Gel electrophoresis of proteins: a practical approach
Gespeichert in:
| Format: | Buch |
|---|---|
| Sprache: | English |
| Veröffentlicht: |
Oxford [u.a.]
Oxford Univ. Press
1998
|
| Ausgabe: | 3. ed. |
| Schriftenreihe: | The practical approach series
197 |
| Schlagworte: | |
| Online-Zugang: | Inhaltsverzeichnis |
| Beschreibung: | XX, 352 S. Ill., graph. Darst. |
| ISBN: | 0199636419 0199636400 |
Internformat
MARC
| LEADER | 00000nam a2200000 cb4500 | ||
|---|---|---|---|
| 001 | BV012149349 | ||
| 003 | DE-604 | ||
| 005 | 19990224 | ||
| 007 | t | ||
| 008 | 980910s1998 ad|| |||| 00||| eng d | ||
| 020 | |a 0199636419 |9 0-19-963641-9 | ||
| 020 | |a 0199636400 |9 0-19-963640-0 | ||
| 035 | |a (OCoLC)245700087 | ||
| 035 | |a (DE-599)BVBBV012149349 | ||
| 040 | |a DE-604 |b ger |e rakddb | ||
| 041 | 0 | |a eng | |
| 049 | |a DE-20 |a DE-M49 |a DE-703 |a DE-19 | ||
| 050 | 0 | |a QP551 | |
| 082 | 0 | |a 572/.6 | |
| 084 | |a VG 8400 |0 (DE-625)147221:253 |2 rvk | ||
| 084 | |a WC 3440 |0 (DE-625)148085: |2 rvk | ||
| 084 | |a CHE 808f |2 stub | ||
| 084 | |a CHE 820f |2 stub | ||
| 084 | |a CHE 810f |2 stub | ||
| 084 | |a CHE 228f |2 stub | ||
| 245 | 1 | 0 | |a Gel electrophoresis of proteins |b a practical approach |c ed. by B. D. Hames |
| 250 | |a 3. ed. | ||
| 264 | 1 | |a Oxford [u.a.] |b Oxford Univ. Press |c 1998 | |
| 300 | |a XX, 352 S. |b Ill., graph. Darst. | ||
| 336 | |b txt |2 rdacontent | ||
| 337 | |b n |2 rdamedia | ||
| 338 | |b nc |2 rdacarrier | ||
| 490 | 1 | |a The practical approach series |v 197 | |
| 650 | 0 | 7 | |a Gelelektrophorese |0 (DE-588)4123267-7 |2 gnd |9 rswk-swf |
| 650 | 0 | 7 | |a Proteine |0 (DE-588)4076388-2 |2 gnd |9 rswk-swf |
| 689 | 0 | 0 | |a Proteine |0 (DE-588)4076388-2 |D s |
| 689 | 0 | 1 | |a Gelelektrophorese |0 (DE-588)4123267-7 |D s |
| 689 | 0 | |5 DE-604 | |
| 700 | 1 | |a Hames, B. D. |e Sonstige |4 oth | |
| 830 | 0 | |a The practical approach series |v 197 |w (DE-604)BV007921321 |9 197 | |
| 856 | 4 | 2 | |m HEBIS Datenaustausch |q application/pdf |u http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=008228598&sequence=000002&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |3 Inhaltsverzeichnis |
| 999 | |a oai:aleph.bib-bvb.de:BVB01-008228598 | ||
Datensatz im Suchindex
| _version_ | 1804126758366085120 |
|---|---|
| adam_text | Gel Electrophoresis
of Proteins
A Practical Approach
THIRD EDITION
Edited by
B D HAMES
School of Biochemistry and Molecular Biology
University of Leeds, Leeds LS2 9JT, UK
Oxford New York Tokyo
OXFORD UNIVERSITY PRESS
Contents
List of contributors xvii
Abbreviations xix
1 One-dimensional polyacrylamide gel
electrophoresis 1
Qinwei Shi and George Jackowski
1 Introduction 1
2 The polyacrylamide gel matrix 2
Chemical structure and mechanism of polymerization 2
Factors affecting polymerization 3
Resolving range of polyacrylamide gels 7
3 Gel apparatus 8
Electrophoresis apparatus 8
Power pack 10
Apparatus for gradient gels 10
Photopolymerization equipment 10
4 Choice of a gel system 11
Non-dissociating systems 11
Dissociating systems 11
Transverse gradient gels 13
5 SDS-polyacrylamide gel electrophoresis 13
Introduction 13
Reagent preparation 14
Gel preparation 16
Sample preparation 21
Sample loading and electrophoresis 27
Molecular mass estimation 29
Troubleshooting 33
6 Non-denaturing polyacrylamide gel electrophoresis 35
Introduction 35
Choice of buffers and polymerization catalyst 37
Preparation of non-denaturing slab gels 38
Sample preparation and electrophoresis of native proteins 39
7 Variations of standard polyacrylamide gel electrophoresis 40
Blue native polyacrylamide gel electrophoresis 40
Acid-urea polyacrylamide gel electrophoresis 42
