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B Multicolumn Purifications
Protein purification can involve challenging separations of complex mixtures that
might not provide sufficient resolution of target proteins from their impurities in a
single chromatographic step. Such cases may require multiple discrete experiments
involving multiple columns and different column chemistries.
Multicolumn purifications combine a series of columns, often with different or
orthogonal chemistries in sequence or in tandem, in a single separation pr
otocol.
Most common protein purification techniques can be incorporated into a single
protocol utilizing multiple columns, providing convenience through automation for
proteins purified on a routine basis. A typical example of a multicolumn
chromatographic scheme consists of an affinity purification step, via a histidine (His)
or glutathione-S-transferase (GST) tag, followed by one or more additional
experiments involving size exclusion columns for desalting or aggregate removal, or
ion exchange columns for separating complex mixtures.
To streamline the combination of such disparate chromatographic methods, an
automated chr
omatography system with a flexible design and versatile control
software is essential. With their flexibility and scalability NGC™ chromatography
systems can be easily customized to meet your multicolumn purification
requirements.
- Chromatography Systems 1
- Software 1
- User Guide 1
- NGC™ Chromatography 3
- Systems and 3
- User Guide 7
- User Guide 9
- Table of Contents 10
- User Guide 11
- 1 Introduction 13
- Main NGC Features 14
- User Guide 15
- NGC Chromatography Systems 16
- User Guide 17
- Finding Out More 18
- 2 The Workspace 19
- The Home Window 20
- User Guide 21
- User Guide 23
- The System Control Window 25
- The Method Editor Window 26
- The Evaluation Window 27
- 2 28
- Touch Screen Menu Commands 29
- Touch Screen Toolbar Commands 30
- 3 System Control 31
- 3 32
- Manual Menu Commands 34
- Tools Menu Commands 34
- Context Menu Commands 35
- Chromatogram View 37
- Table 1. Trace Definitions 39
- Trace Module Explanation 40
- Showing or Hiding Traces 41
- Changing Trace Color 42
- Changing the Y-Scale Values 43
- Autoscaling the UV Trace 43
- To add annotations 45
- To edit an annotation 45
- To delete an annotation 45
- User Guide 47
- Fluidic Scheme Pane 48
- User Guide 49
- Sample Inject Valve 51
- Sample Pump 52
- Column Switching Valve 52
- UV/Conductivity Monitors 52
- Signal Import Module 52
- Fluidic Scheme Configurations 53
- User Guide 55
- User Guide 57
- Working with Fluidic Schemes 59
- To change module settings 61
- Fluidic Scheme Mapping 62
- Color Label valve 63
- To unmap valves 64
- To cancel unmapping valves 64
- To map valves 65
- Calibrations 66
- User Guide 67
- Calibrating Pressure Settings 68
- User Guide 69
- To verify the plumbing path 70
- System Settings 71
- To set the delay volume 72
- Orange 0.02" (0.5 mm) 73
- Green 0.03" (0.75 mm) 73
- Clear 0.062" (1.6 mm) 73
- Control Flow Tab 74
- To control the flow rate 75
- Remote Access Tab 76
- Trace Settings Tab 76
- User Guide 77
- Device Input Tab 78
- User Guide 79
- Device Output Tab 80
- To disable the connection 81
- Air Sensors Tab 82
- To deactivate an air sensor 83
- Email Notifications Tab 84
- User Guide 85
- System Name Tab 87
- System Information 88
- Detector Tab 89
- Multi-Wavelength Detector 90
- Preferences 91
- User Guide 93
- 4 Performing a Manual Run 95
- User Guide 97
- User Guide 99
- Preparing the System 101
- User Guide 101
- User Guide 103
- Cleaning the System 104
- Running an Experiment 105
- User Guide 107
- User Guide 109
- To run an experiment step 110
- Changing Module Settings 111
- Stopping a Manual Run 111
- Clearing Run Data 111
- Saving a Manual Run 111
- Viewing Run Data 113
- 5 Method Editor 115
- Before You Begin 116
- Opening a Method 117
- 5 118
- File Menu Commands 119
- Edit Menu Commands 120
- View Menu Commands 121
- Help Menu Commands 121
- Toolbar Commands 121
- Phase Library Pane 122
- Adding a Phase to a Method 123
- Method Outline Pane 124
- Phase Parameters Pane 124
