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FPLC_Optional_Config

1. purge valve and the air cell using the connectors and ferrules supplied Purge E na Note Thelength of L1 is dependent on which flow sensitivity valve 5 and type of instruction alarm or watch is used in UNICORN Theoptimal length of L 1 must betested in the user configuration 3 Routethesignal cablefrom theair cell to the Air cell 1 connector on edil the Air 900 control box and plug it in L2 300 mm Inlet filter EE KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 89 E Air sensors A 132 4 Air sensor Air 912N AirSensorP960 in the sample inlet flowpath Air sensor Air 912N kit contains the necessary items required for this application Air sensor Air 912N can only be connected to Pump P 960 To sample pump inlet 1 Remove the sample inlet capillary Pump P 960 betw een the sample valve or sample l and the sample pump inlet connector Outlet connection 2 Cutthei d 1 0 mm tubing supplied and 25 cm connect to Air 912N using the fingertight connectors and union supplied 3 Connect the air sensor cable to the Air 912N socket underneath Pump P 960 Air sensor socket Inlet connection 25 cm Inlet filter or sample valve Note Make surethat no air bubbles are trapped in the sample pump flow path before using the pump r
2. ssss G general function valve installation reer es preparing for operation using in a run ssssseeenm l ID code MalVGS art aeria a EIS a ere N injection flushed injection methods KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA A a aa aa aE 117 E 118 A001 uas utu mctu met d cete censo frac 33 ADO TELE 53 injection partial alan ree ia ere pend 42 injection pICK Up 2 eerie ei temo obi iens 42 53 installation general guidelines sssseee nnn 18 Installing Pump P 960 ssssssseeeeeeeneememneennnnnnnnn nnns 68 installing the sample valve sssssseemmmee en 68 INV 907 ACCESSOS aun raei 112 IV 908 acGessorleo arancel ea 112 TeatUIES ES 13 M main fuses A 0900 su ie a trn degeret ter edere ada wn ara dina 37 A 005 aaa 49 Method Wizard overview sssssssee eee 107 mountinig railS retten te cere retener reiten 18 0 overview components instruction sssssssssseeeeeeem enne 20 optional configurations sssssssseeeeeeeneenns 11 P P 900 REMOTE connection een ener 14 P 960 ACGOSSOMES xu detected pepe re i e ur eren nne eaa de shane 112 description 1a eet ent eee rae dns 66 usciti 13 installiert teret a de etr uns 68 partial loopfill in eed eee 335 09 pH electrode installation ict tpe e or PR err ndr Lr ee 63 MOUNUNG irritar Hd T cbe redeat 63
3. From the Component list select the optional component s you have installed by checking the box es Click OK twice and then Close Theinstructions associated to the new component appears when creating a new method or editing an existing method AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 21 ED Installation guidelines 757727772 9 Thecorrect tubing dimension for the column you intend to use 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 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 22 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Fraction collector Frac 901 GE 3 Fraction collector Frac 901 3 1 General Frac 901 can be used for both small scale and preparative scale purifications with AKTAFPLC It collects up to 175 fractionsin 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 T
4. Fraction UniNet 2 y collector n Termination plug UniNet 1 chain 8 Connect a mains cable supplied between Frac 901 and a mains Socket at the rear of AKTAFPLC To UPC 900 Free mains outlet for optional equipment To Frac 901 From mains outlet 24 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Fraction collector Frac 901 3 3 3 Operation 3 3 1 Preparing for operation 1 Start your KTAFPLC optional configuration system as described in 2 6 Preparation for use 2 n Method Editor go to the Instruction field and select Frac 3 Check out the instructions available for Frac 901 to familiarize yourself with thenew functions available R efer also to the Fraction collector Frac 900 901 U ser M anual to learn more about using Frac 901 3 3 2 Fractionation principles and definitions Fraction collector Frac 901 is used for flowthrough fractionation before the elution and for collecting fixed volumes and or peak fractions during elution Fixed volume 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 fixed volume fractionation is valid for collecting fractions during
5. Superloop permits the introduction of larger volumes of sample 1 150 ml onto the column Superloop must not be used with columns generating a column pressure higher than that stated in thetable 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 the variable Pressure_limit Superloop is an accessory available in three sizes Volume Max allowed column pressure Code no a AMA 16 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 However Superloop can also be filled automatically refer to section 12 5 Loading a Superloop on page 82 16 1 Preparation Prepare the injection valve and connect Superloop as follows 1 Connect the supplied Pos 1 LOAD Union Luer female 1 16 Column maleto port 3 of the injection valve Pump Sample psi 2 Ensure that tubing for the syringe Top waste is connected to port 4 pus of the injection valve il Bottom KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Superloop 16 E
6. CAUTION Make sure KTAFPLC is switched off before installing the optional components The mains power to AKTAFPLC must be switched OFF before disconnecting or connecting the UniNet 2 cables 1 Mark the optional function valve with V7 V8 or V9 as required Check that the ID code switch is set to 7 8 or 9 respectively 2 Mountthe bracket and attach the valve to the system rack Choose the best location to facilitatethe routing of thetubing that connects the valveto the main system flowpath 3 Cut and mount the necessary capillaries PEEK i d 0 5 mm see also the Instructions of the valve Restrict the length of the capillaries as much as practically possible to reduce band broadening effects 4 Usetheattached UniN et 2 cable to connect the valve to the UniN et 2 communication link The valve can be connected anywhere between Pump P 920 and the termination plug Check that all capillary and electrical connections are carried out correctly T he following example shows how an optional IN V 907 is connected when used as a reversed flow valve Reversed flow configuration DOWNWARD FLOW position 1 UPWARD FLOW position 2 60 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA General function valve 10 10 2 Operation 10 2 1 Preparing for operation 1 Startyour KTAFPLC optional configuration system as described in 2 6 Preparation for use M ake sure that Valve V7 Valve V8 or Valve V9 is se
7. KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 29 E Fraction collector Frac 950 30 Fixed volume fractionation is terminated by issuing the command FractionationStop 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 or by setting the peak fraction sizeto zero using the parameter PeakSize This will not terminate fixed volume 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 fixed volume or peak fractionation The new fixed volume or peak fraction size is used instantly If a new peak fractionation command is issued during fixed volume fractionation peak fractionation is valid during peaks and fixed volume fractionation before between and after peaks If a new fixed volume fractionation command is issued during peak fractionation peak fractionation is valid during peaks and fixed volume fractionation before between and after peaks this means that the new command becomes effective when the peak is ended If the parameter Start at is set to Next line or Next group for peak fractionation and fixed volume fractionation the fractionation during the next elution will start at the next line or group
8. KTA design Optional Configurations AKTAFPLC User Manual Amersham e Biosciences Important user information All users must read this manual to fully understand the safe use of the components WARNING The WARN ING sign highlights instructions A that must befollowed to avoid personal injury Do not proceed until all stated conditions are clearly understood and met Caution The Caution sign highlights instructions that must be followed to avoid damage to the product or other equipment Do not proceed until all stated conditions are clearly understood and met Note The Note sign is used to indicate information important for trouble free and optimal use of the product Terms and Conditions of Sale Unless otherwise agreed in writing all goods and services are sold subject to the terms and conditions of sale of the company within the A mersham Biosciences group which supplies them A copy of these terms and conditions is available on request Should you have any comments on this product we will be pleased to receive them at Amersham Biosciences AB SE 751 84 Uppsala Sweden AKTAFPLC User M anual 18 1174 46 Edition AA Trademarks FPLC Superloop UNICORN KTA KTA explorer AKTAbasic AKTA purifier and AKTAFPLC are trademarks of Amersham Biosciences Limited Amersham and Amersham Biosciences are trademarks of Amersham plc Office Addresses Amersham Biosciences AB SE 75
9. Set Selected Parameter To Strategy Default Value Cancel Help The parameter Time between samples is used to set the scan cycle for curve updating The shortest possible interval is 1 s AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA o EN 15 AD 900 15 1 General AD 900 is an A D converter for connecting an external instrument to AKTArPLC system The module has one high resolution analog input for monitoring e g pressure UV monitor signals or other signals available as voltage outputs AD 900 communicates with the UN ICORN control system via UniN et 2 15 2 Installation CAUTION Make sure KTAFPLC is switched off before installing the optional components The mains power to KTAFPLC must be switched OFF before disconnecting or connecting the UniNet 1 and UniNet 2 cables 1 ConnecttheM ini DIN cable connector to socket Analog In at AD 900 Note Thelength of theanalog cable must not exceed 3 m 2 Removethe protection cover from the wires marked 1 and 2 at the other end of theM ini DIN cable 3 Connect wire 1 brown to the external device high output Connect wire 2 red to the external device low output Code 0 5 Set the ID switches as shown in the figure 1 2 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 99 ED 100 6 7 8 Connect the module with two UniN et cables as a part of the UniN et 2 chain On
10. deo mr piece hoic s capillary connectors ssssseenm description nens A nian reve raea installation rrr preparing a method escsccssessecseeesesseeseeeees preparing for operation ssseeenee Sensitivity iii ere ene te deeds KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 115 E 116 setting up instructions for function sssssssen 94 testing cachet teen ee dee cha er e de erts 93 Air 900 aCGOSSOLlGo Taldea 112 Control DOX sous ter ta P te eem ee OR HD e RUE 88 Air 912N using in the sample inlet flowpath sssem 90 Air 925 using in the system pump inletflowpath 89 Autosampler A 900 aCCessoriGs eere crece spere bedded ELE Dea dav eL E UG 111 Autosampler A 905 ACCOSSOMES iaa dada dida dada 111 auxiliary equipment ICE METTE 96 preparing for operation ssssssee eee nnns 97 signal handling setup sssss Hem 97 B buffer selection function USING IN AT N aia caeterae ea Tm PP Pp o er n a 59 buffer selection valves HEEL 58 preparing for operation ssssssss eene nnne 59 C calculate the delay volume sssssseee eee 104 capillary connections A900 coher irs eae ete ERR d tete 38 A 005 a aat te ats 50 checking the needle height A900 a tete ste ee A Pod crea cet 41 non idilio taa 52 column selection valves INStallavOn Nasa 56 OPEN aria di 57 component and c
11. If the parameter Start atis set to Nexttube thefractionation startsin 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 H owever 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 e 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 anew fractionation command and Continue is ordered the fraction collector continues with Start at tube number AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Fraction collector Frac 950 EN Note Before executing a fractionation instruction the outlet valve optional must manually be set to the position where the fraction collector is installed 4 3 3 Using Frac 950 in a method Set the fraction collector parameters in the M ethod Wizard when creating the new method e Flowthrough fractionation and the fractionation volume are selected in the Frac 950 Settings Flowthrough Fractionation dialog During the
12. AirSensorP960 Sample container AirSensorP960 in the sample inlet tlow when applying sample directly onto the column with a sample pump 13 2 Installation CAUTION Make sure KTAFPLC is switched off before installing the optional components The mains power to AKTAFPLC must be switched OFF before disconnecting or connecting the UniNet 2 cables 13 2 1 Fitting the Air 900 control box 1 Makesurethatthe KTAFPLC chromatography system is switched off 2 Check that theID 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 EK Code 0 O Q 3t 666 mee AIR CELL gt o UniNet 2 12 88 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Air sensors ety u 7 1 Attach the Air 900 control box vertically to a suitable mounting slot Lock in position by pushing up the snap lock 13 2 2 Connecting to the UniNet 2 communication network Usethe attached UniN et 2 cableto connect the Air 900 control box to the UniN et 2 communication link It can be connected anywhere between the system pump P 920 and the termination plug 13 2 3 Air sensor Air 925 AirSensor1 in the system pump inlet Remove the capillary from the switch valve inlet A 1 To system pump inlet 2 Cut new Teflon capillaries as shown in the figure and attach the
13. For 220 240 VAC usetwo 2 5 AT fuses slow CAUTION DO NOT switch on any of the equipment yet 6 When thevoltage selection and fuses are correct for thelocal mains power source plug in the supplied power cord in a freeoutlet in the base platform of the AKTAFPLC separation unit KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 37 EB A 900 m 777777 8 33 Capillary connections The capillaries to be connected between A 900 and the KTAFPLC separation unit areincluded in AKTA XT Tubing kit no 18 1122 92 Theflow diagram below showsthelocation of A 900 in theAKTArPLC flow path Autosampler Injection valve For Installation test Column Injection Valve V1 Fraction Collector 1 Remove the capillary between the injection valve V1 and the column top inlet 2 Select the capillaries matching the tubing kit to be used in AKTAFPLC after the column Use the capillaries marked G A1 and G A2 with the i d 0 25 mm tubing kit G A1 and H A2 with the i d 0 50 mm tubing kit and F A1 and F A2 with the i d 0 75 and 1 0 mm tubings Note See KTAFPLC System M anual for general recommendations on selecting column and tubing kit 38 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Autosampler A 900 ea SS 3 Connect the capillary marked G A1 F A1 I 2 1 3 m from the tubing kit between the injection valve V1 port 1 and the A 900 injection valv