Transverse gradient gel electrophoresis 42
Contents
Tricine SDS-polyacrylamide gel electrophoresis 46
Use of cationic detergents for PAGE 47
Acknowledgements 49
References 50
2 Protein detection methods 53
Carl R Merril and Karen M Washart
1 Detection of proteins in gels 53
Introduction 53
Organic dyes 55
Silver stains 57
Negative staining 61
Fluorescent stains 62
Direct detection of proteins 64
Detection of radioactive proteins 65
Detection of specific proteins 69
Immunological detection methods 70
2 Western blots 71
Choice of blotting matrix 71
General detection of proteins on Western blots 72
Detection of radioactively labelled proteins 78
Detection of glycoproteins _ 78
Immunological detection of proteins 78
3 Enzyme staining 83
Categories of enzymes detected 83
Specific localization methods 84
Protein renaturation assays 84
4 Quantitative analysis of electrophoretically separated proteins 85
References 86
3 Preparative gel electrophoresis 93
Kelvin H Lee and Michael G Harrington
1 Introduction 93
2 Preparative isoelectric focusing 95
Technology options—immobilized pH gradient versus carrier
ampholyte isoelectric focusing 95
Preparative liquid isoelectric focusing 96
3 Preparative SDS-PAGE 97
Technology options—the Bio-Rad Prep Cell versus scale-up 97
Preparative SDS-PAGE using the Bio-Rad Prep Cell 97
Contents
4 Micropreparative two-dimensional electrophoresis 98
5 Protein recovery from gels 100
Elution of proteins from blotting membranes 101
Elution of proteins from gels 102
6 Conclusions 103
References 104
4 Polymer solution-mediated size separation
of proteins by capillary SDS electrophoresis 105
Andras Gunman, Paul Shieh, and Barry L Karger
1 Introduct ion 105
2 Fundamenta ls of capillary gel electrophoresis 106
3 Operat ional variables 108
Applied electric field and temperature 109
Capillary dimensions 109
Capillary coating 110
Gels and polymer solutions 112
4 Separations 113
Cross-linked polyacrylamide gel 113
Linear polyacrylamide solutions 115
Other entangled polymer solutions 116
The Ferguson method 121
Detection 122
Ultrafast separations 123
5 Conclusions 123
Acknowledgements 124
References 124
5 Conventional isoelectric focusing in gel
slabs, in capillaries, and immobilized
pH gradients 127
Pier Giorgio Righetti, Alessandra Bossi, and Cecilia Gelfi
1 Introduction 127
2 Conventional IEF in amphoter ic buffers 128
General considerations 128
Equipment 132
The polyacrylmide gel matrix 135
Gel preparation and electrophoresis 140
xi
Contents
General protein staining 150
Specific protein detection methods 153
Quantitation of the focused bands 154
Troubleshooting 154
3 Immobil ized p H gradients 156
General considerations 156
IPG methodology 166
Troubleshooting 175
Analytical results with IPGs 177
4 Capillary isoelectric focusing (cIEF) 177
General considerations 177
cIEF methodology 179
5 Conclusions 183
Acknowledgements 183
References 183
6 Two-dimensional gel electrophoresis 189
Samir M Hanash
1 Introduction 189
2 World Wide Web (WWW] resources 189
3 2DE reagents 190
4 Equipment 191
First-dimension isoelectric focusing apparatus 191
Second-dimension slab gel apparatus 192
Apparatus for electroblotting 193
Power supply 1 93
Platform shaking table 193
Apparatus for gel drying 194
5 Sample preparat ion and solubil ization 194
Sample preparation 194
6 Firs t-dimension electrophoresis 195
First-dimension IEF with carrier ampholytes 195
Immobilized pH gradient gels 198
7 Second-dimension SDS-polyacrylamide gel electrophoresis 200
Casting gradient SDS gels 200
Application of the first-dimension gel to the SDS slab gel and
electrophoresis 201
8 Preparative 2DE, special gradients, and gel formats 203
Preparative IPG 203
Special gel formats 205
