provided with
mounting rails for quick mounting of components.
The upper part of the system rack can be raised and locked with screws
in optional positions along its vertical rods. This is practical when using
long columns.
When selecting mounting locations for the optional equipment, it is

important to choose a location which minimizes the length of the
capillaries used to connect the optional components to the rest of the
system.
Many components are attached to the system rack mounting rails with
a snap-in bracket. The bracket is supplied separately with each
component and needs to be fitted as shown below before the
component can be attached.
2. Fit the attached

the two screws
1. Loosen the
two screws
Removing components
Capillaries are connected using unions as specified in section

6.1 Connections, chromatography – standard selection.

________________________________________________ ÄKTAFPLC 5
installation. The ID codes for the different
function valves and the corresponding
instructions used in UNICORN to control them
are as follows:

valves)
Outlet fractionation 4 PV-908 OutletValve, OutletFractions (two
instructions used for controlling
different types of fractionation
functions)
Buffer selection A 5 IV-908 BufferValveA
Buffer selection B 6 IV-908 BufferValveB
Optional function 7 INV-907/IV/PV-908 Valve7

UniNet-2 data communication chain in standard configuration are

To PC
Conductivity
Optical
Unit
Analogue
out 0-1 V UniNet 1
controller board
Flow Cell
pH-GroundpH-Probe
Mains
Lamp
Voltage Frequency Power, max

100-200 V- 50-60 Hz 30 VA
Pressure
Analogue out 0-1 V
Remote
UniNet 2
UniNet 2
UniNet 1
chain
Fraction
collector
Mains output
Leakage current, max

3,5 mA
PC running UNICORN
Mains
Voltage Frequency Power, max Fuse

100-200 V- 50-60 Hz 600 VA T 6,3 AL 5 4 3 2 1
9 8 7 6
WARNING! For continued protection against

risk of fire, replace only woth fuse
of the specified type and current ratings
Termination
plug
UniNet 1
chain
*
Frac-901, Frac-950 and A-900 are connected via the UniNet-1 chain. All other optional compo-
nents are connected via the UniNet-2 chain.

5 ÄKTA FPLC ________________________________________________

To control the optional components, the user must insert UNICORN
control instructions into a method. UNICORN must also be informed
about the insertion of an optional component. This is known as
selecting the component, which makes its corresponding instructions
available. A list of the most commonly used instructions is given below.
The default values are underlined.

component
Column selection PV-908 Manual:Flowpath* ColumnPosition Position (1-8)
valve
Outlet valve PV-908 Manual:Flowpath OutletValve Position (Feed, F1-F8)
Manual:Frac* OutletFractions How Many? (0-7)
Volume (5-10000 ml)
Start at (F2-F8)
Buffer valve IV-908 Manual:Flowpath* BufferValveA Position (A1-A8)
BufferValveB Position (B1-B8)
Optional valve 7 INV-907/ Manual:Flowpath* Valve7 Position (Feed, 1-8)
IV/PV-908
IV/PV-908
IV/PV-908

HighAlarm (0-1 MPa)
LowAlarm (0-1 MPa)

Volume (0-1000 ml)
REMOTE I/O P-920 System:Settings:Special* AuxOut1 Signal (0, 1)
Manual:Pump* AuxOut1 Signal (0, 1)

View:Windows:Instruction Watch_AuxIn1-4 Test Equal (Value 0, 1)

Action (<Block Name>/
box:Watch**
READY/PAUSE/
HOLD/END_BLOCK/
END_METHOD/
CONTINUE)

________________________________________________ ÄKTAFPLC 5
component
collectors TubeChange (Tube, Drop-
Sync, WasteBetween-
Tubes)
Peak_FracParameters_900 StartSlope, EndSlope,
MinWidth

FeedTube_900 -
Peak_FracStop_900 -
TubeChange (Accumula-
tor, DropSync, Tube)
Peak_FracParameters StartSlope, EndSlope,
MinWidth
Method/Manual:Frac Fractionation Tube type (name from rack

definition)
FracSize (0-99999.999
base)
Start at (First tube, Next
tube, Next line, Next
group, Tube number)
Base (Volume, Time)
FractionationStop -
FeedTube -
Peak_Fractionation Tube type (name from rack
definition)
PeakSize (0-99999.999
base)
group, Tube number)
Base (Volume, Time)
Peak_FracStop -
Frac-950 Manual:Frac Man_Fractionation Rack (name from rack def-
inition)
Frac_order
(Serpentine_row, Row-by-
row, Serpentine-column,
Column-by-column)
Tube type (name from rack
definition)
FracSize (0-99999.999
base)
group, Tube number)
Base (Volume, Time)

5 ÄKTA FPLC ________________________________________________

component
Frac-950 Manual:Frac Man_Peak_Fractionation Rack (name from rack def-
(contin- inition)
ued) Frac_order
(Serpentine_row, Row-by-
row, Serpentine-column,
Column-by-column)
Tube type (name from rack
definition)
PeakSize (0-99999.999
base)
group, Tube number)
Base (Volume, Time)
Ignore_LastTube -
Air sensor1 A-9001 System:Settings:Monitors* AirSensor1-3 Mode ( High_Sensitivity,
SamplePump)
Manual:Alarms&Monitors* Alarm_AirSensor1-3 Mode (Enabled,

or System:Settings:Alarms* Disabled)1

box:Watch** Action (<Block Name>/
READY/PAUSE/
HOLD/END_BLOCK/
END_METHOD/
CONTINUE)
*
UNICORN module System Control
**
UNICORN module Method editor/Text instructions

________________________________________________ ÄKTAFPLC 5
Preparation for use
Carry out the following instructions to start-up your ÄKTAFPL C
optional configuration system:
1 Switch on ÄKTAFPLC with the mains switch located at the front left
on the system pump P-920.
3 Login (see Chapter 2 in Making your first run).
4 Make sure you have the correct strategy installed in your system.
A new strategy is installed according to the software installation

creating a new method or editing an existing method.
must be installed. For most columns, the standard tubing (0.50 mm
i.d., PEEK, orange) can be used. In complex configurations using
high flow rates, it may be necessary to replace the standard
0.50 mm i.d. PEEK tubing with 0.75 mm i.d. PEEK tubing.
is used, the back-pressure from the tubing might become higher than
the maximum pressure for the column and the run will stop
immediately after it is started.