14. ssssem 86 T termination plug ainia atada nee 20 U UNIN CEL ee eterne E rr te rn rt Pepe PR ERE 19 A 900 iristallatiofi voii cine 36 48 Frac 901 installation eme 24 Frac 950 installation sssseseeeneeennenennennns 28 UNINCEZ E 19 A900 taa 89 buffer selection valve nenne 58 column selection valve sssssseeeneemeemeen nenne 57 general function valve sssseeeeennnnnnnnn nnne 60 sample pump and sample valve sissies 68 sample pump and sample valve installation sess 68 119 ee V Valve INV 907 ACCESSOMIES uasa aa Rad E Rode a red ra Rd 112 valves features 13 function 19 instruction in UNICORN 19 120 KTAFPLC Optional Configurations U ser M anual 18 1174 46 Edition AA Short instructions Thefollowing 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 1 Wash the KTAdesign flow path with distilled water Switch off KTAdesign Prepare the component s Attach the components to AKTAdesign Prepare the necessary capillaries and connectors Ui BP WN Mount the capillaries and connectors YX Oo Connect the components to the UniNet chain and if required to the mains power supply 8 Ch
15. A 905 has two transportation safety devices which must be removed Removethelocking screw holding the front cover It is located at the right side of the front cover Savethe 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 905 U ser M anual Chapter 2 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 47 6 Autosampler A 905 for KTA A 62 2 Electrical connections WARNING A 905 must be connected to a grounded mains socket It is recommended to use the mains socket in the system WARNING For continued protection against risk of fire replace only with fuses of specified type and rating See Technical specifications in the A 905 User Manual for fuse data CAUTION The mains power to KTAFPLC must be switched OFF before disconnecting or connecting the UniNet 1 cables 1 Makesurethat the KTAFPLC chromatography system is switched off 2 DisconnecttheUniN et 1 cablerunning between M onitor UPC 900 and the computer at the computer end 3 Connect this UniN et 1 cable to any of thetwo UniN et 1 connectors in A 900 Autosampler i A Fraction collector Termination plug UniNet 1 chain To c
16. Instructions E xl Instructions r AirSensorl Parameters c AirSensorP960 Mode Alame Mode High_Sensitivity AirSensorl C Specials Flow from SystemPump Flow from c AirSensor2 c C SampleP Monitors Mode High Sensitivity SystemPump amplePump Flow from SamplePump AirSensor3 Mode High Sensitivity PIS Flow from SystemPump ET Set Selected Parameter To Strategy Default Value Cancel Help Note Thesensitivity of theair cells can be selected at three levels high medium or low sensitivity We recommend you use the high sensitivity setting as standard Check that the connected air cell s are indicated in the flow scheme in UNICORN Verify that the air cells are functioning properly by issuing manual commands from UNICORN asfollows AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Air sensors ety es 13 3 2 Testing AirSensorl located in the main system flow path 1 Confirm that the air sensor AS1 is shown in the flow scheme 2 Fill a sample container with distilled water and submergetheinlet filter of theA 1 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 n System Control select Manual Pump 4 Selectthe instruction Flow in the instruction list 5 Set Flowrate to 5 ml min and click Execute 6 While the system p
17. Set the fraction collector parameters in the M ethod Wizard when creating the new method Flowthrough fractionation and the fractionation volume is selected in the Wash Out Unbound Sample dialog D uring the wash out before elution fractionation a specified volume is collected Jtlution fractionation Fixed Volume Fractionation and or Peak Fractionation is selected in the Elution Fractionation dialog If fixed volume fractionation is selected a fixed volume size set in this dialog is collected during the complete elution If peak fractionation is selected only the peaks will be collected If fixed volume fractionation and peak fractionation is selected the fixed volumes will be collected before between and after the detected peaks The peak fractionation parameters are specified in the Peak Fractionation dialog Either the level or the slope of the UV curve or the signal level from A D 900 if used can be used for detecting a peak W hen using the UV curve level or the AD 900 signal level the level for starting and finishing the fractionation fraction size and minimum peak width if not specified a column should be specified W hen using the UV curve slope the slope value for starting and finishing the fractionation fraction size and minimum peak width if not specified a column should be specified Note Werecommend a maximum flow rate of 3 5 ml min when using Frac 901 with the i d 0 25 mm tubing kit At higher fl
18. 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 KTAFPLC is switched off before installing the optional components The mains power to KTAFPLC must be switched OFF before disconnecting or connecting the UniNet 1 and UniNet 2 cables AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Installation guidelines cu 2 Installation guidelines 2 1 General To use optional equipment in a chromatographic run suitable instructions is selected in the M ethod Wizard when creating a new method The general procedures for creating and editing methods are described in theUNICORN User M anuals Note Thesystem configuration delay volume must berecalculated and reset when the standard configuration after the UV flow cell is changed D etails are given in section 17 Configuration dependent delay volume 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 AKTAFPLC optional equipment is installed correctly 2 3 Pre requisites UNICORN version 4 12 or higher
19. The upper alarm level is set by the parameter HighAlarm and the lower alarm level by LowAlarm 5 5 11 Warning AutoSampler The Waming AutoSampler parameter is used to set warning temperatures when using the cooling function The upper alarm level is set by the parameter HighWarn and the lower alarm level by LowWarn KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 45 6 Autosampler A 905 for KTA 6 Autosampler A 905 for KTA 6 1 General Autosampler A 905 for KTA is intended for automated multiple sample injections of volumes down to 1 0 ul Cover in open position Buffer tubing High pressure valve Tubing guide Syringe waste _ Needle arm Syringe dispenser _ Transport solvent wash vial _t Microplate Wash solvent bottle Wash position Drain wash position Support plate Condensed water and leakage drain 46 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Autosampler A 905 for AKTA 6 7777 62 Installation 6 2 1 Preparations A 905 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 AKTAFPLC base platform The recommended position isto the left of the separation equipment of AKTAFPLC with approximately 3 cm spacing Locking screw Red cap
20. again Stop the pump by clicking End in the System Control toolbar 10 Setup instructions as described below to monitor the AirSensorP960 signal to suit your needs 13 3 4 Setting up instructions for the air sensor function For AirSensor1 3 the presence of air in theflow path is detected by the Air 900 control box which reports its status to UNICORN For AirSensorP960 air is detected by the control software in Pump P 960 The desired control action based on the status report is governed by instructions in the method used Instructions for handling the air sensors must be inserted manually in a method Alarm AirSensor1 3 and Alarm AirSensorP960 Enables disables alarm generation for theair cells An active alarm sets the chromatography system to Pause mode Alarms can be activated deactivated in System Settings or inserted as instructions in a method They can also be activated deactivated manually in Manual Alarms amp Mon mr 1 Parameters Breakpoint T instructi ies WES Alam AjSensorP360 E Mode Insert poo v vi Alarm AlrSensor Va C Disabled Enabled C Flowpath lam Asesor Va x SetCondScale0 ce AlamstMon SetCondScale1 00 WatchPar UV1 C Frac WatchPar Cond Replace WatchPar_Cone Watch WalchPar pH WatchPar Pressure Other WatchPar Flow El eee Ready num VA Stop AirP960 When the instruction is enabled Pump P 960 will stop when air is detected The method execution then
21. cable to connect the valve to the UniN et 2 communication link The valve can be connected anywhere between the system pump P 920 and the termination plug connected to the injection valve 5 Check that all capillary and electrical connections have been carried out correctly 54 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Outlet valve 7 2 1 Preparing for operation 1 Start your AKTArPLC optional configuration system as described in section 2 6 Preparation for use 2 Theoutlet 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 manual commands from UNICORN as follows n System Control select Manual FlowPath Select instruction OutletValve Set theoutlet valveto position F4 Manually run Pump P 920 with distilled water at a flow rate of approximately 2 ml min and check that water is coming out from port 4 of the outlet valve 7 2 2 Using the outlet valve for fractionation To usethe outlet valve in a method select the desired outlet valve options in the Wash Out Unbound Sample and or Elution Fractionation dialogs when creating the method in the M ethod Wizard 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 ava
22. cell 2 mm 15 UV flow cell 5 mm 10 Conductivity cell 24 pH flow cell 100 Accumulator Frac 950 40 Union 1 16 male 1 16 male 0 25 mm i d 1 8 Union 1 16 male 1 16 male 0 5 mm i d 7 Flow restrictor FR 902 5 Capillary i d 0 25 x 100 mm 4 9 Capillary i d 0 50 x 100 mm 19 6 Capillary i d 0 75 x 100 mm 44 2 Capillary i d 1 0 x 100 mm 78 5 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 105 ED Strategies and Method Wizard 18 Strategies and Method Wizard 18 1 Strategies overview FPLC with Frac 950 FPLCF400 FPLC with Frac 901 FPLC_400 The last three digits in the strategy name is the version number 106 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Strategies and Method Wizard 18 18 2 Method Wizard M ethod Wizard Configuration file version 2 0 Thefollowing components are supported by the M ethod Wizard Fraction collector Frac 901 Fraction collector Frac 950 Autosampler A 900 Autosampler A 905 Column valve Outlet valve Buffer valveA1 e Buffer valve A2 Sample pump P 960 Superloop partly Air sensor 1 3 Air sensor P 960 AD 900 e Conductivity monitoring pH monitoring KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 107 ED Strategies and Method Wizard 108 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Accesso
23. continues immediately with the next instruction i e the system will not be set to Pause mode The instruction can be enabled disabled in System Settings or inserted as instructions in a method It can also be enabled disabled manually in Manual Alarms amp Mon l SS Os eee Rae Instructions 1 Wavelength Insert C Pump Imm n Q0 e E Ariana TimeLIV Vat Disabled C Enabled VES Flowpath Alarm UV1 AlamskMon C Frac C Watch C Other Ready NM y 94 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Air sensors 13 A Watch AirSensorl 3 and Watch AirSensorP960 M onitors the air sensor status signals from the air cells Performs a user defined action if the test condition is fulfilled M onitoring is cancelled automatically once the condition has been fulfilled The user defined action can be for example PAUSE or a call to a block ls 0 00 Block Eluent 4 Inlet Breakpoint Instructions Parameters TE A Watch Conc READY Insert 0 00 al Pump Watch pH Sample Inlet BuffeiValve B cv al Start Conc B 0 pc C Flowpath ich Flow Start Instructions Var e Change E E Start_with_PumpWash_E xplore Watch AirSenP960 System Volume Comp ion T a bs PAUS Replace Watch AirSensor2 Watch Watch AirSensor3 C Other ao E Delete Ready is C NM z 13 3 5 Preparing a method for using air sensors The following two examples show ho
24. elution between peaks Theinstruction Fractionation 900 startsfixed volumefractionation The instruction Peak Fractionation 900 starts peak fractionation Fixed volume 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 sizeto zero usingthe parameter PeakSize T his will not terminate fixed volumefractionation if thetwo 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 fixed volume or peak fractionation The new fixed volume or peak fraction size is used instantly KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 25 EP Fraction collector Frac 901 If a new peak fractionation command is issued during fixed volume fractionation peak fractionation is valid during peaks and fixed volume fractionation before between and after peaks If a new fixed volume fractionation command is issued during peak fractionation peak fractionation is valid during peaks and fixed volume fractionation before between and after peaks this means that the new command becomes effective when the peak is ended 3 3 3 Using Frac 901 in a method
25. helatter function is based on peak detection using slope or level sensing Fraction marks and fraction numbers make it easy to identify fractions and peaks Drop synchronization eliminates sample loss below flow rates of 3 ml min during tube change 3 2 Installation 1 Unpack and assembleFrac 901 according to the instructions given in the Fraction Collector Frac 900 901 User M anual 2 Place Frac 901 to the right of the system On line filter Mixer Pump A Pump B A A ae Injection valve V1 Flow restrictor Fraction collector Frac 901 3 Connect a tubing from the outlet of the flow restrictor to the IN porton ro delivery the flow diversion valve arm NC 4 Connect a tubing from theN C normally closed port on the flow diversion valve to the delivery arm on Frac 901 Follow the From flow instructions given in the restrictor IN Frac 900 901 User M anual To waste NO AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 23 EP Fraction collector Frac 901 5 Connect a tubing from the NO normally open port on the flow diversion valve to waste 6 Extend the drain tubing from Frac 901 34 to a suitable waste bottle 7 Modify the UniN et 1 data communication chain by Connecting a UniN et 1 cable between Pump P 920 and Frac 901 Connecting a termination plug in theother UniN et 1 connector in Frac 901 To PC controller board
26. in AKTAFPLC enables Automated multiple sample injections from 1 ul to 1 ml of sample solution Sample stored in tubes in a circular rotating rack Quantification of amount concentration and molecular size External standard quantification nternal standard quantification Standard addition Recovery calculations Molecule size calculations Q uantification 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 Autosampler A 900 with Cooling provides internal cooling of the sample solutions Autosampler A 905 used in AKTAFPLC enables Automated multiple sample injections from a few microliters up to 1 ml of sample solution Sample stored in microplates thus allowing samples collected in microplates in Frac 950 to be used directly in A 905 Autosampler A 905 also provide cooling of the sample solutions AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Introduction qu REESE 1 3 4 Valves Five PV IV 908 valves can be connected as dedicated function valves These valves are used as follows Two PV 908 rotary valves used in conjunc
27. must be installed in the computer connected to your AKTA rPLC system AKTAFPLC must be installed and fully tested before the optional components are installed See the Installation G uidefor your AKTArPLC chromatography system thoroughly with distilled water before removing any capillaries or WARNING Ensure that the entire system has been flushed components CAUTION Make sure KTAFPLC is switched off before installing the optional components The mains power to KTAFPLC must be switched OFF before disconnecting or connecting the UniNet 1 and UniNet 2 cables AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 17 ED Installation guidelines 7 77 24 Principle installation of components CAUTION Make sure KTAFPLC is switched off before installing the Eu optional components The mains power to AKTAFPLC must be switched OFF before disconnecting or connecting the UniNet 1 and 0 UniNet 2 cables a Optional equipment can be mounted on the system rack with the rack X door closed or open Both the inside and the outside are provided with mounting rails for quick mounting of components Theupper part of the system rack can beraised and locked with screws in optional positions along its vertical rods Thisispractical when using long columns When selecting mounting location for the optional equipment it is important to choose a location which m