xii
Contents
9 Electrophoretic transfer of proteins (Western blotting) 207
Electroblotting 207
Protein staining of electroblots 207
10 Gel analysis 208
Causes of variability in 2D patterns 208
Computerized 2D gel scanning and analysis 210
References 211
7 Peptide mapping 213
Anthony T Andrews
1 Introduction 213
2 Apparatus 217
3 The standard technique 217
Using pure protein samples 217
Standard in situ hydrolysis procedure 220
4 Variations of the standard technique 223
5 Peptide mapping of protein mixtures 225
6 Characterization of proteinases 229
7 Interpretation of results 231
References 233
8 Sequence analysis of gel-resolved proteins 237
Richard J Simpson and Gavin E Reid
1 Introduction 237
2 Visualization and concentration of protein gel spots 239
Protein detection in primary gels 239
Protein concentration 242
3 N-terminal sequence analysis 244
4 Internal sequence analysis 248
General considerations 248
Choice of proteolytic enzyme 249
Reduction/alkylation and proteolytic digestion of protein spots 249
Reverse-phase HPLC of peptides 255
5 Mass spectrometric methods for identifying gel-separated
proteins 257
General concepts 257
Sample preparation 259
xiii
Contents
Peptide-mass fingerprinting search algorithms 261
Peptide sequence tag search algorithms 261
Peptide fragment ion search algorithms 262
Acknowledgements 263
References 263
9 Fluorophore-labelled saccharide
electrophoresis for the analysis of
glycoproteins 269
Peter Jackson
1 Introduction 269
2 The release of oligosaccharide from glycoconjugates 270
Enzyme release of glycans 271
Release of saccharides using hydrazinolysis 273
3 Labelling of saccharides with fluorophores 275
4 Electrophoretic standards 277
5 Polyacrylamide gel electrophoresis of ANTS- and AMAC-
labelled saccharides 279
Electrophoresis of ANTS-labelled neutral and acidic saccharides
and AMAC-labelled acidic saccharides 279
Electrophoresis of AMAC-labelled neutral saccharides 282
Oligosaccharide profiles 283
Preparative elution of fluorophore-labelled saccharides from gels 285
6 Enzymatic structural analysis of N-glycans 286
7 Viewing and imaging 289
References 291
10 Analysis of protein:protein interactions by
gel electrophoresis 293
Vincent M Coghlan
1 Introduction 293
2 Isolation of protein complexes 294
Co-immunoprecipitation 294
Affinity purification 297
Chemical cross-linking 298
3 Identifying binding partners 300
Protein overlay 301
Two-dimensional diagonal analysis 305
xiv
Contents
4 Characterizing protein interactions 306
Mobility-shift assay 307
Quantitative methods 309
5 Limitations 310
References 311
Appendices
A l List of suppliers 313
A2 Bibliography of polypeptide detection methods 319
Index 345
XV
|
| any_adam_object | 1 |
| building | Verbundindex |
| bvnumber | BV012149349 |
| callnumber-first | Q - Science |
| callnumber-label | QP551 |
| callnumber-raw | QP551 |
| callnumber-search | QP551 |
| callnumber-sort | QP 3551 |
| callnumber-subject | QP - Physiology |
| classification_rvk | VG 8400 WC 3440 |
| classification_tum | CHE 808f CHE 820f CHE 810f CHE 228f |
| ctrlnum | (OCoLC)245700087 (DE-599)BVBBV012149349 |
| dewey-full | 572/.6 |
| dewey-hundreds | 500 - Natural sciences and mathematics |
| dewey-ones | 572 - Biochemistry |
| dewey-raw | 572/.6 |
| dewey-search | 572/.6 |
| dewey-sort | 3572 16 |
| dewey-tens | 570 - Biology |
| discipline | Chemie / Pharmazie Biologie Chemie |
| edition | 3. ed. |
| format | Book |
