Características

  • Género del libro Literatura y ficción
  • Tipo de narración Novela

Descripción

Libro: Ion Exchange Chromatography & Chromatofocusing

Contenido:

Principles of ion exchange ..................................................................................... 11
Net surface charge and pH ......................................................................................... 11
Steps in an IEX separation ......................................................................................... 12
Resolution ................................................................................................................ 14
Efficiency ...............................................................................................................................................15
Selectivity ...............................................................................................................................................17
Components of ion exchange media ............................................................................ 21
Matrix .....................................................................................................................................................21
Functional groups ....................................................................................................................................23
Binding capacity and recovery .................................................................................... 25
Chapter 2
Ion exchange in practice ........................................................................................ 26
Introduction .............................................................................................................. 26
Media selection ......................................................................................................... 26
Capture ..................................................................................................................................................26
Intermediate purification ..........................................................................................................................27
Polishing ................................................................................................................................................27
Fast media selection and method development ............................................................. 30
Automated media selection, method development and optimization ..............................................................30
Manual media selection, method development and optimization ...................................................................31
Practical considerations for IEX separation ................................................................... 34
pH and ionic strength ...............................................................................................................................34
Anion or cation exchanger ........................................................................................................................35
Strong or weak ion exchangers ..................................................................................................................36
Buffer selection and preparation ...............................................................................................................37
Column and media preparation .................................................................................................................39
Sample preparation ..................................................................................................................................39
Sample application ..................................................................................................................................40
Sample load ............................................................................................................................................40
Elution ...................................................................................................................................................42
Linear gradient elution .............................................................................................................................42
Step elution ............................................................................................................................................44
pH elution ...............................................................................................................................................46
Flow rates ...............................................................................................................................................46
Flow control ............................................................................................................................................47
Wash and re-equilibration .........................................................................................................................48
Detergents, denaturing agents and other additives ......................................................................................48
Analysis of results and further steps ............................................................................ 50
Scaling-up ................................................................................................................ 51
Equipment selection .................................................................................................. 52
Care of ion exchange media ........................................................................................ 52
Troubleshooting ......................................................................................................... 53
BioProcess Media – made for bioprocessing ................................................................. 58
Custom Designed Media ............................................................................................. 58
3
Chapter 3
Ion exchange media ............................................................................................... 59
Introduction .............................................................................................................. 59
MiniBeads: purification or analysis of microgram-milligram quantities
with highest resolution ............................................................................................... 60
Purification options ..................................................................................................................................61
Purification examples ...............................................................................................................................62
Performing a separation ............................................................................................................................63
Cleaning .................................................................................................................................................65
Media characteristics ...............................................................................................................................65
Chemical stability .................................................................................................................................... 66
Storage ................................................................................................................................................... 66
MonoBeads: purification of milligram quantities with highest resolution .......................... 67
Purification options ..................................................................................................................................68
Purification examples ...............................................................................................................................69
Performing a separation ............................................................................................................................71
Cleaning .................................................................................................................................................72
Media characteristics ...............................................................................................................................73
Chemical stability .................................................................................................................................... 73
Storage ................................................................................................................................................... 73
SOURCE: purification at high throughput with high resolution and easy scale-up ............. 74
Purification options ..................................................................................................................................75
Purification examples ...............................................................................................................................76
Performing a separation ............................................................................................................................80
Cleaning .................................................................................................................................................82
Media characteristics ...............................................................................................................................82
Chemical stability .................................................................................................................................... 83
Storage ................................................................................................................................................... 83
Sepharose High Performance: purification with high resolution ...................................... 84
Purification options ..................................................................................................................................84
Purification examples ...............................................................................................................................86
Scaling-up ..............................................................................................................................................86
Performing a separation ............................................................................................................................89
Cleaning .................................................................................................................................................91
Media characteristics ...............................................................................................................................92
Chemical stability .................................................................................................................................... 92
Storage ................................................................................................................................................... 92
Sepharose Fast Flow: purification with good resolution and easy scale-up ........................ 93
Purification options ..................................................................................................................................94
Purification examples ...............................................................................................................................97
Performing a separation .......................................................................................................................... 100
Cleaning ............................................................................................................................................... 102
Media characteristics ............................................................................................................................. 103
Chemical stability .................................................................................................................................. 104
Storage ................................................................................................................................................. 104
Sepharose XL – for selected proteins that require very high binding capacity
to increase productivity, easy scale-up ....................................................................... 105
Purification options ................................................................................................................................ 105
Purification examples ............................................................................................................................. 107
Performing a separation .......................................................................................................................... 109
Cleaning ............................................................................................................................................... 111
Media characteristics ............................................................................................................................. 111
4
Chemical stability ..................................................................................................................................112
Storage .................................................................................................................................................112
Sepharose Big Beads: purification from crude, viscous samples at large scale ............... 113
Purification options ................................................................................................................................ 114
Performing a separation .......................................................................................................................... 115
Cleaning ............................................................................................................................................... 116
Media characteristics ............................................................................................................................. 116
Chemical stability ..................................................................................................................................117
Storage .................................................................................................................................................117
Chapter 4
Ion exchange in a Purification Strategy (CIPP) ........................................................ 119
Applying CIPP .......................................................................................................... 120
Selection and combination of purification techniques ................................................. 120
Ion exchange as a capture step ................................................................................. 123
Ion exchange for intermediate purification ................................................................. 125
Ion exchange as a polishing step ............................................................................... 127
Alternative techniques for polishing steps .................................................................. 128
Chapter 5
Chromatofocusing: principles and methods ........................................................... 129
Introduction ............................................................................................................ 129
Media selection .....................................................................................................................................132
Buffer selection .....................................................................................................................................133
Purification options ................................................................................................................................ 134
Buffer options ....................................................................................................................................... 134
Purification examples ............................................................................................................................. 134
Packing a column ..................................................................................................................................136
Selection of pH gradient and buffers .......................................................................................................137
Buffer preparation ................................................................................................................................. 140
Sample preparation ................................................................................................................................ 141
Performing a separation .......................................................................................................................... 141
Optimization ......................................................................................................................................... 143
Troubleshooting .....................................................................................................................................144
Removing Polybuffer .............................................................................................................................. 148
Cleaning ............................................................................................................................................... 148
Media characteristics ............................................................................................................................. 149
Chemical stability ..................................................................................................................................150
Storage .................................................................................................................................................150
Chromatofocusing and CIPP ....................................................................................... 150

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