28. range from a few microliters to 1 ml in a simple and reproducible manner gt Buffer tubing From system pump _ High pressure valve Til Tubing guide Sample loop Needle arm Tray segment Wash solvent y f 7 Drain wash bottle position Wash position Condensed water and leakage drain KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Autosampler A 900 ea u S 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 0 396 Partial loopfill The sample loop is partially filled with sample giving low sample loss and allowing programmable injection volumes wl 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 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 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 33 EB A 900 77 5 2 Autosampler A 900 with Cooling
29. 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 S pecials FracParameters DelayVol 17 1 Standard configuration The standard configuration of an KTAFPLC system has the following delay volumes AKTAFPLC e Including Frac 901 375 ul Including Frac 950 Including accumulator Tubing kit with tubing No accumulator Standard mode 348 ul 197 yl Prep mode 865 ul Not applicable 17 2 Optional configuration Calculate the contribution from each new component and capillary according to the table below and add this additional volume to the standard configuration delay volume value The example below shows a calculation for including a pH flow cell in an AKTAFPLC system with Frac 901 Capillary volume flow cell volume 90 100 x 19 6 100 ul 128 ul The length of the new 0 5 mm i d capillary is 90 mm the volume of 100 mm tubing is 19 6 ul and the volume of the pH flow cell 100 ul The new delay volume value is 375 128 403 ul AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Configuration dependent delay volume FERE LLL A 17 3 Component and capillary volumes The volumes are theoretical values Component capillary Volume pl Valve PV 908 7 Valve 1V 908 21 Valve INV 907 9 Valve FV 903 113 UV flow
30. useful when large amounts of sample is to be loaded The sample is pushed onto the column Note In this configuration some columns generate a back pressure above 2 0 M Pa at their recommended flow rates If the back pressure exceeds 2 0 M Pa the upper pressure limit of Pump P 960 the pump will stop immediately To achieve a back pressure below 2 0 M Pa when using these columns we recommend decreasing the flow rate We particularly recommend decreasing the flow rate of the system pump during equilibration before sample application and before switching the sample pump in line The sample pump will not start if the back pressure exceeds 2 0 M Pa 12 4 1 Connecting the capillaries 1 Makesurethat the components are properly installed according to section 12 2 Installing the components 2 Cutthecapillaries to suitable lengths and connect them using connectors and ferrules as indicated in the table and in the figures Those lengths are maximum values 3 Check that all capillary and electrical connections have been carried out correctly gg Fingertight connector 1 16 Pump P 960 i Union fingertight female M6 male Outlet connection pe E aA Union 5 16 female HPLC male FS HE fi i Wi Tye _ Tubing connector for 1 8 o d tubing with S 7 Pa ferrule for 1 8 o d tubing i WTO Inlet connection 76 KTAFPLC Optional Configurations User M anual 18
31. 1 84 Uppsala Sweden Amersham Biosciences UK Limited Amersham Place Little Chalfont Buckinghamshire England H P7 9NA Amersham Biosciences Corp 800 Centennial Avenue PO Box 1327 Piscataway NJ 08855 USA Amersham Biosciences Europe GmbH M unzinger Strasse 9 D 79111 Freiburg Germany Amersham Biosciences K K Sanken Building 3 25 1 H yakunincho Shinjuku ku Tokyo 169 0073 Japan Amersham Biosciences Limited 13 F Tower Ever Gain Plaza 88 Container Port Road Kwai Chung N ew Territories Hong Kong www amershambiosciences com Copyright Amersham Biosciences AB 2003 All rights reserved Contents Introduction LW zGeneraliersss A et decet eee ER s ER e RETE a RE 1 2 Scope of Manual si ihe edet ete np ene actes 1 3 Summary of optional configurations sssssssss O nene Installation guidelines 2 1 A pr tee ese pies 22 Unpacking eere eere 2 3 O Prerequisites senri ria Ta ec ete eh 2 4 Principle installation of components sssr 2 5 Components instruction overview see 2 6 Preparation for use ssssssssssseeeeeeeene Fraction collector Frac 901 A tco als terit beendet Dee vro beber e e o en 3 2 Installation senes O oed ea t eet pen forts Fraction collector Frac 950 CA EHE 4 2 Installation casada a 4 3 Operation eine eredi des Autosampler A 900 BETS SGM All s t emet res Ep is 5 2 Autosampler A 900 wit
32. 1120 48 o d 7 mm Sample needle stainless steel 1 A 18 1148 15 Wash solvent bottle 250 ml 1 A 18 1120 33 Autosampler A 905 for KTA Autosampler A 905 for KTA complete 1 A 18 1175 93 Sample needle fused silica 5 ul 1 A 18 1158 68 PEEK loop 20 yl 1 A 18 1160 49 Common autosampler accessories AKTAdesign XT upgrade kit 1 A 18 6083 19 AKTA XT Tubing kit 1 A 18 1122 92 Sample needle PEEK 15 pl 1 A 18 1119 99 Buffer tubing 500 pl 1 A 18 1120 32 Buffer tubing 2000 ul 1 A 18 1120 31 Syringe 100 ul 1 A 18 1120 34 Syringe 250 ul 1 A 18 1120 35 Syringe 500 ul 1 A 18 1120 36 Syringe 1000 pl 1 A 18 1120 37 Luer lock connection female 1 A 18 1120 38 PEEK loop 100 ul 1 A 18 1120 40 Syringe waste tubing 1 A 18 1120 44 Syringe tubing 1 A 18 1120 45 111 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 19 fuere and consumables 112 Item Quantity A C Code no pack Valves Valve INV 907 including 1 A 18 1108 40 one UniNet cable fill port needle and syringe holder are not included Valve IV 908 including 1 A 18 1108 42 one UniNet cable Valve PV 908 including 1 A 18 1108 41 one UniNet cable 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 18 1112 92 dummy electrode Dummy electrode round tip 1 A 18 1111 03 Pump P 960 Pum
33. 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 Tefzel i d 1 0 mm o d 1 16 3m 18 1142 38 Miscellaneous Extra rack panel KTAFPLC 1 18 1128 64 Communication cable P 920 P 50 1 18 1129 60 A accessory C consumable 113 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 19 fuere and consumables 114 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Index Symbols PI PICKUP coacalco A A 900 checking the needle height electrical connections features ira injection methods ov ee installation uere ote terere nes malti FUSES lite ebd operation mii ra purging the syringe ssssee site requirement ssssesseeeenn unpacking aaaeeeaa aaa iaaa AEEA a aE A 905 checking the needle height electrical connections ssssseeess feat res iii injection methods installation caca mail TUSES decerni on rg Operation cocer enne purging the syringe sssssm site requirement ssssssseeeen UNPACKING insisto aireado AD 900 A A TN description aee cedens l1 Tr MEE adi installato cerne ie preparing for operation sssesseeeenneeenn eene using for monitoring a UV signal air sensors AISOTI2 uisus Pitra Poco dun Sten aren Dieu AI 025
34. 1174 46 Edition AA Sample pump P 960 and sample valve c On line filter Column Injection Valve V1 Fraction Collector Restrictor Capillary max length Connector Connection points 1 Tefzel i d 1 0 mm x 50 cm Fingertight connector 1 16 18 1112 55 Sample valve center port Sample inlet tubing oooooo c l Iy Fingertight connector 1 16 18 1112 55 Sample pump inlet Union fingertight female M6 male 18 1112 57 2 Tefzel i d 1 0 mm x 50 cm Fingertight connector 1 16 18 1112 55 Sample pump outlet Fingertight connector 1 16 18 1112 55 Injection valve port 2 3 PEEK i d 0 75 mm x15cm FPingertight connector 1 16 18 1112 55 Injection valve port 3 Fingertight connector 1 16 18 1112 55 Injection valve port 6 4 Tefzel i d 1 0 mm x130 cm Fingertight connector 1 16 18 1112 55 Injection valve port 5 5 Tefzel i d 1 0 mm x 10 cm Fingertight connector 1 16 18 1112 55 Injection valve port 4 Fingertight connector 1 16 18 1112 55 Flow restrictor inlet 6 Tefzel i d 1 0 mm x120 cm FPingertight connector 1 16 18 1112 55 Flow restrictor outlet 7 Teflon i d 1 6 mm x 130 cm Union 5 16 female HPLC male 18 1142 08 Sample valve Tubing connector for 1 8 o d tubing 18 1121 17 ports 1 8 with ferrule for 1 8 o d tubing 18 1121 18 Only if you have a sample va
35. 1174 46 Edition AA pH flow cell and pH electrode mounting 211 113 Operation 1 Start your KTA optional configuration system as described in 2 6 Preparation for use 2 If fraction collection should be used remember to change the delay volume This setting is found in System Control in System Settings Specials Seealso 17 Configuration dependent delay volume Calibratethe pH electrode and set up the additional monitor functions desired for pH measurement These procedures are described in the M onitor UPC 900 User M anual and in the UNICORN User M anuals KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 65 12 Sample pump P 960 and sample valve 12 Sample pump P 960 and sample valve 66 12 1 General Pump P 960 isa laboratory pump for use as a sample pump to apply samples directly on the column and or to fill sample loops including Superloop Pumping action is provided by plungers 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 at a pressure up to 2 M Pa An IV PV 908 valve can be used as sample valvein combination with the sample pump It is a motorised 8 way valve used for automatic sample application of up to 8 samples Note The Method Wizard does not support the sample valve function It is added as a general function valve to a method in the M et
36. 3 Ensuyrethat Superloop is filled with liquid see separate Superloop Instruction 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 Makesureall connections are fingertight 16 2 Filling Superloop Filling Superloop is achieved as follows 1 Set the injection valve to position INJECT 2 Start Pump P 920 and let it run until the movable seal has reached the bottom of Superloop 3 Stop Pump P 920 and settheinjection valve to position LOAD 4 Load a large volume syringe with sample 5 Gently load the syringe contents into Superloop through port 3 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 7 IntheMethod Wizard select Manual as sample application technique The volume to inject is set by parameter Empty Loop with 8 Thesampleis applied to the column when the injection valve is set to position INJECT When the required volume has been injected set the valveto LOAD lower than the maximum allowed pressure of the Superloop WARNING Make sure that the upper pressure limit of the system is AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 103 Configuration dependent delay volume 17 Configuration dependent delay volume 104 The
37. 4 Gently move the sample inlet tubing to the sample vessel AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Sample pump P 960 and sample valve c u 7 7 12 3 6 Preparing a method for filling a sample loop To usethe sample pump for filling a sample loop select Sample Pump Loop Filling P 960 as sample application technique in the M ethod Wizard when creating the new method To use an additional valve as sample valve for example Valve 7 add the Valve7 instruction to the method in the M ethod Editor The M ethod Wizard does not support the sample valve function The sample pump properties are designated P960 Flow and P960 Press in the flow scheme and run data windows The valve properties are designated for example Valve7 in the flow scheme and run data windows Note When filling the sample loop the resistance in theflowpath may cause cavitation if the flow rate exceeds 0 5 ml min Note For complete filling an overfill of 2 5 times the loop volume is needed for maximum reproducibility between the runs 12 3 7 Emptying the sample loop W hen emptyingthe sample loop usea buffer volume of approximately 5 times the sample loop volume to flush the loop and ensure that all sample is injected onto the column KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 75 12 Sample pump P 960 and sample valve FEE 124 Direct loading onto the column Loading sample directly onto the column is
38. 4 in equipment controlled via Pump P 920 REMOTE connector 98 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 pane in UNICORN Theinstructions also monitor the signal status for the input signals and perform user defined actions if test conditions arefulfilled M onitoring is cancelled automatically once a condition has been fulfilled Pre selected action can be for example call to a block Mi o 00 Block Wash Out Unbound Sample hal Breakpoint 0 00 Zw Var J Instructions END BLOCK Pump Flowpath Watch P960 Press Fractionation C Alams amp Mon Watch AirSenP960 Gradient_Delay C Frac C Other Watch Watch Auxnd Watch pH END METHOD Watch Pressure Flow Rate Watch Flow Flowthrough_Fractionation Insert Watch P960 Flow Fractionation_Stop Lhange Watch AuxIn2 Replace Watch Off FREE Y Delete Ready mom 4 Curve representation of input output signals The status for signals AuxIn1 and AuxOutl can be stored as curves in the chromatogram T his is selected in System Settings by setting Curves Store ON Instructions Auslnil Parameters C Alarms BIN C OFF E ON C Specials Time between samples Time between samples 1 000 s 1 000 AusQutl Store ON Time between samples 1 000 s Bi C Monitors E s gie Store ON zl Time hehueen samnles 1 NNN
39. 902 If thesample vessel is placed at a higher level than the end of thesample waste tubing sample might accidently flow through the sample flow path to waste Flow restrictor FR 902 supplied is used in the sample flow path to eliminate this effect by creating a back pressure of 0 2 M Pa Thereby sample vessels can be placed for example on top of the system Theposition of theflow restrictor in theflow path dependson how the sampleis applied with the sample pump see Table 12 1 Theinjection valve position refers to when sample otherwise might flow straight through to waste Sample application type Flow restrictor port IN Injection valve connected to position Filling a sample loop Sample pump P 960 LOAD outlet port Direct loading onto a column Injection valve V1 port 4 INJ ECT Loading a Superloop Injection valve V1 port 5 INJ ECT Usea 10 cm long tubing between the flow restrictor and the port T he flow restrictor is required only if filling the Superloop several times during the run Table 12 1 Position of the flow restrictor KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 69 12 Sample pump P 960 and sample valve o 70 12 3 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
40. 92 Auxiliary equipment controlled via Pump P 920 REMOTE connector 14 1 Installation nme etg 96 14 2 Operation ecco d eei erc etre ente idee gt 97 AD 900 15 1 General eate e et dee t wien 99 15 2 Installation 5 2 erret eibi He eia 99 15 3 Operation ani ed adil eb iere een 100 Superloop 16 1 Preparati rasini annas tel 102 16 2 Filling Superloop eenm narrar 103 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Configuration dependent delay volume 17 1 Standard configuration sene 104 17 2 Optional configuration sse 104 17 3 Component and capillary volumes esee 105 18 Strategies and Method Wizard 18 1 Strategies overview ssssssennemes 106 18 2 Method Wizard ssssssssseeeemeeemnnns 107 19 Accessories and consumables 19 1 Connections chromatography standard selection 109 19 2 Optional components and consumables 110 Index AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Introduction qu Introduction 1 1 General KTA FPLC standard system configurations can be changed to optional configurations This flexibility in KTAFPLC system strategies allowsthe user to enhance already used purification methods and also to develop new more complex methods W h