5 ÄKTA FPLC ________________________________________________
General
purifications with ÄKTAFPL C . It collects up to 175 fractions in 12 mm
diameter tubes, up to 95 fractions in 10–18 mm diameter tubes, and up
to 40 fractions in 30 mm diameter tubes.

fractions and peaks. Drop synchronization eliminates sample loss
below flow rates of 3 ml/min during tube change.
Installation
On-line filter
Mixer
Column
Injection
valve (V1)
17
2 6
34 5
Pump A Pump B UV
Flow restrictor
COND Switch valve
A B W2 W1
Fraction W3
collector
Frac-901
3 Connect a tubing from the outlet of

the flow restrictor to the IN port To delivery
on the flow diversion valve. arm (NC)
4 Connect a tubing from the NC

(normally closed) port on the flow
diversion valve to the delivery arm
on Frac-901. Follow the From flow To waste
(NO)
instructions given in the restrictor (IN)
Frac-900/901 User Manual.
5 Connect a tubing from the NO (normally open) port on the flow

diversion valve to waste.

________________________________________________ ÄKTAFPLC 5
bottle.

34
• Connecting a UniNet-1 cable between Pump P-920 and

Frac-901.
• Connecting a termination plug in the other UniNet-1 connector

in Frac-901.
To PC
Conductivity
Optical
Unit
Analogue
out 0-1 V UniNet 1
controller board
Flow Cell
pH-GroundpH-Probe
Mains
Lamp

100-200 V- 50-60 Hz 30 VA
Pressure
Analogue out 0-1 V
Remote
Fraction
UniNet 2
UniNet 2
collector UniNet 1
chain
Mains output

3,5 mA
1 2
Mains

100-200 V- 50-60 Hz 600 VA T 6,3 AL 5 4 3 2 1
9 8 7 6

Termination
plug
UniNet 1
chain
8 Connect a mains cable supplied between Frac-901 and a mains

socket at the rear of ÄKTA)3/&.
Optical Analogue
Conductivity UniNet 1
Unit out 0-1 V
Flow Cell
pH-GroundpH-Probe
Mains
Lamp
To
100-200 V- 50-60 Hz 30 VA
Pressure
Analogue out 0-1 V
UPC-900
Remote
UniNet 2
Free mains
UniNet 1
Mains output
outlet for
3,5 mA
optional
equipment
1 2
Mains

100-200 V- 50-60 Hz 600 VA T 6,3 AL
To
Frac-901
From
mains
outlet

5 ÄKTA FPLC ________________________________________________
Operation
1 Start your ÄKTA)3/& optional configuration system as described in
section Preparation for use.
2 In UNICORN, go to Instruction box in the Method Editor, and

select Frac.
Check the instructions available for Frac-901 to familiarize yourself

with the functions available. Refer also to the Fraction collector
Frac-900/901 User Manual to learn more about using Frac-901.



elution.


peak.



________________________________________________ ÄKTAFPLC 5
Note: A maximum flow rate of 3 ml/min is recommended for collecting

fractions in Frac-901 when the 0.25 mm i.d. tubing kit is used in
combination with Dropsync.
One fixed volume size before the eluate fractionation:
one fixed volume size during eluate between detected peaks:
within the elution size window set by parameters:
and finally, the peak fractionation volume:
• Peak_Frac_Size
within the peak size window set by parameters:
The peak slope characteristics are controlled with the variables:
Select a method template with “FP” in the middle of the template name
and fill in the fractionation parameters as indicated on the variables
page:

5 ÄKTA FPLC ________________________________________________
General
purifications with ÄKTAFPL C . Up to
392 fractions can be collected in time
or volume base, as fixed volume and/
or peak fractionations.
The fractionation order can be

by-column.
Six standard sizes of tube racks for

tubes, and 96-well microplates are
available.
Sample loss during tube change can

be eliminated by using drop synchronization, or an automatically
controlled accumulator reservoir.
Installation

go to step 2.
4 Disconnect the capillary from the Flow restrictor output in

ÄKTAFPLC .
6 Put Frac-950 on the workbench to the right of ÄKTAFPL C .

________________________________________________ ÄKTAFPLC 5
On-line filter
Mixer
Column
Injection
valve (V1)
17
2 6
34 5
Pump A Pump B UV
Flow restrictor
COND
Fraction
A B W2 W1 collector
Frac-950 W3

UniNet-2 cables.
7 Locate the capillary kit included with Frac-950 and decide if you
want to include the accumulator in Frac-950 or not. According to
your choice, connect a capillary from the flow restrictor output in
ÄKTAFPL C to Frac-950 as described in the Frac-950 User Manual.
8 Connect the supplied waste tubing to the waste outlet on Frac-950

9 Connect the UniNet-1 cable and the termination plug to Frac-950.


5 ÄKTA FPLC ________________________________________________
Operation
1 Start your ÄKTAFPLC optional configuration system as described in
Note: If you have upgraded from Frac-901, your new Frac-950 is used
in method templates in very much the same way as the previously used
Frac-901. However, additional functions in the form of new
instructions and parameters are now available in the instructions list,
and all instructions/parameters valid for Frac-901 are removed.
2 In UNICORN, go to System Control:Manual and select Frac.
3 Check the instructions available for Frac-950 to familiarize

yourself with the functions available in Frac-950. Refer also to the
Fraction Collector Frac-950 User Manual to learn more about
using Frac-950.
Note: A maximum flow rate of 3 ml/min is recommended for collecting

fractions in Frac-950 when the 0.25 mm i.d. tubing kit is used in
combination with Dropsync.



elution.


peak.

