GE HEALTHCARE AKTAprime plus Quick Reference Instructions

Also see for AKTAprime plus: Operating instructions

Contents

GE HEALTHCARE AKTAprime plus Quick Reference Instructions Manual pdf 16 page image

11-0027-48 AC, 2007-09 • p16GST-tagged protein purification1 Preparing the buffers• Use high purity water and chemicals.• Filter all buffers through a 0.45 μm filter before use.Binding buffer (port A1):20 mM sodium phosphate, 0.15 M NaCl, pH 7.3Elution buffer (port B):50 mM Tris-HCl, 10 mM reduced glutathione, pH 8.0Prepare at least 500 ml of each eluent.2 Preparing the samplea) Adjust the sample to composition of binding buffer by:• diluting the sample in binding buffer or• by buffer exchange using HiTrap Desalting or HiPrep26/10 Desalting.b) Pass the sample through a 0.45 μm filter.3 Preparing the systema) Place the inlet tubing from port A1 (8-port valve)in the binding buffer and the tubing from port B(2-port valve) in the elution buffer.b) Place the three brown waste tubings in waste.c) Connect the column between port 1 on the injectionvalve (7-port valve) and the UV flow cell (see Orderinginformation on next page for suitable columns).d) Fill the fraction collector rack with 18 mm tubes(minimum 10) and position the white plate on thefractionation arm against the first tube.e) Connect a sample loop large enough for your samplebetween port 2 and 6 on the injection valve. Use asyringe to manually fill the loop.Note: If a Superloop is needed, additional informationis supplied in the instructions for Superloop.GST-tag PurificationGSTrapSet Sample Inj. Vol(00.0 ml) 00.0Run Application TemplatePress OK to startRun data displayedApplication TemplateTemplates4 Selecting Application Template andstarting the methoda) Check the communication to PrimeView. At the lowerright corner of the screen the text Controlled By:prime should be displayed.b) Use the arrow and OK buttons to move in the menutree until you find GST-tag Purification GSTrap.c) Enter the sample volume and press OK to start thetemplate.Note: If a 5 ml column is preferred, see cue card onp.36.Theoretical gradient in GST-tag Purification GSTrap ApplicationTemplate.Re-equilibrationWashElutionPriming& Equilibration1005011 10 11 6 MinSampleTotal separation time = 37 min + sample application time%B