| fullrecord | <?xml version="1.0" encoding="UTF-8"?><collection xmlns="http://www.loc.gov/MARC21/slim"><record><leader>01727nam a2200469 cb4500</leader><controlfield tag="001">BV012149349</controlfield><controlfield tag="003">DE-604</controlfield><controlfield tag="005">19990224 </controlfield><controlfield tag="007">t</controlfield><controlfield tag="008">980910s1998 ad|| |||| 00||| eng d</controlfield><datafield tag="020" ind1=" " ind2=" "><subfield code="a">0199636419</subfield><subfield code="9">0-19-963641-9</subfield></datafield><datafield tag="020" ind1=" " ind2=" "><subfield code="a">0199636400</subfield><subfield code="9">0-19-963640-0</subfield></datafield><datafield tag="035" ind1=" " ind2=" "><subfield code="a">(OCoLC)245700087</subfield></datafield><datafield tag="035" ind1=" " ind2=" "><subfield code="a">(DE-599)BVBBV012149349</subfield></datafield><datafield tag="040" ind1=" " ind2=" "><subfield code="a">DE-604</subfield><subfield code="b">ger</subfield><subfield code="e">rakddb</subfield></datafield><datafield tag="041" ind1="0" ind2=" "><subfield code="a">eng</subfield></datafield><datafield tag="049" ind1=" " ind2=" "><subfield code="a">DE-20</subfield><subfield code="a">DE-M49</subfield><subfield code="a">DE-703</subfield><subfield code="a">DE-19</subfield></datafield><datafield tag="050" ind1=" " ind2="0"><subfield code="a">QP551</subfield></datafield><datafield tag="082" ind1="0" ind2=" "><subfield code="a">572/.6</subfield></datafield><datafield tag="084" ind1=" " ind2=" "><subfield code="a">VG 8400</subfield><subfield code="0">(DE-625)147221:253</subfield><subfield code="2">rvk</subfield></datafield><datafield tag="084" ind1=" " ind2=" "><subfield code="a">WC 3440</subfield><subfield code="0">(DE-625)148085:</subfield><subfield code="2">rvk</subfield></datafield><datafield tag="084" ind1=" " ind2=" "><subfield code="a">CHE 808f</subfield><subfield code="2">stub</subfield></datafield><datafield tag="084" ind1=" " ind2=" "><subfield code="a">CHE 820f</subfield><subfield code="2">stub</subfield></datafield><datafield tag="084" ind1=" " ind2=" "><subfield code="a">CHE 810f</subfield><subfield code="2">stub</subfield></datafield><datafield tag="084" ind1=" " ind2=" "><subfield code="a">CHE 228f</subfield><subfield code="2">stub</subfield></datafield><datafield tag="245" ind1="1" ind2="0"><subfield code="a">Gel electrophoresis of proteins</subfield><subfield code="b">a practical approach</subfield><subfield code="c">ed. by B. D. Hames</subfield></datafield><datafield tag="250" ind1=" " ind2=" "><subfield code="a">3. ed.</subfield></datafield><datafield tag="264" ind1=" " ind2="1"><subfield code="a">Oxford [u.a.]</subfield><subfield code="b">Oxford Univ. Press</subfield><subfield code="c">1998</subfield></datafield><datafield tag="300" ind1=" " ind2=" "><subfield code="a">XX, 352 S.</subfield><subfield code="b">Ill., graph. Darst.</subfield></datafield><datafield tag="336" ind1=" " ind2=" "><subfield code="b">txt</subfield><subfield code="2">rdacontent</subfield></datafield><datafield tag="337" ind1=" " ind2=" "><subfield code="b">n</subfield><subfield code="2">rdamedia</subfield></datafield><datafield tag="338" ind1=" " ind2=" "><subfield code="b">nc</subfield><subfield code="2">rdacarrier</subfield></datafield><datafield tag="490" ind1="1" ind2=" "><subfield code="a">The practical approach series</subfield><subfield code="v">197</subfield></datafield><datafield tag="650" ind1="0" ind2="7"><subfield code="a">Gelelektrophorese</subfield><subfield code="0">(DE-588)4123267-7</subfield><subfield code="2">gnd</subfield><subfield code="9">rswk-swf</subfield></datafield><datafield tag="650" ind1="0" ind2="7"><subfield code="a">Proteine</subfield><subfield code="0">(DE-588)4076388-2</subfield><subfield code="2">gnd</subfield><subfield code="9">rswk-swf</subfield></datafield><datafield tag="689" ind1="0" ind2="0"><subfield code="a">Proteine</subfield><subfield code="0">(DE-588)4076388-2</subfield><subfield code="D">s</subfield></datafield><datafield tag="689" ind1="0" ind2="1"><subfield code="a">Gelelektrophorese</subfield><subfield code="0">(DE-588)4123267-7</subfield><subfield code="D">s</subfield></datafield><datafield tag="689" ind1="0" ind2=" "><subfield code="5">DE-604</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Hames, B. D.