________________________________________________ ÄKTAFPLC 5




position.
• Ignore last tube and Continue are ordered the fraction collector

When the last position in the rack is reached, an alarm is generated and
the fraction collector goes to position waste. If:

position.

Note: This instruction resets the tube type that has run out, but not
the other tube type used.

5 ÄKTA FPLC ________________________________________________

Start setting up the rack characteristics on Run Setup page Frac-950:
30 mm tubes
18 mm tubes
30 mm tubes
Select Rack type, Fractionation order, Last tube and mark the last tube
position in which you intend to put a tube by clicking on the
corresponding coloured spot in the figure.

One fixed volume size before the eluate fractionation:


within the window set by parameters:

________________________________________________ ÄKTAFPLC 5

within the window set by Start_Frac_at and End_Frac_at
The peak slope characteristics are controlled with the variables:
Select a method template with “FP” in the middle of the template name

5 ÄKTA FPLC ________________________________________________
Installation
Column
Selection
On-line filter Valve
(V2)
Mixer
18
2 7
3 6
4 5
Injection
Valve (V1)
17
2 6
34 5
Pump A Pump B
Restrictor
Fraction
A B W2 W1 Collector
W3
18
2 7
3 6
4 5 UV
COND
Column
Selection
Valve (V3)

UniNet-2 cables.
2 Mount the brackets and attach the two valves to the system rack.
3 Cut and mount the new 0.50 mm i.d. capillaries. Mount one
capillary between ports 1 on Column valve 2 and Column valve 3.
This is the column bypass position used as default position for the
valves in UNICORN.



________________________________________________ ÄKTAFPLC 5
Operation
1 Start your ÄKTAFPL C optional configuration system as described in
2 The column selection valves are automatically recognized by


from port 4 of column valve 2.
• Connect a capillary between port 4 of column valve 2 and port

4 of column valve 3 and check that water is coming out from the
centre port of column valve 3.

scheme and V2_ColPos and V3_ColPos in the run data window. To use
the column selection valves in a method, proceed as follows:


column selection instruction, e.g. Start_Conditions_zz. Highlight
the instruction below which you want to insert the valve instruction.

5 ÄKTA FPLC ________________________________________________

3 Click on the Flowpath button.



8 Save the method.

________________________________________________ ÄKTAFPLC 5
5.8 Outlet valve
Installation
On-line filter
Mixer
Column
Injection
Valve (V1)
17
2 6
34 5
Pump A Pump B UV
COND
Restrictor
A B W2 W1 Outlet
Valve (V4)
1 8
2 7
3 6
45
Fraction
Collector W3 F2 F3 F4 F5 F6 F7 F8

UniNet-2 cables.
1 Mark the PV-908 valve with Outlet Valve 4. Check that the ID code
switch is set to 4.
2 Mount the bracket and attach the valve to the system rack.
3 Cut and mount the new 0.50 mm i.d. capillaries.



5 ÄKTA FPLC ________________________________________________
Operation
2 The outlet valve is automatically recognized by UNICORN at

system start-up. Check that the valve added is indicated in the flow
scheme in UNICORN.
3 Verify that the outlet valve is functioning properly by issuing

• Select instruction OutletValve.
• Set the outlet valve to position 4.

from port 4 of the Outlet valve.
Using the outlet valve for fractionation

Two different fractionation instructions are available to use with the
outlet valve:
• OutletValve
This instruction parameter turns the outlet valve to one of the eight
positions available. This makes the instruction suitable for
controlling flow-through fractionation before elution starts.
• OutletFractions
This instruction has three parameters, How_many?, Volume and
Start_at. This makes the instruction suitable for controlling large
volume fractionation (min. 5 ml) during elution. Both instructions
are independent of using Fraction collector Frac-901 or Frac-950.
Note: Installing an outlet valve in the flow path affects the system delay
ÄKTAFPLC for instructions on how to re-calculate and change the delay
volume.

________________________________________________ ÄKTAFPLC 5
Collecting flow-through fractions with the outlet valve
This instruction is designated OutletValve in the flow scheme and
V4_Outlet in the run data window. To use the instruction in a method,
proceed as follows:


Outlet valve instruction, e.g. Flowthrough_Fractionation. Highlight
the instruction below which you want to insert the flowthrough
fractionation instruction.
Delete the
FractionCollector
instruction.

5 From the Instructions list, select OutletValve. The valve instruction

parameter Position allows you to change all eight positions of the
outlet valve.

5 ÄKTA FPLC ________________________________________________
6 Define a variable name for the outlet valve instruction parameter,

e.g. OutletValve_Pos. This variable allows you to control the
position of the outlet valve to the position you want.
8 Insert an instruction to reset the valve to the desired position after

flowthrough fractionation is terminated.

variable OutletValve_Pos to the desired position.
10 Save the method.

________________________________________________ ÄKTAFPLC 5
Collecting large fractions with the outlet valve
This instruction is designated OutletFractions. To use the instruction in
a method, proceed as follows:


fractionation instruction, e.g. Eluate_Fractionation. Highlight the
instruction below which you want to insert the fractionation
instruction.
Delete the old

FractionCollector
instruction and add the
new OutletFractions
instruction.
Click here to define

variable names for the
4 Click on the Frac button.
5 From the Instructions list, select OutletFractions. The instruction

parameters allow you to select manually how many fractions to
collect, the volume of each fraction and which position to start
from.
6 Click the upper Var button to define a variable name for how many
fractions to collect, e.g. No_of_Fracs. This variable allows you to
control the number of fractions to collect. When the number of
fractions have been collected, the outlet valve is set to start position.

