Packing AxiChrom 50 and 70 columns with ÄKTA avant 150
Contents
1. 2 Select System and Predefined Method Intelligent Packing in the dialog Click OK New Method System MDH P3 02 Create a new method by using the Predefined Method Intelligent Packing v Empty Method Method Description Packing AxiChrom columns with a predetermined column type by a flow of hydraulic liquid that pushes the adaptor down The user initiates the start of compression at the exact point when the adapter reaches the consolidated bed surface The adapter compresses the bed according to the packing factor or target bed height as selected Two column performance tests upflow downflow are automatically performed Result The phases included in the chosen method are displayed in the Method Outline pane to the left and the default settings for each of the phases are shown in the Phase Properties pane to the right AxiChrom 50 100 columns Addendum 28 9864 93 AA Step Action 2 In the Phase Properties pane of the Method Settings phase edit general settings like Column type and Method Base Unit UNICORN automatically calculates correct settings for volume flow rate and pressure limits based on the selected column type Note Only AxiChrom columns can be selected in order to be able to run the method The illustration below shows the Method Outline pane to the left and the Phase Properties pane of the Method Settings phase to the right Phase Properties T2. If the column hardware and medium are made by GE Healthcare select the name of the hardware and medium to have most of the parameters filled in automatically Show hardware types by diameter cr Show medium types by technigue GE Healthcare hardware type GE Healthcare medium type Ary Ary Run Parameters Details Ordering Information Parameters Technique Column volume ral Mas pre column pressure MPa Max delta column pressure MPa Default flowrate ml min Max flowrate ml min Default linear flowrate cmh Max linear flowrate omh Min pH value short tern Maz pH value short term Min pH value long term Maz pH value long tern Required information Global Personal Note The Intelligent Packing method is only compatible with AxiChrom columns AxiChrom 50 100 columns Addendum 28 9864 93 AA Step Action e Select the appropriate AxiChrom hardware in the drop down list GE Healthcare hardware type for the new column type To filter the drop down list to only show hardware types with certain diameters enter the diameter range in cm in the Min and Max fields for Show hardware types by diameter cm above e Select the GE Healthcare medium type for the new column type in the drop down list To filter the drop down list to only show medium types for a specific separation technique choose the appropriate technique in the Show medium types by technique drop do
3. 3 25 1 Hyakunincho Shinjuku ku Tokyo 169 0073 Japan 28 9864 93 AA 12 2010
4. sure that the settings are correct After the equilibration two Column Performance tests are run one downflow and one upflow Ensure that the Upflow check box is checked in one of the Column Performance Test phases The test will then be run upflow Ensure that the Upflow check box is unchecked in the other Column Performance Test phase and this test will be run downflow It is not necessary to run the tests but is recommended to ensure the quality of the packed bed A solution of either acetone or NaCl can be used to give a good indication of the column packing quality The eluate is monitored by measuring the UV absorption at 280 nm or the conductivity and the curve can be evaluated in Evaluation See Performance eveluation of the column in AxiChrom 50 70 and 100 columns User Manual for more information The illustration below shows the Method Outline pane to the left and the Phase Properties pane of the Column Performance Test phase to the right Phase Properties Text Instructions iT Method Settings Column Performance Test C Use the same flow rate as method settings Fl t 30 0 Hh 0 0 458 3 Intelligent Packing ea __ 300 E C Up flow y Equilibration Test note Packing test Down flow y Sample application technique Column Performance Test Manual loop filling Sample pump loop filling v Inject sample directly onto column Column Performance Test Note The sample pump will be washed with sample pr