- Method Steps Pane 125
- To view step details 126
- Method Settings Parameters 127
- Column Selection 128
- User Guide 129
- To add user-defined columns 131
- Detector Settings 133
- Unit Selection 133
- Fraction Collection 134
- Buffer Selection 135
- Pump Inlets 136
- Pump + Inlet Valves 136
- Pump + Buffer Blending 136
- To select a buffer 137
- Phase Controls and Parameters 138
- User Guide 139
- Standard Phases 140
- User Guide 141
- User Guide 143
- Column Wash Parameters 145
- Elution Parameters 146
- Special Considerations 147
- NGC Scout Systems 148
- User Guide 149
- User Guide 151
- To clean the pH valves 152
- User Guide 153
- User Guide 155
- System Preparation Parameters 156
- Create New Phase Parameters 157
- Step Library Pane 158
- Scout Parameters Pane 159
- 6 Creating a Method 161
- Standard Method Templates 162
- User Guide 163
- 6 164
- User Guide 165
- User Guide 167
- To create a method 168
- To change the fluidic scheme 168
- To specify general settings 169
- User Guide 171
- Adding Phases 172
- Editing Methods and Phases 172
- Renaming Phases 173
- Rearranging Phases 173
- Deleting Phases 174
- User Guide 175
- To run a method 177
- To add runs to the run queue 177
- Saving a Method 179
- Renaming a Method 180
- Deleting a Method 181
- The Scouting Wizard 183
- User Guide 185
- User Guide 187
- User Guide 189
- Creating a Scouting Method 191
- User Guide 193
- Running a Scouting Method 194
- To run a scout method 195
- 7 Evaluating Results 197
- Managing Analysis Projects 198
- To open a saved analysis 199
- 7 200
- Analysis Menu Commands 203
- Tab Toolbar Commands 204
- Customizing the Chromatogram 205
- Selecting the Active Trace 206
- Showing or Hiding a Trace 207
- Changing the Axes 207
- Changing Trace Colors 208
- Zooming In and Out 209
- Chromatogram 210
- To resize the selected region 211
- Annotating the Chromatogram 212
- Copying the Chromatogram 213
- Changing Table Grouping 214
- Sorting Table Columns 215
- Copying a Table 216
- User Guide 217
- Managing Runs 218
- To close a tab 219
- To close all tabs 219
- Comparing Traces 220
- Stacked View 221
- Overlay View 223
- Hiding All Traces in a Group 224
- Showing All Traces in a Group 224
- To offset a trace 225
- Managing Analyses 226
- Copying an Analysis 227
- Renaming an Analysis 227
- Exporting Run Data 228
- Peak Integration 229
- Baseline Parameters 230
- Peak Parameters 231
- Peak Filtering 231
- Default Parameters 231
- Starting Peak Integration 232
- User Guide 233
- Changing Peak View Options 234
- User Guide 235
- Showing or Hiding Columns 237
- Copying the Peaks Table 238
- User Guide 239
- Adjusting Peaks Manually 240
- To add a peak 241
- Saving Peak Integration Data 242
- Evaluating Fractions 243
- Rack Display 244
- Outlet Valve Display 244
- Fractions Table 245
- User Guide 247
- Viewing Fraction Details 248
- Pooling Fractions 248
- To create a fraction pool 249
- To clear a pool 249
- Viewing Heat Map Details 251
- To hide the rack display 252
- Column Performance Analysis 253
- Calculate 254
- Analyzing Column Performance 255
- Click Delete 256
- Column Performance Table 258
- User Guide 259
- 8 Importing and Exporting 261
- To import a method or run 262
- Importing Unicorn Data Files 263
- User Guide 265
- Exporting Data 266
- Exporting Data as an NGC File 267
- To export only the method 268
- To export only the run data 268
- User Guide 269
- To export multiple runs 270
- User Guide 271
- Exporting Diagnostic Logs 273
- Change the date range 274
- User Guide 275
- 9 Reports 277
- Producing a Report 278
- Method Reports 279
- Run Reports 279
- Printing a Report 280
- Saving a Report 281
- 9 282
- A Database Management 283
- Back Up the NGC Database 284
- Restore the NGC Database 285
- A 286
- B Multicolumn Purifications 287
- Method base unit 288
- Sample volume 288
- Step duration and length 288
- User Guide 289
- Template Name Description 291
- Plumbing the NGC System 293
- Purification Templates 294
- User Guide 295
- Priming the NGC System 296
- User Guide 297
- Life Science 300
- Bio-Rad 300
- Laboratories, Inc 300
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