5 ÄKTA FPLC ________________________________________________
Note: The instruction parameter How_many? can also be used to

terminate outlet fractionation by setting it to 0. The outlet valve is then
reset to start position.
8 Click the middle Var button to define a variable name for the size
(volume) of the fractions to collect, e.g. FracSize_Vol. This variable
allows you to control the volume of the fractions to collect.
10 Click the lower Var button to define a variable name for the port to
start from, e.g. Start_at_Pos. This variable allows you to control
the position of the outlet valve from which you start collecting
fractions from.

variables created above to the desired values.
13 Save the method.

________________________________________________ ÄKTAFPLC 5
Installation
On-line filter
Mixer
Column
Injection
Valve (V1)
17
2 6
34 5
Pump A Pump B UV
COND
Restrictor
Fraction
Buffer Buffer Collector
Valve A (V5) Valve B (V6) W2 W1
2
18 18 W3
7 2 7
3 6 3 6
45 45
A1 A2 A3 A4 A5 A6 A7 A8 B1 B2 B3 B4 B5 B6 B7 B8

UniNet-2 cables.
1 Mark the IV-908 valves with Buffer Valve A and Buffer Valve B.
Check that the ID code switches are set to 5 and 6 respectively.
2 Mount the brackets and attach the valves to the system rack.
3 Cut and mount the required teflon tubings (i.d. 1.6 mm).



5 ÄKTA FPLC ________________________________________________
Operation
2 The buffer selection valves are automatically recognized by

indicated in the flow scheme in UNICORN.
3 Verify that Buffer Valve A and Buffer Valve B are functioning

properly by issuing manual commands from UNICORN as
follows:
• Select instruction BufferValveA.
• Set Buffer Valve A to position A4.

from port 4 of the buffer valve.
• If applicable, repeat these steps for Buffer Valve B.

________________________________________________ ÄKTAFPLC 5
The buffer valve instructions are designated BufferValveA and
BufferValve in the flow scheme. In the run data window, they are
designated V5_BufferA and V6_BufferB. To use the valves in a method,
proceed as follows:



3 Click on the Flowpath radio button.
4 From the Instructions list, select BufferValveA or BufferValveB. The

valve instruction parameter Position allows you to change all eight
positions of the buffer valves individually.

5 ÄKTA FPLC ________________________________________________

parameters, e.g. BufferValveA_Pos/BufferValveB_Pos. These
variables allow you to control the position of the buffer valves
individually to the position you want.

variable BufferValveA_Pos/BufferValveB_Pos to the desired
positions.
8 Save the method.

________________________________________________ ÄKTAFPLC 5
Installation

UniNet-2 cables.
1 Mark the optional function valve with Valve 7, Valve 8 or Valve 9

as required. Check that the ID code switch is set to 7, 8 or 9
respectively.
2 Mount the bracket and attach the valve to the system rack. Choose
the best location to facilitate the routing of the tubing that connects
the valve to the main system flow path.
3 Cut and mount the necessary capillaries (PEEK, 0.50 mm i.d.).

Restrict the length of the capillaries as much as practically possible
to reduce band broadening effects.

plug connected to the injection valve V1.

The following example shows how an optional INV-907 is connected

when used as a reversed flow valve.
1 1
2 7 Pump 7 Pump
2
To monitor To monitor 3
3 6 6
4 5 4 5
Stop plug Stop plug

5 ÄKTA FPLC ________________________________________________
Operation
2 A connected valve is automatically recognized by UNICORN at

system start-up. Check that the added valve is indicated in Flow
scheme in UNICORN.

• Set the valve to position 3 (Waste for INV-907).
• Make sure that the inlet capillary to the general function valve is
connected to the centre port on an IV/PV-908 valve, or port 7 on
an INV-907 valve.

from port 3 of IV/PV-908, or port 5 for INV-907.

General function valve instructions are designated Valve7, Valve8 and
Valve9 in the flow scheme and run data windows. To use a valve in a
method, proceed as follows:


________________________________________________ ÄKTAFPLC 5
general function valve instruction, e.g. Start_Conditions_zz.
Highlight the instruction below which you want to insert the valve
instruction.

4 From the Instructions list, select the valve you intend to use
(Valve7, Valve8 or Valve9). The valve instruction parameters allow
you to change the position of the valves. For an INV-907 injection
valve, only three positions can be selected.

5 ÄKTA FPLC ________________________________________________

Valvex_pos, where x=7, 8 or 9 depending on the ID code for the
valve used. This variable allows you to turn the position of the
valve to the port you want.

8 Save the method.

________________________________________________ ÄKTAFPLC 5
General
CAUTION ! Never use 0.25 mm i.d. tubing (blue) when the pH

flow cell is mounted in the system. The electrode may rupture due to
high back-pressure.
The pH electrode used in ÄKTAFPLC is mounted in a separate pH flow

cell housing the pH electrode itself, as well as a dummy pH electrode.
The dummy pH electrode replaces the pH electrode whenever the pH
electrode is to be removed from the system, e.g. for storage.
Note: Always use the 0.50 mm tubing kit when the pH flow cell is
mounted.

refilled, an internal electrolyte bridge of 4 M KCl saturated with
Ag/AgCl, an outer electrolyte bridge of 2 M KNO3 , an annular ceramic
Installation
1 Remove the flow restrictor and the male/male union from the
conductivity flow cell.
2 Install the pH flow cell and pH electrode as described in the

Monitor UPC-900 User Manual. The pH flow cell should be
mounted directly to the right of the Optical unit.
On-line filter
Mixer
Column
Injection
valve (V1)
17
2 6
34 5
Pump A Pump B UV
Flow restrictor
COND
Fraction
A B W2 W1 collector
pH Flow cell W3
New capillary of Male/male union

90 mm

5 ÄKTA FPLC ________________________________________________
3 Cut a new capillary of 90 mm (0.50 mm i.d.) and connect the outlet

(the screw head end) from the conductivity flow cell to the inlet on
the pH flow cell (lower connection marked IN).
4 Connect the outlet from the pH flow cell (marked OUT) to the inlet
on the flow restrictor (marked IN) using the male/male union.
5 Connect the pH electrode BNC connector to the pH-Probe

connector on the rear of Monitor UPC-900. Should interference
from static fields occur, connect the pH flow cell to the pH-Ground
connection as well using a standard laboratory 4 mm “banana
plug” cable.