41. Amersham Biosciences volumes between 0 1 2 0 ml can be applied The sample is drawn into the sample loop by the sample pump Note Theflow rate should not exceed 0 5 ml min when filling the sample loop O therwise cavitation might occur dueto the resistance in the flow path 12 3 1 Connecting the capillaries 1 M akesurethat the sample pump and the sample valve if included are properly installed according to section 12 2 Installing the components 2 Cut the capillaries to suitable lengths and connect them using connectors and ferrules as indicated in the table and in the figures The lengths in the table are maximum lengths 3 Check that all capillary and electrical connections have been carried out correctly Fingertight connector 1 16 Pump P 960 Outlet connection Union fingertight female M6 male Union 5 16 female HPLC male Q d Tubing connector for 1 8 o d tubing with o ferrule for 1 8 o d tubing IN Inlet connection AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Sample pump P 960 and sample valve c On line filter Fraction Collector Restrictor Capillary max length Connector Connection points 1 PEEK i d 0 75 mm x24 cm Fingertight connector 1 16 18 1112 55 Injection valve port 4 Sampie init tubing Fingertight connector 1 16 18 1112 55 Sample valve cen
42. Autosampler A 900 with Cooling is purchased separately It provides internal cooling of the sample vials This is done by a Peltier element and a fan inside the autosampler and a plastic hood covering the vials Cooling hood 34 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Autosampler A 900 ea A 53 Installation 5 3 1 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 AKTAFPLC base platform The recommended position isto the left of the separation equipment of AKTAFPLC with approximately 3 cm spacing ng screw A 900 has two transportation safety devices which must be removed Removethelocking 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 A utosampler A 900 User M anual Chapter 2 35 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA ED Autosampler A 900 57777 5 3 2 Electrical connections WARNING A 900 must be connected to a grounded mains socket It is recommended to use the mains socket in the system WARNING For continued protection against risk of fire repl
43. E connector on rear of P 920 Auxiliary equipment using digital input output signals can be connected to the 9 pole D SUB female REMOTE connector on the rear panel of Pump P 920 5 V TTL signals only Pins 1 4 handle digital input signals and pins 6 9 handle digital output signals Pin 5 is signal ground Pin Signal Function 1 4 Auxin 1 4 Active state can be selected to 0 or 1 in UNICORN Min duration 50 ms Max frequency 1 Hz ee Se 5 OV Signal ground ee e 6e e e e el 6 9 AuxOut 1 4 Active state can be selected to 0 or 1 in UNICORN 5 V TTL signals Use a shielded cable with a 9 pin D SUB male connector for the P 920 REMOTE connector The other end of the cable connects to the auxiliary equipment and must be adapted to ts relevant connector The connection example below shows the principle for connecting input output signals P 920 Digital signal interface Pin1 Pin2 Pin3 Pin4 Pin 5 Pin 6 Pin 7 Pin 8 Pin 9 REMOTE Connector Auxin 1 Auxin 2 Auxin 3 Auxin 4 AuxOut 1 AuxOut2 AuxOut3 AuxOut 4 From external equipment outputs To external equipment inputs Signal ground in external equipment 96 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Auxiliary equipment controlled via Pump P 920 REMOTE connector 14 14 Operation 14 2 1 Preparing for operat
44. FPLC are Connecting Fraction collector Frac 901 Connecting Fraction collector Frac 950 Connecting Autosampler A 900 and A 900 with Cooling and Autosampler A 905 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 Theactual useis completely user defined For example they can be used as a reversed flow valve or a sample selection valve Connecting sample pump P 960 Connecting up to four air sensors Connecting external equipment using digital input output signals through the system pump P 920 REMOTE connector Connecting AD 900 e Connecting a Superloop An extra rack panel is available as an accessory to accommodate optional equipment 1 3 1 Fraction collector Frac 901 Fraction collector Frac 901 is an automated fraction collector for use in AKTArPLC chromatography systems under UN ICORN control The functionality and performance is aimed for mid range usage po AS AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 11 ER Introduction 12 1 3 2 Fraction collector Frac 950 Fraction collector Frac 950 is an automated fraction collector for use in AKTArPLC chromatography systems under UNICORN control The functionality and performance is aimed for high end usage 1 3 3 Autosampler A 900 and Autosampler A 905 Autosampler A 900 used
45. ODGrallOni a ice 65 pH flow cell installation i ce rte ene nen pr tr oat 63 ies iio Aereas a n a a UM ainda 63 pH measurement accessories and consumables ssssesseeeee 112 pre requisites 1 cssc Ana Der cde 17 purging the syringe A 900 sss 40 purging the syringe A 905 sss 52 PV 908 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA ACCESSOMES aa 112 features ni Ac 13 R reversed flow configuration ssssssseee mes 60 S sample injection techniques ssssssseeeememmnns 42 InjectionF luShed 2 tete aaa 43 InjectionPartial cocinar 43 InjectionPick Up niece tr tret ne erp Lone 44 sample loop CM PLYIN Gs stet digerere pa ep roe PUE IDE 75 TOW Fate 21 niece tee het er knee ect ate Fix era iaa 70 I II 70 sample pump connecting the capillaries s 82 Teat t8s A ain eec erbe ae En cen eh aes 13 Sample pump P 960 Kit sss eene 67 sample valve installifig 425 citer ep tt ht tee ERE PER ER EVER Er EET 68 snap in brack t uso ntt eir Ie RO eO PO e Er ias 18 strategies OVerVieW see ecrire ca 106 Superloop ACTOSSOMES sistere nt fe Ap tin erpap ge t Tu 113 CONNECT sirsiran cei entr tbe t n RE RR RAE FR TEUER REEL a araa E dei 102 description 3 t te ai 102 feat resi ER 14 OS is recep acte n pa dt 103 A 82 preparing a method for loading
46. ORN must be informed about the insertion of an optional component Thisisknown as selecting the component which makes its corresponding instructions available To print a list of the general instructions and the instructions corresponding to the selected components 1 In the Method Editor select File Print 2 Check the Instruction set box and clear the other boxes 3 Click OK to print the instructions 20 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Installation guidelines cu 2 2 6 Preparation for use To start your AKTArPLC optional configuration system 1 Switch on KTAFPLC with the mains switch located at the front left on the system base Check that the computer and printer are switched on Log in see KTAFPLC M aking your first runs M ake sure you have the correct strategy installed in your system see 18 Strategies and M ethod Wizard A new strategy is installed according to the software installation instructions given in the UNICORN User M anuals In UNICORN Main Menu select Administration S ystem Setup Select System and then click Edit Click Component O ColumrValve V2 CO ColumrValve v3 O OutletValve V4 M BufferValveA V5 M BufferValveB V6 O Valve V O Valve V8 O Valve V8 C pH Cell M Fraction collector 900 Sample pump P 950 E CO AutoSampler 4 900 AutoSamplerThermo CO AutoSampler 4 905 O ADS00 O Airsensor P 960 C Airsensor 1 Cancel Help
47. Valve7 IV PV 908 Optional function 8 INV 907 Valve8 IV PV 908 Optional function 9 INV 907 Valve9 IV PV 908 Components are connected to the UNICORN control system using either UniN et 1 or UniN et 2 cables Both the UniN et 1 and the UniN et 2 data communication chain in standard configuration is routed from the rear of P 920 via their respective components to the last component in the chain where it is terminated with a plug Note When Pump P 960 is used it is always installed as the last component in the UniN et 2 chain Since the pump has an internal termination no termination plug is needed Frac 901 Frac 950 A 900 and A 905 are connected via the UniN et 1 chain All other optional components are connected via the UniN et 2 chain KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 19 ED Installation guidelines 222277z70z3z Both theUniN et 1 and the UniN et 2 chain can be interrupted anyw here betw een P 920 and the termination plug to interconnect the optional components in the chain Thetermination plug can be moved to thelast component furthest away from P 920 if motivated by cable routing considerations To PC controller board UniNet 2 Fraction chain collector i PC running UNICORN P 3 Exp Cc Terminati ermination plug E UniNet 1 2 5 Components instruction overview UNIC
48. ace only with fuses of specified type and rating See Technical specifications in the A 900 User Manual for fuse data CAUTION The mains power to KTAFPLC must be switched OFF before disconnecting or connecting the UniNet 1 cables 1 Makesurethat the KTAFPLC chromatography system is switched off 2 DisconnecttheUniN et 1 cablerunning between M onitor UPC 900 and the computer at the computer end 3 Connect this UniN et 1 cable to any of thetwo UniN et 1 connectors in A 900 Autosampler i A Fraction collector Termination plug UniNet 1 chain To computer 36 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Autosampler A 900 ea 227777 4 Conecta new UniN et 1 cable supplied with the A 900 from the other UniN et 1 connector on the A 900 to the computer The UniN et 1 connection to the computer must be made to the board with four green LED s Four green LEDs 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
49. apillary volumes sssseseemmm tetaee 105 components tii a E dip dea 21 cooling A2900 5 2 cnet e riter mate ter cred ee dede 34 cooling parameters sssssssseeeeeeene eene nnns 42 D delay vol mes comio cnet le treu 104 direct loading onto column preparing a method sssssssssee eene enne nnns 81 E electrical connections AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA E OO A 900 F filling a sample loop preparing a method flow path NEO sat votre cde rece rcp eg A 905 ins Flowrestrictor FR 902 ssssseemHee emn flushed loop semem Frac 901 accessories sssssee nennen features easet ete bte oett ef aas installation ssee ene Operation aiii ries using in a method ssseee Frac 950 dCCeSsSOF es inserite reme ERE UE feal leS uidet cine prie m npn s installation sseeeemn Operation sorcar nanasa a a ts optional components and consumables A standard mode sseee non ncan ona upgrading from Frac 901 ee using in a method ssseeeee fractionation order Frac 950 sssesess fractionation principles and definitions fixed volume fractionation Frac 901 fixed volume fractionation Frac 950 accu MEE Efacs950 tin se eter eh dea peak fractionation Frac 901 s peak fractionation Frac 950
50. bing to be filled Note If the syringe needs to be emptied switch the sample valve before removing the syringe to prevent sample from flowing back to the vessel Repeat step 6 and 7 for the remaining sample inlet tubings To fill the buffer inlet tubing in port 8 1 2 Set the sample valveto port 8 Draw buffer with the syringe until the liquid level has passed through the sample pump The check valves in the sample pump will prevent the liquid from being withdrawn when removing the purge tubing Disconnect the purge tubing Fit the original connector to the port Flush the sample flow path tubing with buffer to remove any trapped air bubbles according to the procedure in section 12 3 4 Removing air by running the sample pump Procedure for system not including IV PV 908 as sample valve 1 Put the sample inlet tubing in a buffer vessel Disconnect the connector fitted to the outlet port on the connection part of Pump P 960 i e before the pressure sensor Connect the purge tubing to the same port Set the injection valve V 1 to LOAD Draw buffer with the syringe until the liquid level has passed through the sample pump The check valves in the sample pump will prevent the liquid from flowing back when removing the purge tubing Disconnect the purge tubing Fit the original connector to the port Flush the sampleinlet tubing with buffer to remove any trapped air bubbles according to the procedure in section 12 3
51. d instructions are used when the syringeis 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 surethe syringe is run to the H ome position before leaving the manual function AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Autosampler A 900 ea e 5 5 7 RotateTray The RotateTray instruction is used to rotate the tray e g when new vials are loaded into the tray segments 5 5 8 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 5 5 9 AutoSampler Temperature W hen using A 900 with cooling function the AutoSampler Temperature parameter is used to enable the cooling function and to set the desired temperature The cooling function is enabled by deselecting the OFF check box The temperature range is 4 40 C The cooling starts immediately when clicking OK Note To use Autosampler A 900 with Cooling both AutoSampler and AutoSamplerT hermo should be selected in the Components list in System Setup 5 5 10 Alarm_AutoSampler The Alarm_AutoSampler parameter is used to set alarm temperatures when using the cooling function
52. d to detect air when automatic sample application using P 960 is performed Oneair cell can beused to detect air when applying large amounts of sample with the system pump Therearefour types of air cells available They aredesignated Air 912 Air 912N Air 925 and Air 925N and differ in inner diameter This makes them suitable to use with different sizes of capillary Air 912 has a smaller inner diameter and is suitableto usein the P 960 sample flow path Air 925 has a larger inner diameter and is suitableto use in the inlet flow path to the system pump P 920 Air 912N and Air 925N can be connected directly to P 960 Up to four detection points can be used in the process flow path The detectors are designated AS1 AS2 AS3 and ASP960 in the flow scheme 1 3 7 Auxiliary equipment via Pump P 920 REMOTE connection The system pump P 920 is provided with a digital input output connection comprising four output and four input channels The channels are opto isolated and accept standard T TL level signals Examples of use include External alarm input output Start stop of external equipment 1 3 8 AD 900 AD 900 is an A D converter for connecting an external instrument to the AKTAFPLC system The module has one high resolution analogue input for monitoring e g pressure UV monitor signals or other signals available as voltage outputs 1 3 9 Superloop Superloop permits the introduction of larger volumes of sampl
53. e 1 150 ml onto the column AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Introduction qu u S77 13 10 Extra rack panel Theextra rack panel is positioned on the vertical rods in thefree space between the lower and upper part of the system rack when the upper part is raised to a higher position Locking screws E O n h Extra rack panel for mounting gt ERE optional equip Ms MS ment 2 Ole El Ee E Ie KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 15 ED Introduction 16 gt gt gt gt e The components are designed for indoor use only Do not use in a dusty atmosphere or close to spraying water O perate 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