5. the Intelligent Packing phase make sure that the GE approved packing settings radio button is clicked This button should be clicked if an AxiChrom column type with GE approved media was chosen in the Method Settings phase Default parameter values for packing will automatically be filled in into the Intelligent Packing phase The illustration below shows the Method Outline pane to the left and the Phase Properties pane of the Intelligent Packing phase to the right Phase Properties Text Instructions iT Method Settings Intelligent Packing GE approved packing settings Intelligent Packing Custom packing settinas Pack Factor 1 15 1 00 1 50 y GE Approved Media i h 0 00 120 00 Equilibration Adapter velocity 60 00 cmh h ee Flow rate cmh 0 0 458 3 Column Performance Test Time 1 00 min h Column Performance Test Seescima lds oh ae lected ethanol ao 40 80 Target bed height 20 cm Estimated time until bed contact warning 26 3 min Inlet for hydraulic chamber liquid Inlet for mobile phase Inlet A AQ Use the same inlets as in Method Settings Fill the system with the selected Inlet A l v hydraulic chamber liquid as x 7 i Inlet B B1 v 0 0 B 0 0 100 0 Column position for hydraulic chamber Position 2 v Note If the user wants to make changes in parameter values Custom packing settings can be used See Creating an Intelligent Packing method using custom media
6. 0 columns Addendum 28 9864 93 AA 13 Step Action q In the Phase Properties pane of the Equilibration phase make sure that the settings are correct The default flow rate is set to 30 cm h and the total volume of the Equilibration phase is 1 3 column volumes The illustration below shows the Method Outline pane to the left and the Phase Properties pane of the Equilibration phase to the right Phase Properties Text Instructions iT Method Settings Equilibration Reset UY monitor recommended if the equilibration occurs before the purification Intelligent Packing h C Use the same flow rate as in Method Settings Use the same inlets as in Method Settings Flow ate 30 0 cm h 0 0 458 3 Inlet At z InletB BI 00 B 00 100 0 y L s al Fill the system with the selected buffer Column Performance Test v Equilibrate until Column Performance Test the total volume is 1 30 CY O the following condition is met Conductivity greater than 000 mS em 0 00 1000 00 Accepted pH fluctuation 010 0 00 14 00 Accepted UV fluctuation m4U 0 00 6000 00 Accepted conductivity fluctuation mS cm 0 00 300 00 Signal stable for 7 00 min 0 02 1000 00 Maximum equilibration volume cy Note All parameters in this phase can be changed AxiChrom 50 100 columns Addendum 28 9864 93 AA Step Action 8 In the Phase Properties pane of the Column Performance Test phase make
7. GE Healthcare Life Sciences Addendum 28 9864 93 AA AxiChrom 50 70 columns Packing AxiChrom 50 and 70 columns with AKTA avant 150 1 Introduction This document is an Addendum to AxiChrom 50 100 User Manual 28 9331 08 and describes packing of AxiChrom 50 and 70 columns with AKTA avant 150 Specifications Compatible systems AxiChrom 50 and 70 columns are compatible with the KTA avant 150 system and integrated in UNICORN control software with a predefined method for Intelligent Packing Packing flow rates The table below describes the packing flow rates of columns connected to KTA avant Column Packing flow rateat30cm h Packing flow rate at 60 cm h AxiChrom 50 8 8 ml min 17 7 ml min AxiChrom 70 17 7 ml min 35 4 ml min 3 Connect AxiChrom columns to AKTA avant Introduction This chapter describes the basic configuration for connecting AxiChrom 50 and 70 columns to AKTA avant 150 Connection to AKTA avant 150 Connect the column to the system as described in the illustration in the Intelligent Packing phase and according to those column valves chosen in the method see Creating an Intelligent Packing method using GE approved media step 6 on page 13 Note Use a T connection on the outlet tubing Tubing used between column and system Column diameter mm KTA system Tubing i d mm KTA avant 150 KTA avant 150 2 AxiChrom 50 100 columns Addendum 28 9864 93 AA
8. The illustration shows tubing connections for AKTA avant 150 28 9888 89 Tubing kit AxiChrom 50 AKTA avant desk id 1 7 L 1200 x2 28 9888 92 Tubing kit AxiChrom 50 70 AKTA avant floor id 1 7 L 1600 x2 id 1 7 L 800 x1 id 1 7 L 1200 x1 id 2 9 L 1600 x1 id 2 9 L 1600 x1 id 1 7 L 200 x2 id 1 7 L 200 x2 AxiChrom 50 100 columns Addendum 28 9864 93 AA 4 Create column type Introduction Before creating the Intelligent Packing method in UNICORN a column type needs to be created with the Column Handling tool This type consists of e AxiChrom column hardware e Medium e Pressure flow parameters e Bed height For more information on Column Handling refer to the documentation for your UNICORN software Open the Column Handling dialog To open the Column Handling dialog e select Tools Column Handling in any of the UNICORN modules or e click the Column Handling icon in the Toolbar where available Ei AxiChrom 50 100 columns Addendum 28 9864 93 AA Create a new column type The table below describes how to add a new column type with the Column Handling tool Step Action 1 In the Column Type Parameters tab in the Column Handling dialog click New Result The New Column Type dialog opens New Column Type