Operation
Note: Installing a pH flow cell in the flow path affects the system delay
ÄKTAFPLC for instructions on how to re-calculate and change the delay
volume.
Note: With peak volumes less than 1 ml, resolution decreases due to
the volume in the pH flow cell.
2 Calibrate the pH electrode and set up the additional monitor

functions desired for pH measurement. These procedures are
described in the Monitor UPC-900 User Manual and in the
UNICORN User Manual.

________________________________________________ ÄKTAFPLC 5
5.12 Sample pump
General
laboratory pump for use as a sample
pump to apply samples directly on P-950
the column and/or to fill sample

loops, including Superloop. Pumping
action is provided by a membrane
driven by a stepper motor. The pump
is a low pulsation pump and features
a motor speed independent of
temperature and load to give
accurate and reproducible flow rates
up to 50 ml/min.
A Sample valve IV/PV-908 can be

used in combination with the sample pump. It is a motorized 8-way
valve used for automatic sample application of up to 8 samples.
There are three ways of using Pump P-950 in the ÄKTAFPLC system:

procedures to remove air from the flow path inside the sample pump,
as described in the following.


5 ÄKTA FPLC ________________________________________________

by the sample pump.
Note: When filling the sample loop, the resistance in the flowpath may
cause cavitation if the flow rate exceeds 0.5 ml/min.
Installation
1 Assemble Pump P-950 according to the instructions given in the
2 Use the supplied screws to attach the mounting bracket to the upper
rear of the pump housing.
3 Attach the sample pump to the system rack.

5 Use the attached UniNet-2 cable to connect the sample pump to the
UniNet-2 communication link. The sample pump can be connected
plug.

Outlet connection


Inlet connection

________________________________________________ ÄKTAFPLC 5
On-line filter
Mixer
Column
Injection
Valve (V1) 3
17
2 6
34 5
Pump A Pump B UV
COND
Sample
Pump
Restrictor
4
Fraction
2 Collector
A B 1 W2 W1
W3
5
S1

1
port*
2
3
4
5
port 1-8
*
Only if you have a sample selection valve in your configuration.

5 ÄKTA FPLC ________________________________________________

Preparation for use in section 5.4 General guidelines.
2 Check that the sample pump is indicated in the flow scheme

window.

section Installation on page 332.




1 Make sure the system is prepared according to section Preparing
for operation.

If the running conditions are changed, calibrate the sample pump
according to the instructions given in the Pump P-950 User Manual.

the sample pump.

________________________________________________ ÄKTAFPLC 5
Preparing a method for filling a sample loop
shown below.


3 Select Block:New.
manually.

*
*
*
added manually.

to LOAD.

5 ÄKTA FPLC ________________________________________________

sample pump.


breakpoint variable
loop.
12 Save the method.


________________________________________________ ÄKTAFPLC 5
This page is intentionally blank.

5 ÄKTA FPLC ________________________________________________

of sample are to be loaded. The sample is pushed onto the column.
Note: Due to the back-pressure in the flowpath, the flow rate should
not exceed 20 ml/min when loading sample directly onto the column.
Installation

plug.

Pump P-950
Outlet connection

Inlet connection

________________________________________________ ÄKTAFPLC 5
On-line filter
Mixer
Column
2 Injection
Valve (V1)
17
2 6
Pump A Pump B
34 5 3 UV
COND
4
Restrictor
Sample Fraction
Pump Collector
A B W2 W1
1
6 5 W3
S1

1
(Sample inlet tubing) Tubing connector for 1/8" o.d. tubing, 18-1121-17 port*
2
connector
3
4
5
6

replaced with PEEK i.d. 0.75 mm tubing. The sample inlet tubing may be adjusted to a suitable
length.

5 ÄKTA FPLC ________________________________________________


window.





If a sample selection valve is used in your configuration, go directly to
the next section. Otherwise, proceed as follows:


________________________________________________ ÄKTAFPLC 5




sample container.

when using a sample selection valve
If a sample selection valve is used (assumed to be connected as Valve8
in this example), proceed as follows to remove air from the sample
flowpath:




5 ÄKTA FPLC ________________________________________________

below.


9 Save the method and run it. It takes approximately 12 minutes to

complete.
Preparing a method for direct loading onto column

method as follows:


3 Select Block:New.

________________________________________________ ÄKTAFPLC 5
Blue square
Set Base Volume
and



flow rate to the viscosity of the sample.

5 ÄKTA FPLC ________________________________________________


the column.

________________________________________________ ÄKTAFPLC 5
This page is intentionally left blank

5 ÄKTA FPLC ________________________________________________
Loading a Superloop

ml
the seal moves towards either end piece of the glass tube.
10
5
0
Installation

Note: It is not possible to operate the Injection valve to position Waste

when the sample pump is running (the sample pump will stop).
plug.

Pump P-950
Outlet connection

Inlet connection

________________________________________________ ÄKTAFPLC 5
On-line filter
Mixer
Column
Injection
1 Valve (V1)
17
2 6
34 5
Pump A Pump B UV
3 2 COND
Restrictor
Buffer
Sample Fraction
Pump Collector
A B W2 W1
4 Sample
W3
S1
Superloop

1
connector
2
3
4

5 ÄKTA FPLC ________________________________________________


window.







ethanol container.