54. e UniN et cableis supplied with the module The valve can be connected anywhere between the system pump P 920 and the termination plug connected to the injection valve Note Thepower on LED on AD 900 flashes for a few seconds when connecting the U niN et cable The light gets steady when the communication with UNICORN is established Attach the module to the system rack Check that all electrical connections have been made correctly 15 3 Operation 15 3 1 Preparing for operation 1 Start your AK TAFPLC optional configuration system as described in section 2 6 Preparation for use M akesurethat AD900 is selected in the Components list Check that AD 900 isindicated in theflow schemein UNICORN A steady light from the green power on LED on AD 900 indicates that the communication is established Verify that the moduleis functioning by running the system pump manually and checking that a signal is shown in theRun Data pane 15 3 2 Using AD 900 for monitoring an external signal The AD 900 instructions are designated AD900 in the flow scheme In the run data window they are designated AD900 To use AD 900 for peak fractionation in a method instead of using the UV measurement in M onitor UPC 900 1 4 In theMethod Wizard select a fractionation option that includes Peak Fractionation In the Peak Fractionation dialog select AD900 from the Peak Identification menu Type suitable values in AD900 Level Peak Fractiona
55. e instruction under System Control Manual Flowpath AutosamplerControl Note If the syringe holder is not fully tightened i e no gap is obtained the plunger tip moves forcefully into the syringe end which can damage the syringe KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 39 EB A 900 Fill the wash solvent bottle with 2096 ethanol in water If the sample contains large amounts of salt or if the sample is very sensitive to ethanol use water as wash solvent instead degas and change the water every day Screw the bottle into the wash solvent holder and placethe holder on the A 900 Put the wash solvent tubing in the wash solvent bottle 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 5 4 1 Purging the syringe in A 900 1 40 M anually run the SyringeEnd and SyringeHome instructions alternately Repeat until no more air is left in the syringe more than 5 r
56. e port 1 marked pump Theend that is pre bent is connected to the AKTAFPLC injection valve V 1 4 Connectthe capillary marked G A2 H A2 F A2 I 21 15 m from the tubing kit between the A 900 injection valve port 6 marked column and the UV flow cell inlet Note This connection the dotted line is only temporary When the installation test is successfully performed G A2 H A2 F A2 is connected to the top inlet of the column and the capillary connecting the column outlet and theU V flow cell inlet is refitted 5 Check additional internal and waste tubing connections on A 900 according to the A 900 User M anual instructions 5 4 General preparation 1 Startyour AKTArPLC optional configuration system as described in 2 6 Preparation for use M ake sure that AutoSampler A 900 is selected in the Component list 2 Press the System Control button and select System Settings Specials 3 Check that the default sample loop volume 100 ul syringe volume 250 ul and needle height 2 mm are selected 4 Manually run the SyringeEnd instruction under System Control Manual Flowpath AutosamplerControl 5 Removethesyringefrom thesyringe valve M akesurethat the Luer connector in the valve remains in place 6 Fill thesyringe with 20 ethanol in water from a vessel 7 Replace the filled syringe Fully tighten the syringe so that a gap between theplunger tip and the syringe end isobtained after having executed the SyringeHom
57. e tubing using the SyringeEnd_905 and SyringeHome_905 instructions SyringeEnd_905 draws a syringe volume of wash solvent from the wash solvent bottle and fills the wash solvent tubing SyringeHome_905 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 905 instruction to perform a standard wash All tubing connected to the syringe valve will be filled and flushed with wash solvent 6 3 1 Purging the syringe in A 905 1 3 M anually run the SyringeEnd 905 and SyringeHome 905 instructions alternately Repeat until no more air is left in the syringe more than 5 repeats can be necessary Finish by performing the InitialWash 905 instruction 6 3 2 Checking the needle height in A 900 Refer to the Autosampler A 905 for KTA User M anual 52 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Autosampler A 905 for AKTA 6 6 4 Operation The parameters for controlling Autosampler A 905 are set in the Sample Injection dialog in the M ethod Wizard when creating the method The A 905 can be used for automatic sample injection from 1 0 ul to 1 ml To achieve optimal performance selection of sample loop volume syringe size and speed plus the washing method between injections must be considered Three different injec
58. eck all connections 9 Start up AK TAdesign 10 Select the component s in System Setup 11 Test the function of the optional equipment manually 12 Create a new method with the Method Wizard and make the appropriate selections to support the optional configuration If the component is not supported by the Method Wizard insert the required 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 verify the optional configuration in the new or modified method 15 Correct any faults and store the method Reverting to standard configuration 1 Flush AKTAdesign with distilled water 2 Disconnect the system in System Control 3 Switch off KTAdesign 4 Disconnect equipment to be removed from the UniNet chain and reconnect the UniNet chain for the standard configuration 5 Remove capillaries and reconnect the system flow path to the standard configuration Check all connections Start up KTAdesign Select a method supporting the standard configuration 9 Set up parameters to suit your needs 10 Perform a test run with distilled water to verify the standard configuration in the selected method Amersham e Biosciences TC information ab Uppsala Printed in Sweden by TK i Uppsala AB
59. efer to section 12 Sample pump P 960 and sample valve 90 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Air sensors ety A 13 2 5 Capillary connectors Thefollowing capillary connectors are recommended for connecting the liquid tubings to the air sensors Airsensor Air 925 and Air 925N ES Connector for 3 16 o d tubing no 18 1112 49 i with ferrule 18 1112 48 Da Connector for 1 8 o d tabing no 18 1121 17 Eg E with ferrule 18 1121 18 Connector for 1 16 o d nO no 18 1127 07 El with ferrule 18 1127 06 Airsensor Air 912 and Air 912N m E gm Fingertight connector for 1 16 o d tubing ESP 2 no 18 1112 55 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 91 13 Air sensors 13 Operation 13 3 1 Preparing for operation 1 Sh 0 N 92 Start your KTAFPLC optional configuration system as described in 2 6 Preparation for use M ake sure that Airsensor 1 and or 2 and or 3 is selected in the Components list In System Control select System Settings Click the Monitors radio button In the Instruction field select e g AirSensorl In the Parameters field select High Sensitivity for the air cell s to be used Select the flow source for AirSensor1 3 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 xp100Frac Monitors
60. en 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 AKTAFPLC standard system configurations KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 9 ER Introduction 10 O ptional configurations are shown in theflow 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 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 implem
61. enser aspirates a flush volume of sample into the sample line The A 900 injection valveis 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 W hen the sample loop sizeis changed theloop volume setting must be changed in System Settings 100 ul is the default loop volume After each injection the needle is rinsed f WashExt NO is selected in the method the needle is only washed internally f 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 reducethe amount of the flush volume The air segment is not injected 5 5 3 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 After each injection the needle is rinsed f WashExt NO is selected in the method the needle is only washed internally f 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 A
62. entation 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 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 M anual and or Instruction for the respective component as well as the System M anual for your AKTAFPLC system This manual is divided into a number of main sections reflecting the optional configurations available for AKTAFPLC system Select the main 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 AKTAFPLC system AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Introduction qu 1 3 Summary of optional configurations An overview of all optional configurations available for KTAFPLC is summarized below Optional configurations supported by AKTA
63. epeats can be necessary Finish by performing the InitialWash instruction AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Autosampler A 900 ea E 54 2 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 Putavial with a flat bottom filled with e g water in the tray segment directly below the needle arm Thebottom of the vial must be clearly visible 3 Perform a manual injection by issuing the Manual Flowpath InjectionFlushed instruction in System Control 4 Check visually that the needle stops approximately 2 mm above the vial bottom 5 If not go to System Settings Specials 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 T he 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 A djust the final setting in small steps until a needle height suitable for the actual use is obtained type of vial sample volume available etc AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 41 EB A 900 42 5 5 Operation 5 5 1 General The parameters for controlling A utosampler A 900 are set in the Sample Injection dialog in the M ethod Wizard when creating the method The A 900 can be used for automatic sample injection from a few
64. h Cooling eene 5 3 Inistallatiori terrere re rad 5 4 General preparation sss 5 5 Operations sacre tede e ro p P iUe RS Autosampler A 905 for KTA 6 1 General nette eee e inden c 62 Installation talar elem tte 6 3 General preparation sss 6 4 Operation irme tene cete teet Denn Outlet valve Tal Installation 7 2 Operatioti sace evt re ee eee AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 10 11 12 13 14 15 16 Column selection valves Bile Installation 56 9 2 Operation etica 57 Buffer selection valves 9 1 Installation 5 2 oc acn er certc eric nee ne eerte 58 9 2 Operation ei petet irr CLIE Re d eResps 59 General function valve LOD Installation ttt Pet aa 60 10 2 Operation aine Adda et tette 61 pH flow cell and pH electrode mounting III General iiit etd b rr te ED dente 63 112 Installatior ct reri t ent ee te nada rad 63 TAPS Operation e eerte iaa 65 Sample pump P 960 and sample valve 12 1 General eR tA 66 12 2 Installing the components seem 68 12 3 Loading a sample loop sss mm 70 12 4 Direct loading onto the column seseeeeees 76 12 5 Loading a Superloop sssssseee 82 Air sensors 13 1 Generando vir tr enc d ecc ad P dra 87 13 25 Installation csi re cio ns ra n a rre Ye ens 88 13 3 Operation cnc Ue ne pe HH eet e
65. hod Editor Three sample application techniques can be performed by using Pump P 960 in the AKTAFPLC system Loading a sample loop e Direct loading onto the column Loading a Superloop Each technique requires a specific configuration and different procedures to remove air from the sample flow path This is described in the following sections Control of the sample pump can be achieved automatically from a method or manually via the functions availablein UNICORN Note Make sure that any air trapped in the sample flow path is removed before use Presence of air in the sample pump might cause pressure pulsation and inaccurate flow AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Sample pump P 960 and sample valve c SEM Sample pump P 960 kit includes the items needed for upgrading an AKTArPLC system for Direct loading onto the column with Pump P 960 Thefollowing items are included in Sample pump P 960 kit e Pump P 960 UniNet cable Holders for KTA design systems Flow restrictor FR 902 Purgekit e Tefzeltubing i d 1 0 mm 3m e PEEK tubing i d 0 75 mm 2m Finger tight connectors and unions qx AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 67 ET Sample pump P 960 and sample valve 12 2 Installing the components 68 CAUTION The mains power to AKTArPLC must be switched OFF before disconnecting or connecting the UniNet 2 cables Th
66. ilable This makes the instruction suitable for controlling flow through fractionation before elution starts The properties are designated V4 Outlet in the run data window 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 Installingan outlet valvein theflow path affects thesystem delay volume See section 17 Configuration dependent delay volume for instructions on how to re calculate and change the delay volume KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 55 ED Column selection valves 8 Column selection valves 8 1 Installation Column Selection On line filter Injection Valve V1 Restrictor Fraction Collector Selection Valve V3 CAUTION Make sure KTAFPLC is switched off before installing the optional components The mains power to KTAFPLC must be switched OFF before disconnecting or connecting the UniNet 1 and UniNet 2 cables 1 Mark the two PV 908 valves with 2 and 3 Check that the ID code switch is set to 2 and 3 respectively 2 Mount the brackets and attach the two valves to the system rack 3 Cutand mount the new i d 0 50 mm capillaries M ount one capillary between port 1 on column valve 2 and
67. inimize the length of the capillaries used to connect the optional components to the rest of the system M any 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 mounting bracket with the two screws Removing components Capillaries are connected using unions as specified in chapter 19 Accessories and consumables 18 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Installation guidelines En All valves have a unique ID code which identifies them to the UNICORN control system at system start up ThisID code should be checked before installation ThelD 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 mn Sample injection INV 907 InjectionValve PV 908 ColumnPosition combined instruction P308 controlling both valves N Column selection Column selection Ww 5 Outlet fractionation PV 908 OutletValve OutletFractions two instructions used for controlling different type of fractionation functions Buffer selection A 5 IV 908 BufferValveA Buffer selection B 6 1V 908 BufferValveB Optional function 7 INV 907
68. ion Start your KTA FPLC optional configuration system as described in 2 6 Preparation for use M akesurethat Auxiliary Equipment is selected in the Components list Four digital input and four digital output signals can be handled via the P 920 REMOTE connector The input signals are designated AuxIn1 4 and the output signals are designated AuxOut1 4 14 2 2 Instructions for signal handling To set up signal handling in a method thefollowing parameters and instructions must be inserted manually in a method AuxOut1 4 To set up and usethe output signals do as follows 1 In System Control select System Settings Exp100Frac Specials Instructions E Xi m Instructions AuxDutl Parameters C Alarms Press D 0 000 sec a Signal 0 1 Keyboard d Mode KeyLocked cd Specials Out C Monitors C Curves Signal 1 I Set Selected Parameter To Strategy Default Value JL Cancel Help 2 Click on the Specials radio button 3 In the Instruction field select the signal you want to use e g AuxOut1 4 Check in the external equipment User M anual which logical value 0 or 1 that corresponds to inactive mode 5 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 6 Repeat this for all the output signals you want to use AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 97 1