9. e column type Result The Save As dialog opens Save As ad 6 Type in a Column type name and click Save Result The column type is saved in the database and displayed in the Column types list Note If traceability of result is requested a column individual can be created under column type for logging of the data see UNICORN 6 1 Method Manual AxiChrom 50 100 columns Addendum 28 9864 93 AA 5 Creating an Intelligent Packing method using AKTA avant 150 Introduction This chapter contains some additional information to the chapter Intelligent Packing of the column in the AxiChrom 50 70 and 100 columns User Manual where the Intelligent Packing wizard is presented for AKTApilot and AKTAexplorer This chapter describes how to create an Intelligent Packing method for AKTA avant 150 based on either GE approved media or custom media Creating an Intelligent Packing method using GE approved media This section describes the creation of an example Intelligent Packing method based on GE approved media If custom media is selected instead the method settings allow a wider range of variables to be defined to optimize the method Follow the instruction below to create an Intelligent Packing method based on GE approved media Step Action 1 Open the Method Editor module and click on the New method icon ye ns ign Result The New Method dialog opens AxiChrom 50 100 columns Addendum 28 9864 93 AA Step Action
10. ext Instructions IT Method Settings Method Settings Column selection Result Name amp Location Show by technique Intelligent Packing Start Protocol Intelligent Packing Start Protocol Protocol Column t j olumn type AxiChrom 50 300 20 um Capto Q 20 cm v EET y a Column Properties Column volume i 392 699 ml Equilibration Pressure limit pre column 1 00 MPa 0 00 5 00 s X I Unit selection Pi limit delta col 1 00 MPa 0 00 5 00 v tessure limit delta column af Rete Ree lear cy a Use flow restrictor Column Performance Test Flow Rate Unit cmh A Column position Position 1 a wv R A e Monitor settings Wavelengths 190 700 nm Column Performance Test F Flow rate 458 3 cm h 0 0 458 3 uvd 290 nm Control the flow to avoid overpressure Dl uv2 nm gus3 nm Use manually prepared buffers MRA Al J Inlet B BI E C Enable pH monitoring Use BufferPro automatic buffer preparation Enable air sensor alarm Recipe Inlet A Inlet B 5 C Sample inlet pH 2 8 5 4 recommended Conc M 0 050 0 100 Column Logbook Enable logging of Column Performance Test oO cP Make sure that all parameter values are correct Note Ifthe check box Column Performance Testis checked automatic logging of the result will be saved in the column logbook if a column individual is chosen AxiChrom 50 100 columns Addendum 28 9864 93 AA Step Action 4 In the Phase Properties pane of
11. ior to sample application Sample inlet Buffer v Sample volume 0 01 Cy Fill loop with mil Empty loop with mil Elution volume 1 40 Cy Note To check if the test is run upflow or downflow see the Upflow check box If this is checked the test is run upflow AxiChrom 50 100 columns Addendum 28 9864 93 AA 15 Step Action At both tests the flow rate is set to 30 cm h It is important to use a low flow and to use the same flow rate if tests are repeated to get comparable results 9 Click the Save the method icon fo Result The Save As dialog opens 10 In the Save As dialog e Select a target folder to enable the Save button e Type a Name for the method e Select a System from the list e Click the Save button Result The created method is saved in the selected folder Creating an Intelligent Packing method using custom media Intelligent Packing methods may also be used for custom media UNICORN facilitates this by providing an extended range of variables that may be modified to optimize the method For example it is possible to e Pack according to a desired Packing Factor or bed height If selecting the latter option the user stops the compression when the final bed height has been reached e Select flow conditioning with associated variables e Specify desired adapter velocity during packing within defined range In the Phase Properties pane of the Intelligent Packing phase make sure that the Cus