________________________________________________ ÄKTAFPLC 5
distilled water.

If the running conditions are changed, calibrate the sample pump
according to the instructions given in the Pump P-950 User Manual.


sample container.

method as follows:


4 Select Block:New.

5 ÄKTA FPLC ________________________________________________
Blue square
Set Base Volume
and MethodBase
SamplePump

breakpoint variable

to LOAD.

sample pump.

system pump.

16 Save the method.

________________________________________________ ÄKTAFPLC 5
5.13 Air sensors
General
Air sensors (AS1, AS2 and AS3) can be used in the ÄKTAFPLC flow
On-line filter
Mixer
Column
Injection
Valve (V1)
17
2 6
34 5
Pump A Pump B UV
COND
Restrictor
Air cell
AS1 Fraction
Buffer B W2 W1 Collector
A W3
One air cell located in the inlet flow of pump P-920 for checking the buffer supply.
On-line filter
Mixer
Column
Injection
Valve (V1)
17
2 6
34 5
Pump A Pump B UV
COND
Restrictor
Sample
Fraction
Pump Collector
A B W2 W1
Purge valve W3
Air cell
AS2
Sample container
One air cell located in the sample inlet flow when applying sample
directly onto the column with a sample pump.
Air cells can be fitted in other locations in the ÄKTAFPLC main flow

5 ÄKTA FPLC ________________________________________________
Installation

UniNet-2 cables.

1 Make sure that the ÄKTAFPL C chromatography system is switched
off.

ON
1 2 3 12

Release
Fasten


________________________________________________ ÄKTAFPLC 5
Using air cell Air-925 (AS1) in the system pump inlet flow path
To system pump inlet
1 Remove the A capillary from the system pump inlet A.
2 Cut new Teflon capillaries, i.d. 1.6 mm, as shown in the figure and
attach the purge valve and the air cell using the connectors and
ferrules supplied.
Purge
L1=920 mm
valve
Note: The length of L1 is dependent on which flow, sensitivity and
A-925 Note: When running gradients, the air sensor detects smaller air
Air cell bubbles than expected. During a gradient, the flow rate through the
Inlet filter 3 Route the signal cable from the air cell to the Air cell 1 connector
Using air cell Air-925 (AS2) in the sample pump inlet flow path
1 Remove the sample inlet capillary from the Sample pump P-950
To sample pump inlet
Outlet connection
2 Cut the sample inlet capillary and attach the
purge valve and the air cell using the
connectors and ferrules supplied.
Purge
valve 3 Route the signal cable from the air cell to the
Air cell 2 connector on the Air-900 control
box and plug it in.
Note: The length of L1 is dependent on

A-925 which flow, sensitivity and type of Inlet connection
Air cell
instruction (alarm or watch) is used in
UNICORN. The optimal length of L1 must be tested in the user
configuration.
Note: Make sure that no air bubbles are trapped in the sample
Inlet filter or pump flow path before using the pump (refer to section 5.12
Sample valve Sample pump).

5 ÄKTA FPLC ________________________________________________
Operation
2 To set the air sensor sensitivity, go to System Control and select

System:Settings.

in UNICORN.

________________________________________________ ÄKTAFPLC 5
filter of the A tubing. Purge the air cell using a syringe and the purge
valve mounted on the inlet capillary. The purge valve is opened to
the syringe by turning the syringe connector a 1/2 turn counter-
clockwise.
the flow diagram is indicated with a white colour.

3 Fill a container with distilled water and submerge the inlet tubing.
the flow diagram is indicated with a white colour.


5 ÄKTA FPLC ________________________________________________

connected the air cell to, either AS1, AS2 or AS3 should be shown
in the flow scheme.
2 Prepare ÄKTAFPL C to enable distilled water to be pumped through

the optionally located air cell.

the air cell.
5 Set Flow:Flowrate or SampleFlow:Flowrate to 2 ml/min and click

Execute.
flow diagram is indicated with a white colour.

to suit your needs.

used. Instructions for handling the AS1, AS2 and AS3 air cells must be
inserted manually in a method.
Alarm_AirSensor1–3

________________________________________________ ÄKTAFPLC 5
Watch_AirSensor1–3

To use air cells in a method, you should insert instructions to modify an
Example 1


Start_Conditions.

instruction.

radio button.

radio button

5 ÄKTA FPLC ________________________________________________

6 Click on Insert.
7 Save the method.
Example 2


sensor.

*)
CONTINUE. variable

calculation.

________________________________________________ ÄKTAFPLC 5
3 Select Block:New.



clicking on Delete.

10 Save the method.

5 ÄKTA FPLC ________________________________________________
5.14 Auxiliary equipment controlled via Pump P-920 REMOTE

connector
Installation

REMOTE connector the optional components.
on rear of P-920
Pressure
Analogue out 0-1 V
Remote
UniNet 2
UniNet 1

Mains output
3,5 mA
Inputs and outputs are +5 V TTL compatible. Unused inputs can be left
Mains
open and are read as logical “1”.
100-200 V- 50-60 Hz 600 VA T 6,3 AL
Pin Signal Function

1-4 AuxIn 1-4 Active state can be selected to 0 or 1 in UNICORN.
5 4 3 2 1 Min. duration=50 ms. Max. frequency=1 Hz.
9 8 7 6 5 0V Signal ground
6-9 AuxOut 1-4 Active state can be selected to 0 or 1 in UNICORN.

P-920
Connector

________________________________________________ ÄKTAFPLC 5
Operation
1 Start your ÄKTAFPL C optional configuration system as described in

AuxOut1-4
To set up and use the output signals, proceed as follows:
AuxOut1.

0 or 1, corresponds to inactive mode.


5 ÄKTA FPLC ________________________________________________
Watch_AuxIn1-4
selected actions can be, for example, end block.

The status for signals AuxIn1 and AuxOut1 can be stored as curves in
the chromatogram. This is selected in System:Settings by setting
Curves:Store ON.