69. ir is recommended to reducethe amount of the flush volume The air segment is not injected KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 43 EB A 900 44 5 5 4 InjectionPick up The InjectionPickup instruction selects the ul pick up injection modefor A 900 The syringe dispenser aspirates transport liquid from the transport vial into the sample line The A 900 injection valveis 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 After each injection the needle is rinsed f WashExt NO is selected in the method the needle is only washed internally f WashExt YES is selected in the method the needle is also washed 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 5 5 5 lInitialWash ThelnitialWash 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 5 5 6 SyringeHome End The SyringeHome En
70. is section describes how to install the components in the system To connect the capillaries see the section describing that specific sample application method L oading a sample loop Direct loading onto the column or Loading a Superloop 12 2 1 Installing Pump P 960 1 Assemble Pump P 960 according to the instructions given in the Pump P 960 User M anual 2 Attach the sample pump to the system rack 12 2 2 Installing IV PV 908 as sample valve if used 1 Assemble the sample valve see thelV PV 908 Instructions 2 Install the valve as a General function valve see chapter 10 and attach the valve to the system rack 12 2 3 Installing UniNet 2 cables Connecting P 960 only 1 Removethetermination plug from the last component in the UniN et 1 chain 2 Connect anew cable supplied between this component and Pump P 960 the leftmost socket Note Thetermination plug is not required P 960 has an internal termination Connecting P 960 and valve IV PV 908 1 Removethetermination plug from the last component in the UniN et 1 chain 2 Connect a new cable supplied between this component and valve IV PV 908 3 Connect a new cable supplied between valve IV PV 908 and Pump P 960 the leftmost socket Note Thetermination plug is not required P 960 has an internal termination AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Sample pump P 960 and sample valve c 12 2 4 Installing Flowrestrictor FR
71. kpressure 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 U PC 900 User M anual The pH flow cell should be mounted directly to the right of the optical unit KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 63 11 pH flow cell and pH electrode mounting Mixer 64 On line filter Injection valve V1 Fraction collector Flow restrictor New capillary of Male male union 90 mm 3 Cutanew capillary of 90 mm 0 50 mm i d and connect the outlet the screw head end from the conductivity flow cell to theinlet on the pH flow cell lower connection marked IN Connect theoutlet from the pH flow cell marked OUT to theinlet on the FR 902 marked IN using the male male union Connect the pH electrode BN C connector to the pH Probe connector on the rear of M onitor 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 Check that all capillary and electrical connections have been carried out correctly Note When usingthepH electrode the flow restrictor FR 904 must be replaced with the supplied flow restrictor FR 902 O therwise the long term stability and lifetime of the pH electrode will deteriorate AKTAFPLC Optional Configurations User M anual 18
72. lected in the Component list 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 UN ICORN as follows n System Control select Manual FlowPath Select instruction Valve X where X D code for the valve Set the valveto position 3 corresponds to the WASTE position in valve IN V 907 Manually run Pump P 920 with distilled water at a flow rate of approximately 2 ml min and check that water is coming out from port 3 on the valve corresponds to port 5 in valve IN V 907 10 2 2 Using the general function valve in a run 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 1 Create a new method by selecting File Method Wizard in the Method Editor 2 Select View Text instructions to display the text instruction editor Double click on the instruction block where you want to add the general function valve instruction H ighlight the instruction below which you want to insert the valve instruction 3 Click the Flowpath radio button KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 61 EU General function valve 4 From the Instructions list select the valve you intend to use Valve7 Valve8 or Valve9 The valve instruction paramete
73. ll capillary and electrical connections have been carried out correctly 4 3 Operation 4 3 1 Preparing for operation 1 Start your KTAFPLC optional configuration system as described in 2 6 Preparation for use Note Ifyou have upgraded from Frac 901 your new Frac 950 is used in methods in very much the same way as the previously used Frac 901 H owever 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 n System Control select Manual Frac 3 Check out the instructions available for Frac 950 to familiarize yourself with the new functions available R efer also to the Fraction collector Frac 950 U ser M anual to learn more about using Frac 950 4 3 2 Fractionation principles and definitions Fraction collector Frac 950 is used for flowthrough fractionation before the elution and for collecting fixed volumes and or peak fractions during elution Fixed volume fractionation and peak fractionation can have different fractionation sizes If both are activated simultaneuously the size set for peak fractionation is valid for collecting fractions during peaks and the size set for fixed volume fractionation is valid for collecting fractions during elution before between and after peaks The instruction Fractionation starts fixed volume fractionation The instruction PeakFractionation starts peak fractionation
74. ll the sample inlet tubings in port 1 7 1 Put the chosen sample inlet tubings from port 1 7 in the sample valve into the sample tubes 2 Immerse the tubing from port 8 in the valve in a buffer vessel 3 Set the valve to any of the chosen sample inlet ports 1 7 Note The sample inlet tubings should be filled before filling the buffer tubing connected to port 8 4 Disconnect the connector fitted to injection valve V1 port 3 Connect the purge tubing to the same port ae UL d Purge tubing 5 Set theinjection valveV1 to LOAD 6 Draw sample with the syringe until the liquid level has passed through the sample valve i KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 79 12 Sample pump P 960 and sample valve 80 7 8 Switch the sample valve to the next sample inlet tubing to be filled Note If the syringe needs to be emptied switch the sample valve before removing the syringe to prevent sample from flowing back to the vessel Repeat step 6 and 7 for the remaining sample inlet tubings To fill the buffer inlet tubing in port 8 1 2 Set the sample valveto port 8 Draw buffer with the syringe until the liquid level has passed through the sample pump The check valves in the sample pump will prevent the liquid from being withdrawn when removing the purge tubing Disconnect the purge tubing Fit the original connector to the port Flush the sample flow path tubing with buffer to rem
75. lve in your configuration not shown here Note Connect stop plugs to non used ports in the sample valve Note If low sample flows are used x10 ml min sample inlet tubings connected to ports 1 7 can be replaced with Tefzel i d 1 0 mm tubing together with finger tight connectors The sample inlet tubing should be adjusted to a suitable length AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 71 ET Sample pump P 960 and sample valve 78 12 4 2 Preparing for operation 1 Start your AKTArPLC optional configuration system as described in 2 6 Preparation for use M ake sure that Sample pump P 960 and if required Valve V7 or Valve V8 or Valve V9 are selected in the Component list Check that the sample pump and the sample valve if included are indicated in the flow scheme window M ake sure the system is connected as previously described in section 12 4 1 Connecting the capillaries 12 4 3 Testing the pump and the valve Verify that the sample pump is functioning properly by issuing manual commands from UNICORN as follows Note The sample valve is designated Valve 7 in this instruction 1 10 11 Fill a container with distilled water and submerge the sample inlet capillary S1 or for example S7 if the sample valveis included in the container Set the injection valveto position WASTE Remove air from the sample flow path using the purge tubing according to section 12 4 5 In System Con
76. lve to port 8 buffer inlet 3 Settheinjection valveto LOAD 4 Runthepump at 5 ml min for 1 min If the pump fails pumping liquid remove the air using a purge T to section 12 3 5 Removing air by using a purge 12 3 5 Removing air by using a purge tubing If thereis plenty of air in the sample inlet tubings or if the sample pump is dry use a purge tubing to fill the tubings and the pump Procedure for system including IV PV 908 as sample valve This procedure describes how to first fill the sample inlet tubings and then the buffer inlet tubing To fill the sample inlet tubings in port 1 7 1 Put the chosen sample inlet tubings from port 1 7 in the sample valve into the sample tubes 2 Immerse the tubing from port 8 in the valve in a buffer vessel 3 Set the valve to any of the chosen sample inlet ports 1 7 Note The sample inlet tubings should be filled before filling the buffer tubing connected to port 8 4 Disconnect the connector fitted to the outlet port on the connection part of Pump P 960 i e before the pressure sensor Connect the purge tubing to the same port e ee 1 Purge tubing 5 Settheinjection valve V1 to LOAD 6 Draw sample with the syringe until the liquid level has passed through the sample valve KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 73 12 Sample pump P 960 and sample valve 74 7 8 Switch the sample valve to the next sample inlet tu
77. microlitres 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 are as follows InjectionFlushed The sample loop is completely quantitatively filled with sample resulting in extremely good reproducibility InjectionPartial The sample loop is partially filled with sample giving low sample loss and allowing programmable injection volumes njectionPickup After aspiration from the vial the sample volume is transported into the loop with transport liquid mobile phase from another vial resulting in virtually no sample loss M anual functions used for setting up and maintaining the A 900 are found in System Control Manual Flowpath AutosamplerControl Some of the available commands are as follows nitialWash RotateTray SyringeHome End NeedleHome Front The parameters for controlling Autosampler A 900 with Cooling are Set in System Control System Settings Specials desired temperature and in System Control System Settings Alarms alarm and warning levels These functions are described in the following sections AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Autosampler A 900 ea 5 5 InjectionFlushed The InjectionFlushed instruction selects the flushed loop injection mode for A 900 The syringe disp
78. nent list Check that the sample pump is indicated in the flow scheme window M ake sure the system is connected as previously described in section 12 5 1 Connecting the capillaries 12 5 3 Testing the sample pump Verify that the sample pump is functioning properly by issuing manual commands from UNICORN as follows 1 2 Fill a container with distilled water and submerge the sample inlet capillary S1 Set the injection valve to position INJECT WARNING OVER PRESSURE The injection valve must be set to position INJ ECT in the Loading a Superloop configuration If not the Superloop might rupture due to over pressure when running the sample pump resulting in injury 84 Remove air from the sample flow path using the purge tubing according to section 12 5 5 In System Control select Manual Pump Select the instruction SampleFlow 960 in the instruction list Set Flowrate to 5 ml min and click on Execute Check that the sample pump is delivering flow Stop the pump by clicking End in the System Control toolbar AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Sample pump P 960 and sample valve c u S 12 5 4 Removing air by running the sample pump If there are only small amounts of air in the sampleflow path remove the air as follows 1 Immersethe end of the sample inlet tubing in a suitable buffer 2 Settheinjection valve to INJECT position INJ ECT in the Loading a S
79. nfigurations User M anual 18 1174 46 Edition AA Buffer selection valves ex 9 2 Operation 9 2 1 Preparing for operation 1 Start your AKTArPLC optional configuration system as described in 2 6 Preparation for use M ake sure that BufferValveA V5 and BufferValveB V6 are selected in the Component list 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 Buffer Valve A and Buffer Valve B are functioning properly by issuing manual commands from UNICORN as follows n System Control select Manual FlowPath Select instruction BufferValveA Set BufferValveA to position A4 Manually run pump P 920 with distilled water at a flow rate of approximately 2 ml min and check that water is drawn through port 4 in BufferValve A f applicable repeat these steps for Buffer Valve B Using the buffer selection function in a run To usethe buffer selection valves in a method select the desired valve options in the M ethod Wizard when creating the method The buffer valve instructions are designated BufferValveA and BufferValveB in the flow scheme In the run data window they are designated V5 BuffA and V6 BuffB KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 59 ED General function valve 10 General function valve 10 1 Installation To install any optional valve proceed as follows
80. ng the column selection function in a run To usethe column selection valves in a method select the desired valve option in the M ethod Wizard when creating the new method The column selection valves are designated ColumnValves in the flow scheme and V2 ColPos and V3 ColPos in the run data window KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 57 ED Buffer selection valves 9 Buffer selection valves Mixer L Injection 9 1 Installation On line filter p Pump A Pump yo 9 9 E Buffer Valve A V5 Buffer UU Valve B V6 we Wi ia RATES VUUUUUUUO Bi B2 B3 B4 B5 B6 B7 B8 58 Column Restrictor UV COND Fraction Collector v w3 CAUTION Make sure KTAFPLC is switched off before installing the optional components The mains power to AKTAFPLC must be switched OFF before disconnecting or connecting the UniNet 2 cables M ark the IV 908 valves with A and B Check that the ID code switches are set to 5 and 6 respectively M ount the brackets and attach the valves to the system rack Cut and mount the required teflon tubings i d 1 6 mm Use the attached UniN et 2 cablesto connect the valveto the UniN et 2 communication link The valve can be connected anywhere between the system pump P 920 and the termination plug connected to the outlet valve correctly Check that all capillary and electrical connections are carried out AKTAFPLC Optional Co