</subfield><subfield code="e">Sonstige</subfield><subfield code="4">oth</subfield></datafield><datafield tag="830" ind1=" " ind2="0"><subfield code="a">The practical approach series</subfield><subfield code="v">197</subfield><subfield code="w">(DE-604)BV007921321</subfield><subfield code="9">197</subfield></datafield><datafield tag="856" ind1="4" ind2="2"><subfield code="m">HEBIS Datenaustausch</subfield><subfield code="q">application/pdf</subfield><subfield code="u">http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=008228598&sequence=000002&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA</subfield><subfield code="3">Inhaltsverzeichnis</subfield></datafield><datafield tag="999" ind1=" " ind2=" "><subfield code="a">oai:aleph.bib-bvb.de:BVB01-008228598</subfield></datafield></record></collection> |
| id | DE-604.BV012149349 |
| illustrated | Illustrated |
| indexdate | 2024-07-09T18:22:32Z |
| institution | BVB |
| isbn | 0199636419 0199636400 |
| language | English |
| oai_aleph_id | oai:aleph.bib-bvb.de:BVB01-008228598 |
| oclc_num | 245700087 |
| open_access_boolean | |
| owner | DE-20 DE-M49 DE-BY-TUM DE-703 DE-19 DE-BY-UBM |
| owner_facet | DE-20 DE-M49 DE-BY-TUM DE-703 DE-19 DE-BY-UBM |
| physical | XX, 352 S. Ill., graph. Darst. |
| publishDate | 1998 |
| publishDateSearch | 1998 |
| publishDateSort | 1998 |
| publisher | Oxford Univ. Press |
| record_format | marc |
| series | The practical approach series |
| series2 | The practical approach series |
| spelling | Gel electrophoresis of proteins a practical approach ed. by B. D. Hames 3. ed. Oxford [u.a.] Oxford Univ. Press 1998 XX, 352 S. Ill., graph. Darst. txt rdacontent n rdamedia nc rdacarrier The practical approach series 197 Gelelektrophorese (DE-588)4123267-7 gnd rswk-swf Proteine (DE-588)4076388-2 gnd rswk-swf Proteine (DE-588)4076388-2 s Gelelektrophorese (DE-588)4123267-7 s DE-604 Hames, B. D. Sonstige oth The practical approach series 197 (DE-604)BV007921321 197 HEBIS Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=008228598&sequence=000002&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Gel electrophoresis of proteins a practical approach The practical approach series Gelelektrophorese (DE-588)4123267-7 gnd Proteine (DE-588)4076388-2 gnd |
| subject_GND | (DE-588)4123267-7 (DE-588)4076388-2 |
| title | Gel electrophoresis of proteins a practical approach |
| title_auth | Gel electrophoresis of proteins a practical approach |
| title_exact_search | Gel electrophoresis of proteins a practical approach |
| title_full | Gel electrophoresis of proteins a practical approach ed. by B. D. Hames |
| title_fullStr | Gel electrophoresis of proteins a practical approach ed. by B. D. Hames |
| title_full_unstemmed | Gel electrophoresis of proteins a practical approach ed. by B. D. Hames |
| title_short | Gel electrophoresis of proteins |
| title_sort | gel electrophoresis of proteins a practical approach |
| title_sub | a practical approach |
| topic | Gelelektrophorese (DE-588)4123267-7 gnd Proteine (DE-588)4076388-2 gnd |
| topic_facet | Gelelektrophorese Proteine |
| url | http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=008228598&sequence=000002&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| volume_link | (DE-604)BV007921321 |
| work_keys_str_mv | AT hamesbd gelelectrophoresisofproteinsapracticalapproach |