________________________________________________ ÄKTAFPLC 5
5.15 Superloop


(1–150 ml) onto the column. Superloop must not be used with
columns generating a column pressure higher than that stated in the
table below. If the column pressure is higher, the flow must be reduced
in order not to exceed the pressure limit. Set the pressure limit in the
ml 10 method with the variable Pressure_limit.
5

10 ml 4 MPa 18-1113-81
50 ml 4 MPa 18-1113-82
150 ml 2 MPa 18-1023-85


Preparation
1 Connect the supplied

Pos. 1 LOAD
Column
2
Sample 3 6
5
Super

separate Superloop
Instruction).

5 ÄKTA FPLC ________________________________________________

as possible.
Filling Superloop

movable seal has reached
the bottom of Superloop.

35
25
20

Empty_loop_with.

________________________________________________ ÄKTAFPLC 5
5.16 Configuration dependent delay volume in ÄKTAFPLC
altered when the standard configuration after the UV cell is changed to
The delay volume value for the standard configuration including

Frac-901 is 375 µl.
The delay volume value for the standard configuration including

Frac-950 is 197 µl (without accumulator with tubing).

standard configuration delay volume value.
The example below shows a calculation for including the pH flow cell.
Capillary volume + cell volume = 90/100 x 19.6 + 100 µl=128 µl.
(The length of the new i.d. 0.5 capillary is 90 mm, the volume of
100 mm tubing is 19.6 µl, and the volume of the pH flow cell 100 µl.)
375 + 128 = 503 µl (including Frac-901), and



Valve PV-908 7
Valve IV-908 27
Valve INV-907 9
Valve FV-903 113
UV flow cell 2 mm 15
UV flow cell 5 mm 10
pH flow cell 100
Finger-tight male/male (Frac-950) 7

5 ÄKTA FPLC ________________________________________________
5.17 Strategy and template overview in ÄKTAFPLC
Templates overview
Type of ÄKTAFPLC system
Template name*
FPLC
Chromatofocusing X
Installation Test X
Manual Injection FP Isocratic X
Manual Injection FP Segmented X
Manual Injection FP Step X
*
These template versions belong to the strategy versions listed
below.
Strategies overview
ÄKTAFPLC system Strategy name*
FPLC with Frac-901 FPLC_200
FPLC with Frac-950 FPLCF200
*
The last three digits in the strategy name are the version number.

________________________________ Accessories and consumables 6
6 Accessories and consumables
6.1 Connections, chromatography – standard selection
Connector Ferrule Tubing (mm) Material/ Pressure

Description and design Connections limit
cat. no. cat. no. o.d i.d. colour (bar)
Fingertight M6 male/female 18-1112-57 M6/ 0.8 PEEK 100

UNF 10-32 black
Fingertight connector 18-1112-55 UNF 10-32 1.6 PEEK 250

1/16” male (1/16") black
Fingertight male/M6 female 18-1112-58 UNF 10-32 / 0.8 PEEK 100

M6 black
Tubing connector for 18-1112-49 18-1112-48 UNF 5/16-24 4.8 PEEK 70

3/16” o.d. tubing (3/16”) black

1/8” o.d. tubing (1/8") black

1/16” o.d. tubing (1/16") black
Union 5/16” female/ 18-1142-08 UNF 5/16-32 / 1.8 PEEK 70

HPLC male UNF 10-32 black

6 Accessories and consumables _______________________________
6.2 Optional components and consumables

Item Quant./pack A/C* Code no.
Fraction Collector Frac-950
Fraction Collector Frac-950, complete 1 A 18-6083-00
with rack A (18 mm + 30 mm tubes)
Rack A, complete with bowl, tube support 1 A 18-6083-11
and tube holder
Rack B, complete with bowl, tube support 1 A 18-6083-12
and tube holder
Rack C, complete with bowl, tube support 1 A 18-6083-13
and tube holder
Rack D, complete with bowl, tube support 1 A 18-6083-14
and tube holder
Rack E, complete with tube holder 1 A 18-6083-15
Rack F, complete with tube holder 1 A 18-6083-16
Rack G, complete with tube holder 1 A 18-6083-17
Fraction Collector Frac-901

Fraction Collector Frac-901, complete
with 18 mm tube rack 1 A 18-1118-97
Tube racks, complete with bowl,
tube support, holder and guide:
12 mm 1 A 19-8684-03
18 mm 1 A 18-3050-03
30 mm 1 A 18-1124-67
Tube support 1 A 18-3054-02
Tube holder and guide:
12 mm 1 A 19-7242-02
18 mm 1 A 19-8689-02
30 mm 1 A 18-1124-68
Eppendorf tube holder for 12 mm rack 100 A 18-8522-01
Flow Diversion Valve, FV-903 1 A 18-1114-50
incl. mounting bracket
Valve INV-907
Valve INV-907 including
one UniNet cable
(fill port, needle and syringe holder
are not included 1 A 18-1108-40
Valves IV-908, PV-908
Valve IV-908 including
one UniNet cable 1 A 18-1108-42
Valve PV-908 including
one UniNet cable 1 A 18-1108-41
*) A = accessory, C = consumable

________________________________ Accessories and consumables 6
pH measurement
pH electrode, round tip, including 1 C 18-1134-84
flow cell and holder
pH electrode, round tip 1 C 18-1111-26
pH flow cell, round tip, including 1 A 18-1112-92
dummy electrode
Dummy electrode, round tip 1 A 18-1111-03
Pump P-950
Pump P-950 including 1 A 18-6083-01
one UniNet cable, 0.7 m
ÄKTAexplorer 10S Upgrade kit, including 1 A 18-1147-02
Pump P-950, Mounting kit P-950,
Valve PV-908,
Sample holder SH-900
two UniNet cables, 0.3 m
ÄKTAexplorer 10S Tubing kit
Air sensor Air-900
Air-900 control box including
pump tubing and one UniNet 2 cable 1 A 18-1121-22
Air cell
Air-912 (1.2 mm) 1 A 18-1121-23
Air-925 (2.5 mm) 1 A 18-1121-24
Purge valve 1 A 18-1126-33
Superloop 10 ml, 50 ml
Superloop 10 ml, complete 1 A 18-1113-81
Tubing kit for Superloop (10 ml) 1 A 18-1113-83
Tubing kit for Superloop (50 ml) 1 A 18-1113-84
Superloop 150 ml