81. ngertight female M6 male Qutlet connection Inlet connection 82 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Sample pump P 960 and sample valve c On line filter Mixer p Pump A Pump B 9 9 Column Injection Valve V1 UV COND Fraction Collector Restrictor Superloop Capillary max length Connector Connection points 1 Tefzel i d 1 0 mm x 130 cm Fingertight connector 1 16 18 1112 55 Sample pump inlet Sample inlet tubing Union fingertight female M6 male 18 1112 57 2 Tefzel i d 1 0 mm x 50 cm Fingertight connector 1 16 18 1112 55 Sample pump outlet Fingertight connector 1 16 18 1112 55 Injection valve port 3 3 Tefzel i d 1 0 mm x 130 cm Fingertight connector 1 16 18 1112 55 Injection valve port 4 4 Tefzel i d 1 0 mm x 10 cm Fingertight connector 1 16 18 1112 55 Injection valve port 5 Fingertight connector 1 16 18 1112 55 Flow restrictor inlet 5 Tefzel i d 1 0 mm x 120 cm Fingertight connector 1 16 18 1112 55 Flow restrictor outlet SSS AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 83 ED Sample pump P 960 and sample valve pun 1 12 5 2 Preparing for operation Start your KTAFPLC optional configuration system as described in 2 6 Preparation for use M ake sure that Sample pump P 960 is selected in the Compo
82. omputer 48 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Autosampler A 905 for AKTA 6 4 Connecta new UniN et 1 cable supplied with the A 900 from the other UniN et 1 connector on the A 900 to the computer The UniN et 1 connection to the computer must be made to the board with four green LED s Four green LEDs 5 Check that the voltage setting of the A 905 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 V usetwo 5 AT fuses slow For 220 240 V use two 2 5 AT fuses slow CAUTION DO NOT switch on any of the equipment yet 6 When thevoltage selection and fuses are correct for thelocal mains power source plug in the supplied power cord in a freeoutlet in the base platform of the AKTAFPLC separation unit KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 49 6 Autosampler A 905 for KTA E 6 2 3 Capillary connections The capillaries to be connected between A 905 and the KTA FPLC separation unit are included in AKTA XT Tubing kit no 18 1122 92 Theflow diagram below shows the location of A 905 in the KTAFPLC flow path Autosampler Injection valve For Installation test Col
83. onnected as previously described in section 12 3 1 Connecting the capillaries 12 3 3 Testing the pump and the valve Verify that the sample pump is functioning properly by issuing manual commands from UNICORN as follows Note The sample valve is designated Valve 7 in this instruction 1 8 9 Fill a container with distilled water and submerge the sample inlet capillary S1 or for example S7 if the sample valveis included in the container Set the injection valveto position LOAD Remove air from the sample flow path using the purge tubing according to section 12 3 5 In System Control select Manual Flowpath If the sample valve is included select the instruction Valve7 in the list and select 7 Click on Execute Select Manual Pump Select the instruction SampleFlow 960 in the instruction list Set Flowrate to 0 5 ml min and click Execute 10 Check that the sample pump is delivering flow through its outlet 11 Stop the pump by clicking End in the System Control toolbar Thesample valve is designated V7 in this instruction 72 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Sample pump P 960 and sample valve c A 12 3 4 Removing air by running the sample pump If there are only small amounts of air in the sampleflow path remove the air as follows 1 Immersetheend of thesampleinlet tubing in a suitable buffer inlet S8 if using a sample valve 2 If using a sample valve set the va
84. or 9 depending on thelD code for the valve used This variable allows you to turn the position of the valve to the port 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 Valvex_pos to the desired position 8 Save the method ee 62 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA pH flow cell and pH electrode mounting 211 11 pH flow cell and pH electrode mounting 11 1 General 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 electrodeis to be removed from the system e g for storage ThepH electrode contains a sealed A g A gCl reference which cannot be refilled an internal electrolyte bridge of 4 M KCI saturated with Ag AgCl an outer electrolyte bridge of 1 M KN O3 an annular ceramic reference junction and a low profile pH membrane This is commonly known asa sealed combination doublejunction type The pH electrode is delivered with a transparent cover Note Theflow cell should not be used with any other pH electrode Note Always usethe 0 50 tubing kit and flow restrictor FR 902 when the flow cell is mounted 11 2 Installation CAUTION Never use 0 25 mmi d tubing blue when the pH flow cell is mounted in the system The electrode may rupture due to high bac
85. ove any trapped air bubbles and rinsethe pump according to the procedure in section 12 4 4 Removing air by running the sample pump Procedure for system not including IV PV 908 as sample valve 1 Put the sample inlet tubing in a buffer vessel Disconnect the connector fitted to injection valve V1 port 3 Connect the purge tubing to the same port Set the injection valve V1 to LOAD Draw buffer with the syringe until the liquid level has passed through the sample pump The check valves in the sample pump will prevent the liquid from flowing back when removing the purge tubing Disconnect the purge tubing Fit the original connector to the port Flush the sampleinlet tubing with buffer to remove any trapped air bubbles according to the procedure in section 12 4 4 R emoving air by running the sample pump Gently move the sample inlet tubing to the sample vessel AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Sample pump P 960 and sample valve c 12 4 6 Preparing a method for direct loading onto column To usethe sample pump for direct loading onto the column select Sample Pump Direct Loading P 960 as sample application techniquein the M ethod Wizard when creating the new method To use an additional valve as sample valve for example Valve 7 add the Valve7 instruction to the method in the M ethod Editor The M ethod Wizard does not support the sample valve function Thesample pump properties are de
86. ow rates the drops will turn to a continuous liquid stream 26 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Fraction collector Frac 950 EN 4 Fraction collector Frac 950 4 1 Generals Standard mode Prep mode Frac 950 can be used for both small scale standard mode and preparative scale prep mode purifications with AK TAFPLC Up to 392 fractions can be collected in time or volume base as fixed volume and or peak fractionations Thefractionation order can be selected as serpentine row row by row serpentine column or column by column For standard mode four sizes of tube racks for tube diameters of 12 18 and 30 mm tubes and 96 well microplates are available For prep mode two sizes of racks for tube diameter of 30 mm tubes and 250 ml bottles are available A Funnel to flask kit for transporting the fractions to other vessels is also available Using Frac 950 in prep mode requires a dispenser arm specially made for the prep mode racks 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 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 27 E Fraction collector Frac 950 m I 4 2 28 Installation CAUTION Make sure KTAFPLC is switched off before installing the optional components The mains p
87. ower to KTAFPLC must be switched OFF before disconnecting or connecting the UniNet 1 and UniNet 2 cables 1 On Unpack and assemble Frac 950 according to the instructions given in the Fraction Collector Frac 950 User M anual f 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 Disconnect the mains cable from Frac 901 Disconnect the UniN et 1 cables from Frac 901 Disconnect the capillary from the flow restrictor output in AKTAFPLC Remove Frac 901 from the workbench Put Frac 950 on the workbench to the right of KTAFPLC Note Frac 950 requires slightly more space than Frac 901 line filter Injection valve V1 Flow restrictor Fraction collector Frac 950 W3 Locate the capillary kit included with Frac 950 and decide if you want to include the accumulator in Frac 950 or not in prep mode you always use the accumulator According to your choice connect a capillary from the flow restrictor output in AKTAFPLC to Frac 950 as described in the Frac 950 User M anual AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Fraction collector Frac 950 EN Connect the supplied waste tubing to the waste outlet on Frac 950 and route it to a suitable waste container 9 Connect the UniN et 1 cable and the termination plug to Frac 950 10 Connect the mains cable to Frac 950 11 Check that a
88. p P 960 kit including Pump P 960 1 A 18 6727 00 UniNet cable 0 7 m holders for AKTAexplorer AKTApurifier AKTAbasic and AKTArPLC flow restrictor purge kit unions connectors and tubing Sample valve kit including 1 A 18 1175 86 Valve PV 908 UniNet cable 0 7 m unions connectors and tubing Air sensor Air sensors Air 912 1 2 mm 1 A 18 1121 23 Air 925 2 5 mm 1 A 18 1121 24 Air sensor A 912N kit including 1 A 18 1175 84 air sensor holder for AKTAexplorer connectors and tubing Air 925N 1 A 18 1174 16 Air 900 control box including 18 1121 22 pump tubing and one UniNet 2 1 A cable Purge valve 1 A 18 1126 33 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Accessories and consumables EN Item Quantity A C Code no pack AD 900 AD 900 Analog Digital converter 1 A 18 1148 62 Superloops Superloop 10 ml 50 ml Superloop 10 ml complete 1 A 18 1113 81 Superloop 50 ml complete 1 A 18 1113 82 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 Superloop 150 ml complete 1 A 18 1023 85 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
89. port 1 column valve 3 Thisis the column bypass position used as default position for the valves in UNICORN 56 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Column selection valves 8 A 4 Usethe attached UniN et 2 cables to connect the valves to the UniN et 2 communication link The valves can be connected anyw here between the system pump P 920 and the termination plug connected to the injection valve 5 Check that all capillary and electrical connections are carried out correctly 8 2 Operation Preparing for operation 1 Start your AKTAFPLC optional configuration system as described in 2 6 Preparation for use M ake sure that ColumnValve V2 and ColumnValve V3 are selected in the Component list 2 Thecolumn 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 column selection valves arefunctioning properly by issuing manual commands from UNICORN as follows n System Control select Manual FlowPath Select instruction ColumnPosition Set ColumnPosition to Position Manually run the pump P 920 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 valve3 and check that water is coming out from the center port of column valve 3 Usi
90. pport and tube holder 12 mm tube rack complete with bowl 1 A 18 6083 12 tube support and tube holder Microtiter plate 30 mm tube rack 1 A 18 6083 13 complete with bowl tube support and tube holder 30 mm tube rack standard mode 1 A 18 6083 14 complete with bowl tube support and tube holder 30 mm tube rack prep mode complete 1 A 18 6083 15 250 ml bottles rack complete 1 A 18 6083 16 Funnel to flask kit complete with tubing 1 A 18 6083 17 funnels tubing guide and extension legs Dispenser arm kit complete 1 A 18 6083 18 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 including 1 A 18 1114 50 mounting bracket AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Accessories and consumables EN Item Quantity A C Code no pack Autosampler A 900 Autosampler A 900 1 A 18 1116 61 Autosampler A 900 with Cooling 1 A 18 1144 61 Tray segment set 1 5 ml vial 1 A 18 1119 90 o d 12 mm Tray segment set 0 5 ml vial 1 A 18
91. ptional configuration system as described in 2 6 Preparation for use M ake sure that AutoSampler A 905 is selected in the Component list 2 Click the System Control button and select System Settings Specials 3 Check that the default sampleloop volume 1 ml syringe volume 1 ml and needle height 5 mm are selected 4 Run the AutosamplerControl 905 SyringeEnd 905 command in Manual Flowpath 5 Removethesyringefrom thesyringe valve M akesurethat the Luer x connector in the valve remains in place 6 Fill thesyringe with 20 ethanol in water from a vessel 7 Replace the filled syringe Fully tighten the syringe so that a gap between theplunger tip and the syringe end isobtained after having executed the AutosamplerControl 905 SyringeHome 905 command in Manual Flowpath Note If the syringe holder is not fully tightened i e no gap is obtained the plunger tip moves forcefully into the syringe end which can damage the syringe KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 51 6 Autosampler A 905 for KTA Fill the wash solvent bottle with 20 ethanol in water If the sample contains large amounts of salt or if the sample is very sensitive to ethanol use water as wash solvent instead degas and change the water every day Screw the bottle into the wash solvent holder and place the holder on the A 905 Put the wash solvent tubing in the wash solvent bottle Fill th
92. ries and consumables EN 19 Accessories and consumables 19 1 Connections chromatography standard selection Description and design Connector Fertule Connections Tubing o d Material Pressure cat no cat no colour limit bar 5 16 female M6 male 18 1127 76 UNF 5 16 32 PEEK black 100 M6 Fingertight female M6 male 18 1112 57 UNF 10 32 PEEK black 100 y A Fingertight connector 18 1112 55 UNF 10 32 1 6 mm PEEK black 250 1 16 male 1 16 5 glee Fingertight male M6 female 18 1112 58 UNF 10 32 PEEK black 100 M6 Mm Tubing connector for 18 1112 49 18 1112 48 UNF 5 16 24 4 8mm PEEK black 70 3 16 o d tubing 3 16 T E Tubing connector for 18 1121 17 18 1121 18 UNF 5 16 24 3 2 mm PEEK black 70 1 8 o d tubing 1 8 pes y ud Tubing connector for 18 1127 07 18 1127 06 UNF 5 16 24 1 6mm PEEK black 70 1 16 o d tubing 1 16 Union 5 16 female 18 1142 08 UNF 5 16 32 PEEK black 70 HPLC male UNF 10 32 gt De EE E KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 109 19 fuere and consumables PT 19 Optional components and consumables 110 Item Quantity A C Code no pack Fraction Collector Frac 950 Fraction collector Frac 950 complete 1 A 18 6083 00 with 18 mm 30 mm tube rack 18 mm 30 mm tube rack complete 1 A 18 6083 11 with bowl tube su
93. rloop select Sample Pump Loop Filling P 960 as sample application technique in the M ethod Wizard when creating the new method Note Do NOT select the S8 Prerinse Sample Loop 2 ml check box The sample pump properties are designated SampleFlow 960 and P960 Press in the flow scheme and run data windows lower than the maximum allowed pressure of the Superloop WARNING Make sure that the upper pressure limit of the system is 86 AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Air sensors 213 13 Air sensors 13 1 General Air sensors Air 912 Air 912N Air 925 and Air 925N can be used in the AKTAFPLC for detecting air in the system flow path or sample flow path Up to four air sensors can be used with the instruction names AirSensor1 AirSensor2 AirSensor3 and AirSensorP960 Air 912N and Air 925N The examples below show two possible locations Air sensors can be fitted in other locations in the AKTAFPLC system flow path The installation procedure is similar to the ones described in the following sections On line filter Mixer L Injection C Valve V1 S 2 AirSensor1 g E Fraction B Collector AirSensor1 in the inlet flow of Pump P 920 for checking the buffer supply KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 87 NED Air sensors On line filter Column Injection Valve V1 Fraction Collector Restrictor