12. on page 16 AxiChrom 50 100 columns Addendum 28 9864 93 AA 11 12 Step Action In the Phase Properties pane of the Intelligent Packing phase click on the Slurry Recipe button to receive a calculation of the amount of medium to fill in the column based on the slurry concentration The recommended packing buffert for the selected medium is also shown in the dialog Slurry Recipe Result The Suggested Slurry Recipe dialog opens Suggested Slurry Recipe Prepare 1 1 1 slurry with 462 medium Exchange liquid in slurry to 0 4 M Nall and fill into the column Note The Slurry Recipe button is not available in Custom Packing settings Click OK to close the dialog AxiChrom 50 100 columns Addendum 28 9864 93 AA Step Action 6 In the Phase Properties pane of the Intelligent Packing phase click on the Column connection button to display an illustration describing how to connect the column to the system Column Connection Result The How to connect the column to the system dialog opens How to connect the column to the system Connect the column inlet to Position 14 on the column position valve and the column outlet to Position 1B and Position 2B with a T connection Connect the hydraulic chamber inlet to Position 24 on the column position valve Note The text will be updated dependent on the column positions chosen but the illustration will remain the same Click OK to close the dialog AxiChrom 50 10
13. ry priming and packing the column using AKTA avant Follow the instructions in the AxiChrom 50 70 and 100 columns User Manual to prepare the slurry prime the column pour slurry into the column and to pack the column The UNICORN pages may differ slightly for AKTA avant 150 but the concept is the same as for AKTAexplorer and AKTApilot 18 AxiChrom 50 100 columns Addendum 28 9864 93 AA For local office contact information visit www gelifesciences com contact GE Healthcare Bio Sciences AB Bj rkgatan 30 751 84 Uppsala Sweden www gelifesciences com axichrom imagination at work GE imagination at work and GE monogram are trademarks of General Electric Company AxiChrom AKTAexplorer AKTApilot AKTA and Drop Design are trade marks of GE Healthcare companies 2010 General Electric Company All rights reserved First published Dec 2010 All goods and services are sold subject to the terms and conditions of sale of the company within GE Healthcare which supplies them A copy of these terms and conditions is available on request Contact your local GE Healthcare representative for the most current information GE Healthcare Europe GmbH Munzinger Strasse 5 D 79111 Freiburg Germany GE Healthcare UK Limited Amersham Place Little Chalfont Buckinghamshire HP7 9NA UK GE Healthcare Bio Sciences Corp 800 Centennial Avenue P O Box 1327 Piscataway NJ 08855 1327 USA GE Healthcare Japan Corporation Sanken Bldg
14. tom packing settings radio button is clicked 16 AxiChrom 50 100 columns Addendum 28 9864 93 AA The illustration below shows the Method Outline pane and the Phase Properties pane of the Intelligent Packing phase Phase Properties Text Instructions iT Method Settings Intelligent Packing GE approved packing settings Intelligent Packing Custom packing settings y GE Approved Media Equilibration v Column Performance Test kd Selected medium Capto Q Target bed height 20 cm Column Performance Test Estimated time until bed contact warning 30 4 min Inlet for hydraulic chamber liquid Inlet A A2 v Fill the system with the selected hydraulic chamber liquid Column position for hydraulic chamber Position 2 v Column Connection Pack by Packing Factor Pack Factor 1 20 Pack to the target bed height Adapter velocity 50 00 C Flow conditioning Flow rate Time 0 00 Slurry Concentration as measured in 20 ethanol Inlet for mobile phase 1 00 1 50 cmh 0 00 120 00 0 0 cmh 0 0 458 3 min 41 41 80 Use the same inlets as in Method Settings Inlet Al v Inlet B B1 v 0 0 B 0 0 100 0 Make the appropriate changes and save the method in the same way as for GE approved packing AxiChrom 50 100 columns Addendum 28 9864 93 AA 17 6 Preparing the slur
15. wn list above Result The following parameters are automatically filled in can be edited if appropriate Run Parameters Details Parameters Value Unit Parameters Value Unit Technique Anion Exchange K Hardware diameter 5 0 cm Column volume mil Bed height cm Max pre column pressure 1 0 MPa Typical loading range ma Max delta column pressure MPa Total liquid volume t ml Default flow rate malin Void volume Yo ml Max flow rate ral rnin Typical peak width at base ml Default linear flow rate cmh Average particle diameter 90 0 Max linear flow rate cmh Molecular weight range Min pH value short term 2 Ordering Information Max pH value short term 14 Min pH value long term 2 Parameters Value Max pH value long term 12 Name Code number Medium name Capto Q Medium code number 17 5316 02 Hardware name AxiChrom 50 300 glass 20 um steel Hardware code number See user manual for AxiChrom Enter the remaining parameter values for the new column type in the Run Parameters Details and Ordering Information tabs Fields marked with must be filled in Values in the gray fields are calculated and automatically filled in based on entered values for the corresponding parameters Select whether the the new column type should be Global available for all users or Personal only available for the current user AxiChrom 50 100 columns Addendum 28 9864 93 AA Step Action Click Save As to save th
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