6 Accessories and consumables _______________________________
Tubing
PEEK, blue, i.d. 0.25 mm, o.d. 1/16” 2m 18-1120-95
PEEK, orange, i.d. 0.50 mm, o.d. 1/16” 2m 18-1113-68
PEEK, green, i.d. 0.75 mm, o.d. 1/16” 2m 18-1112-53
PEEK, brown, i.d. 1.0 mm, o.d. 1/16” 2m 18-1115-83
Teflon, i.d. 1.6 mm, o.d. 1/8” 3m 18-1121-16
Teflon, i.d. 2.9 mm, o.d. 3/16” 3m 18-1112-47
Tefzel, i.d. 0.25 mm, o.d. 1/16” 2m 18-1121-36
Tefzel, i.d. 0.50 mm, o.d. 1/16” 2m 18-1120-96
Tefzel, i.d. 0.75 mm, o.d. 1/16” 2m 18-1119-74
Miscellaneous
Extra rack panel, ÄKTAFPLC 1 A 18-1128-64
Communication cable P-920–P-50, 1 A 18-1129-60
*) A = accessory, C = consumable
October 2000

Short instructions
The following short instructions are checklists for
users who are fully familiar with the safety
precautions and operating instructions described in
this manual. The instructions assume that optional
equipment is installed according to the installation
instructions.
Converting to optional configurations Reverting to standard configuration

1 Wash the ÄKTAdesign flow path with
1 Flush ÄKTAdesign with distilled water.
distilled water.
2 Switch off ÄKTAdesign.
2 Switch off ÄKTAdesign. 3 Disconnect equipment to be removed from the
3 Prepare the component(s). UniNet chain and reconnect the UniNet chain for
4 Attach the components to ÄKTAdesign. the standard configuration.
5 Prepare the necessary capillaries 4 Remove capillaries and reconnect the system
and connectors. flow path to the standard configuration.
6 Mount the capillaries and connectors. 5 Check all connections.
7 Connect the components electrically. 6 Start-up ÄKTAdesign.
8 Check all connections. 7 Select a method supporting the
9 Start-up ÄKTAdesign. standard configuration.
10 Test the function of the optional equipment 8 Set up parameters to suit your needs.
manually. 9 Perform a test run with distilled water to
11 Plan the software instructions to be added to verify the standard configuration in the selected
a method, or select a ready made method. method.
12 Insert the instructions in their proper positions
in a new or modified method.
13 Set up parameters to suit your needs.
14 Perform a test run with distilled water to
TC information ab, Uppsala. Printed in Sweden by T. K. i Uppsala AB

verify the optional configuration in the new
or modified method.
15 Correct any faults and store the method.

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Optional Configurations

Amersham Pharmacia Biotech Europe GmbH

© Amersham Pharmacia Biotech AB 2000

ÄKTAdesign Optional Configurations

6 Accessories and consumables

Short instructions on back page

ÄKTAdesign Optional Configurations

When optional equipment is connected to a standard chromatographic

Optional configurations are selected, installed and put into operation

Optional configurations are monitored and controlled via methods run

Optional configurations are shown in the flow scheme displayed in the

ÄKTAdesign Optional Configurations

Generally speaking, there are two types of optional configurations.

Components used in completely user-defined configurations are

Components used in dedicated configurations are referred to in plain

ÄKTAdesign Optional Configurations

For specific information regarding function, maintenance, trouble-

This manual is divided into four main sections reflecting the

• ÄKTAexplorer (Main Section 2 ÄKTAexplorer)

• ÄKTApurifier (Main Section 3 ÄKTApurifier)

• ÄKTAbasic (Main Section 4 ÄKTAbasic

• ÄKTAFPLC ™ (Main Section 5 ÄKTAfplc)

ÄKTAdesign Optional Configurations

1.3 Summary of optional configurations

• Connecting Fraction collector Frac-901.

• Connecting Fraction collector Frac-950.

• Connecting Autosampler A-900.

• Connecting a flow direction valve.

• Connecting a sample selection valve.

• Connecting up to 2 motorized multi-port optional function valves.

• Connecting on-line pH measurement.

• Sample pump P-950.

• Connecting up to three air sensors.

• Connecting external equipment using digital input/output signals

Fraction collector Frac-901

Fraction collector Frac-950

ÄKTAdesign Optional Configurations

• Automated multiple sample injections from a few microlitres to

• Quantification of amount, concentration and molecular size

• External standard quantification .

• Internal standard quantification.

• Molecule size calculations.

Quantification uses peak data from standards to produce

Note: Quantification requires additional software not included in

• One PV-908 valve can be connected as a sample selection valve.

• Two optional valves, freely selected from INV-907, a seven-port,

ÄKTAdesign Optional Configurations

The sample pump P-950 is commonly used in combination with a

Auxiliary equipment via Pump P-900 REMOTE connection

Examples of use include:

• External alarm input/output.

• Start/stop of external equipment.

ÄKTAdesign Optional Configurations

• Connecting Fraction Collector Frac-901.

• Connecting Fraction Collector Frac-950.

• Connecting Autosampler A-900.

• Connecting column selection valves.

• Connecting a buffer selection valve.

• Connecting a flow direction valve to control flow direction through

• Connecting a sample selection valve.

• Connecting up to 2 motorized multi-port optional function valves.

• Connecting on-line pH measurement.

• Connecting Sample pump P-950.

• Connecting up to three air sensors.