94. rs allow you to change the position of the valves For IN V 907 only three positions can be selected see the Instructions of the valve t Method Editor UNTITLED File Edit Block View Help Dasa Gals Wi Main 00 Base CV 0 10 Column ml Any Block Start with PumpWash Explorer Block Flow Rate Block Column Pressure Limit Block Start Instructions Block BufferValve A1 Inlet Block Eluent A Inlet Block Eluent B Inlet Block Start Conc B Block Column Valve Column Valve 0 00 Base SameAsMain 0 00 ColumnPosition Position Bypass iiColumn_Position 0 00 End Block Block System Volume Compensation Block Column Equilibration Block AutoZero UV Block Aut PressureFlow Regulation Block Outlet Valve F2 Block Flowthrough Fractionation Block Sample Iniection Block Aut PressureFlow Regulation Block Flow Rate Block Wash ut Unbound Sample Block Stop Flowthrough Fractionation Block utlet Valve Reset Block Aut PressureFlow Reset Block Outlet Valve F2 Block Fractionation Block Flow_Rate Block Linear_Gradient Block Gradient_Delay Block Fractionation_Stop Click here to define a vari able name for the instruc tion parameter Instructions oa C Pump Ponien Var p il Et Feed ea Flowpath wa C MlemstMon onn C Frac Beplace C Watch C Other InjectionMark Delete Ready nom 4 5 Definea variable name for the instruction parameter e g Valvex pos where x 7 8
95. signated P960 Flow and P960 Press in the flow scheme and run data windows The valve properties are designated for example Valve7 in the flow scheme and run data windows Note Make sure that any air trapped in the sample flow path is removed before use KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 81 12 Sample pump P 960 and sample valve 12 5 Loading a Superloop Thesample 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 thetube into two separate chambers D epending on theflow direction the seal moves towards either end piece of the glass tube Superloop is available in three sizes 10 50 150 ml allowing application of 1 10 1 50 and 1 150 ml samples respectively When loading a Superloop a sample valveis usually not used 12 5 1 Connecting the capillaries 1 Makesurethat the sample pump is properly installed according to section 12 2 Installing the components 2 Cut the capillaries to suitable lengths and connect them using connectors and ferrules as indicated in thetable and in the figures Thelengths are maximum values 3 Check that all capillary and electrical connections have been carried out correctly Fingertight connector 1 16 Pump P 960 Union fi
96. ter port 2 Tefzel i d 1 0 mm x 50 cm Fingertight connector 1 16 18 1112 55 Injection valve port 3 Fingertight connector 1 16 18 1112 55 Sample pump inlet Union fingertight female M6 male 18 1112 57 3 Tefzel i d 1 0 mm x 130 cm Fingertight connector 1 16 18 1112 55 Injection valve port 5 4 Tefzel i d 1 0 mm x 10 cm Fingertight connector 1 16 18 1112 55 Sample pump outlet Fingertight connector 1 16 18 1112 55 Flow restrictor inlet port 5 Tefzel i d 1 0 mm x 120 cm Fingertight connector 1 16 18 1112 55 Flow restrictor outlet port 6 Tefzel i d 1 0 mm x 32 cm Fingertight connector 1 16 18 1112 55 Sample valve inlet S1 S7 7 Tubing conn 1 8 o d tubing 18 1121 17 Teflon i d 1 6 mm x 130 cm with ferrule for 1 8 o d tubing 18 1121 18 Sample valve inlet S8 Union 5 16 female HPLC male 18 1142 08 Only if a sample valveis installed Connect stop plugs to the non used ports in the sample valve AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 71 ET Sample pump P 960 and sample valve 1 12 3 2 Preparing for operation Start your AKTArPLC optional configuration system as described in 2 6 Preparation for use M ake sure that Sample pump P 960 and if required Valve V7 or Valve V8 or Valve V9 are selected in the Component list Check that the sample pump and the sample valve if included are indicated in the flow scheme window M ake sure the system is c
97. tion Volume and if no column is selected Minimum Peak Width Save the method when finished Note When adding the instruction in an existing method in the M ethod Editor insert it beforethe fractionation AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA o EN u 7777 The settings of the AD 900 filter can be changed in System Specials Autozero of the AD 900 can be performed in Manual Alarms amp Mon To set specific filter properties in an existing method 1 In the Method Editor select View Text Instructions to display the text instructions editor Double click on the first block in the method where the AD 900 instructions should be inserted Highlight an instruction The instruction will be inserted below this instruction Note Makesurethat theinstruction is inserted beforethe sample injection 2 Click on the Alam amp Mon radio button 3 From the Instructions list select AD900Filter Choose Type and Length and click Insert 4 Fromthelnstructionslist select AutoZeroAD900 and click Insert T he defined instruction is inserted in the method KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 101 ED Superloop 16 Superloop 102 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 might shatter
98. tion methods can be selected Flushed loop The sample loop is completely quantitatively filled with sample resulting in extremely good reproducibility better than 0 396 Partial loopfill The sample loop is partially filled with sample giving low sample loss and allowing programmable injection volumes Hl 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 TheA 905 uses a syringeto aspirate the sample from a microplate into the sample loop To prevent contamination of the syringe the A 905 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 the sample needle and also rinses both components KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 53 Outlet valve 7 Outlet valve 7 1 Installation On line filter Fraction Collector W3 F2 F3 F4 F5 F6 F7 F8 CAUTION Make sure KTAFPLC is switched off before installing the optional components The mains power to KTAFPLC must be switched OFF before disconnecting or connecting the UniNet 1 and UniNet 2 cables 1 MarkthePV 908 valve with 4 Check that thelD code switch is set to 4 2 Mount the bracket and attach the valve to the system rack 3 Cut and mount the new i d 0 50 mm capillaries 4 Usetheattached UniN et 2
99. tion to switch between multiple columns The valves are controlled synchronously by one common instruction OnePV 908 rotary valveused to divert between waste and seven positions to collect flow through or to collect seven large fractions e Oneor two IV 908 rotary valves for buffer selection Used to switch between different buffer solutions Threeoptional valves freely selected from IN V 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 1 3 5 Sample pump P 960 Sample pump P 960 is a single channel laboratory pump which can be connected to accomplish automatic sample application when using the same sample several times It can also be used to apply samples directly on the column or for filling sample loops including Superloop Pump P 960 uses plungers and check valves for transporting the liquid Its flow rate is controlled and the pump pressure monitored from UNICORN The sample pump is commonly used in combination with a multiport motorized valve PV 908 to enable automatic sample application of several different samples KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 13 ER Introduction 14 1 3 6 Air sensors The use and location of the air sensors is user defined For example e Oneair cell can be use
100. trol select Manual Flowpath If the sample valve is included select the instruction Valve7 in the list and select 7 Click on Execute Select Manual Pump Select the instruction SampleFlow 960 in the instruction list Set Flowrate to 1 ml min and click Execute Check that the sample pump is delivering flow through port 4 W 2 in the injection valve Stop the pump by clicking End in the System Control toolbar The sample valve is designated V7 in this instruction AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Sample pump P 960 and sample valve c u S 12 4 4 Removing air by running the sample pump If there are only small amounts of air in the sampleflow path remove the air as follows 1 Immersethe end of the sample inlet tubing in a suitable buffer inlet S8 if using a sample valve 2 If using a sample valve set the valve to port 8 buffer inlet 3 Settheinjection valveto WASTE 4 Runthepump at 40 ml min for 1 min If the pump fails pumping liquid remove the air using a purge tubing according to section 12 4 5 Removing air by using a purge tubing 12 4 5 Removing air by using a purge tubing If thereis plenty of air in the sample inlet tubings or if the sample pump is dry use a purge tubing to fill the tubings and the pump Procedure for system including IV PV 908 as sample valve This procedure describes how to first fill the sample inlet tubings and then the buffer inlet tubing To fi
101. umn Injection Valve V1 Fraction Collector 1 Removethe capillary between the injection valve V1 and the column top inlet 2 Select the capillaries matching the tubing kit to be used in AKTAFPLC after the column Use the capillaries marked G A1 and G A2 with the i d 0 25 mm tubing kit G A1 and H A2 with the i d 0 50 mm tubing kit and F A1 and F A2 with the i d 0 75 and 1 0 mm tubings Note See KTAFPLC System M anual for general recommendations on selecting column and tubing kit 50 KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA Autosampler A 905 for AKTA 6 E 3 Connect the capillary marked G A1 F A1 I 21 3 m from the tubing kit between the injection valve V1 port 1 and the A 905 injection valve port 1 marked pump The end that is pre bent is connected to the AKTAFPLC injection valve V 1 4 Connect the capillary marked G A2 H A2 F A2 21 15 m from the tubing kit between the A 905 injection valve port 6 marked column and the UV flow cell inlet Note This connection the dotted line is only temporary When the installation test is successfully performed G A2 H A2 F A2 is connected to the top inlet of the column and the capillary connecting the column outlet and theU V flow cell inlet is refitted 5 Check additional internal and waste tubing connections on A 905 according to the A 905 User M anual instructions 6 3 General preparation 1 Start your KTAFPLC o
102. ump 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 in 13 3 4 Setting up instructions for the air sensor function to monitor the AirSensor1 signal to suit your needs Note AirSensor2 and AirSensor3 are tested in a similar way 13 3 3 Testing AirSensorP960 located in the sample flow path 1 Makesurethat the sample pump is functioning properly 2 Confirm that air sensor ASP960 is shown in the flow scheme 3 Fill a sample container with distilled water and submerge the sample tubing 4 n System Control select Manual Pump in UNICORN 5 Select the instruction SampleFlow 960 in the instruction list 6 Set Flowrate to 5 ml min and click Execute 7 Whilethe sample pump is running check that the ASP960 symbol in the flow diagram is indicated with white colour po AS KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 93 13 Air sensors A 8 Liftthesampletubinginlet out of theflask and check that the white indication changes to red when air enters the ASP960 flow sensor 9 Submergethe sampletubing inlet and check that the red indication changes back to white
103. uperloop configuration If not the Superloop might rupture due to over pressure when running the sample pump resulting in injury WARNING OVER PRESSURE The injection valve must be set to 3 Run the pump at 40 ml min for 1 min If the pump fails pumping liquid remove the air using a purge tubing according to section 12 3 5 Removing air by using a purge tubing 12 5 5 Removing air by using a purge tubing If thereis plenty of air in the sampleinlet tubings or if the sample pump is dry use a purge tubing to fill the tubings and the pump 1 Putthesample inlet tubing in a buffer vessel 2 Disconnect the connector fitted to the injection valve port 2 Connect the purge tubing to the same port 3 Settheinjection valveto LOAD 4 Draw buffer with the syringe until the liquid level has passed through the sample pump The check valves in the sample pump will prevent the liquid from flowing back when removing the purge tubing 5 Disconnect the purge tubing 6 Fittheoriginal connector to the port 7 Flush thesampleinlet tubing with buffer to remove any trapped air bubbles according to the procedure in section 12 5 4 Removing air by running the sample pump 8 Gently move the sample inlet tubing to the sample vessel AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 85 ED Sample pump P 960 and sample valve A 12 5 6 Preparing a method for loading a Superloop To use the sample pump for loading a Supe
104. w to use air sensors in a method Example 1 In this example AirSensorP960 is used together with sample pump P 960 in a configuration designed to accomplish direct sample loading onto the column and checking the sample supply 1 In the Method Wizard select Sample Pump Direct Loading P 960 as sample application technique when creating the method 2 To enable the air sensor function check the Inject all Sample box 3 Typethemaximum allowed sample volume in the Maximal Injection Volume 0 20000 ml field Example 2 In this example AirSensor1 is used for checking the supply of buffer A 1 In the Method Editor select File Method Wizard and create a new method 2 Select View Text instructions to display the text instruction editor Highlight the instruction below which you want to insert the alarm air sensor instruction 3 Click the Alam amp Mon radio button and select Alarm_AirSensorl 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 4 Select Enabled and click Insert 5 Savethe method AKTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 95 14 iin equipment controlled via Pump P 920 REMOTE connector 14 Auxiliary equipment controlled via Pump P 920 REMOTE connector 14 1 Installation CAUTION Make sure AKTAFPLC is switched off before installing the optional components REMOT
105. wash out before elution fractionation a specified volume is collected Elution fractionation Fixed Volume Fractionation and or Peak Fractionation is selected in the Elution Fractionation dialog If fixed volume fractionation is selected a fixed volume size set in this dialog is collected during the complete elution If peak fractionation is selected only the peaks will be collected If fixed volume fractionation and peak fractionation is selected the fixed volumes will be collected before between and after the detected peaks Thepeak fractionation parameters are specified in the Peak Fractionation dialog Either the level or the slope of the UV curve or the signal level from AD 900 if used can be used for detecting a peak W hen using the UV curve level or the AD 900 signal level the level for starting and finishing the fractionation fraction size and minimum peak width if not specified a column should be specified When using the UV curve slope the slope value for starting and finishing the fractionation fraction size and minimum peak width if not specified a column should be specified KTAFPLC Optional Configurations User M anual 18 1174 46 Edition AA 31 5 e A 900 5 32 Autosampler A 900 5 1 General Autosampler A 900 is intended for automated multiple sample injections A sequence of valve switching and syringe dispenser controlled sample withdrawal enables the A 900 to inject sample volumes in a

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