399

1-D gel evaluation 87 f2-D electrophoresis 42, 92 ff, 100 ff, 275 ff,

379 ffs. a. Two-dimensional electrophoresis

2-D gel evaluation 88 f

aA, C, G, T s. adenine, cytosine, guanine,

thymineAcid violet 17 staining 271 fAcid violet staining 272, 290Acrylamide 16, 339, 352, 359

formula 16gel recipes, DNA gels 243, 300 f

IEF gel 200, 259Immobiline gel 259, 279 fnative PAGE 177SDS gradient gel 218, 236, 284titration curve gel 164ultrathin layer electrophoresis 148vertical gels 233

gradient 36immobilized pH gradient 59polymerization 148, 164, 180, 200, 221,

235, 266, 285Additive gradient 30, 62, 321 ffAdenine 25Adsorption

on blotting membranes 68 ff, 109, 247 ffon capillaries 11

A/D transformer s. analog digital transformerAffinity electrophoresis 20, 151AFLP 331Agarose gel 15, 151, 189, 348 ff

blotting 68clinical routine 17electrophoresis 15immunoelectrophoresis 18native immunoblotting 69

PFG electrophoresis 23preparation 152 ff, 190 ffprotein detection 159 ff, 195 frehydratable 54submarine electrophoresis 12, 22, 125

Agarose IEF 54method 189 ff, 348 ff

Albumin 94Alkaline blotting 74Alkaline phosphatase 20, 76Alkylation 37, 101, 116, 214, 363Allelic ladder 32Amido Black staining 75, 253Ammonium nitrate 162, 171, 187, 196, 206Ammonium persulfate 16, 164, 339,

343, 352gel recipe, DNA gels 243, 303

IEF gel 200Immobiline gel 259, 281native electrophoresis 177SDS gradient gel 218, 236, 284titration curve gel 164, 166ultrathin layer gel electrophoresis 148

Amphoteric buffer 4, 42, 175, 187Amphoteric substance 5, 51, 65Amplification products 27 ff, 299 ff, 385Amplifying enzyme 77Analog digital transformer 111Analysis, quantitative 5, 48 ff, 81 ffAnion 5, 47Antibody 18, 67, 151 ff

detection an blots 76 fimmunoblotting 76, 109, 253quantity 82reactivity 75solution 155, 158, 196, 209

Antigen 18, 67, 159Antigen-antibody titer 82, 159API 114

Index

Electrophoresis in Practice, Fourth Edition. Reiner WestermeierCopyright � 2005 WILEY-VCH Verlag GmbH & Co. KGaA, WeinheimISBN: 3-527-31181-5

400

APS s. ammonium persulfateARDRA 28Automation 12, 25, 78, 128 fAutoradiography 13, 24, 32, 76, 85,

103, 377

bBAC s. bisacryloylcystamineb-Alanine 58Base pair ladder 299Base pairs 27, 32Basic pH gradients 61, 101Bi-directional electrophoresis 24Bi-directional transfer 68, 71Bioinformatics 92, 118Biotin-avidin 24, 77Bis s. NN¢-methylenebisacrylamideBisacryloylcystamine 17Blocking 75, 253, 377 fBlotting 4, 13, 67 ff, 104

blotting equipment 69 ff, 127, 129, 247 ffelectrophoretic 24, 70, 247 ff, 371 ffof IEF 69of SDS electropherograms 250semi dry 70, 247 fftrouble-shooting 371 ff

Blotting membranes 67 ff, 72, 249blocking of – 75staining of – 75, 253

Blue Native PAGE 41, 99Blue toning 226 fBovine serum albumin 75, 90bp s. base pairsBSA s. bovine serum albuminBuffer 7, 34 ff, 73 f, 300 f, 317

agarose strips 128amphoteric 6, 42, 175 ffstrips, paper 14, 127, 149, 183, 222, 292,

307 f, 328, 334strips, polyacrylamide 40system, continuous 248system, discontinuous 5, 22, 34, 45, 74,

218, 228, 235, 238, 248 f, 300 f, 362

cCalibration curve 39, 85, 89Capillary blotting 68Capillary electrophoresis 3, 7, 10 ff, 27, 48,

53, 82Capillary isotachophoresis 48CAPS s. 3-(cyclohexylamino)-propanesulfonic

acidCarboxymethyl 73

Carrier ampholytes 55 ff, 62, 96, 100, 163,189, 197, 268, 277, 339 ffrecipe 166, 191, 201, 283

Catalyst 16, 42, 167, 175, 189Cathodic drift 58, 100, 208, 349

s. a. plateau phenomenonCation 5, 41, 47Cationic detergent 41, 42cDNA 32Cell disruption 276Cellulose acetate 13, 84Cerebrospinal fluid 16, 206Cetyltrimethylammonium bromide 42, 366CHAPS s.

3-(3-cholamidopropyl)dimethylammonio-1-propane sulfate

Charge heterogeneity 21, 81Chemically assisted fragmentation 113Chemiluminescence 24, 77, 843-(3-Cholamidopropyl)dimethylammonio1-

propane sulfate 277, 283Chromatography

of peptides 92of proteins 99

Chromosome separation 23Clinical diagnostic 13, 17, 151Clinical laboratory 17CM s. carboxymethylCo-analytical modifications 96Collagen Molecular Weight 214Collision-induced dissociation 115Colloidal gold 75, 77Colloidal staining, Coomassie 170, 185,

204 f, 224 f, 294, 358Complex formation 7, 82Composite proteins 184Computer

for DNA sequencing 26for evaluation 83 ff, 92, 118

Concentration effect 5, 47s. a. zone sharpening effect

Configurational changes 6, 52, 53, 96Confocal scan head 86Constant denaturing gel electrophoresis 30,

321 ffContinuous buffer 39, 74, 146, 151, 244Continuous buffer system 249Continuous freeflow electrophoresis 10Cooling 14, 29, 54, 122 ff, 318

thermoelectric 128 fCoomassie Blue staining 13, 41, 103

agarose gels 159, 195

Index

401

colloidal 170, 185, 204 f, 224 f, 294, 358,367

fast 170, 185, 205, 224immunoelectrophoresis 161

Counter immunoelectrophoresis 19Counter-ion 45Cross-linking 16, 27, 297, 315Cryo-isoelectric focusing 54, 209Cryoproteins 54CTAB s. cetyltrimethylammonium bromideCurrent 122Cy3 and Cy5 label 26, 104, 3113-(Cyclohexylamino)-propanesulfonic acid

73Cystein gels 101Cysteins 37, 101Cytosine 25

dDa s. DaltonDalton 20, 38Database 89, 108, 118Deep Purple� 104, 295Denaturing conditions

DNA 24, 30, 313 ff, 322 ff, 331 ffproteins 37, 57, 96, 208, 211 ff, 268, 277

Densitometric evaluation 83 ffDensitometry 83 ffDensity gradient 36, 60, 219, 236, 240, 265 f,

269, 281, 324 ffDesoxyribonucleic acid 116

blotting 68 ffSeparation methods 12, 15, 22 ff, 299 ff

Detection 12, 22, 29, 49, 76 ff, 84 ff, 102s. a. autoradiographys. a. blottings. a. staining

Detector 48, 112Detergent 12

anionic 211cationic 31, 366non-ionic 41, 53, 96zwitterionic 53, 277 ff

Dextran gel 62DGGE s. Denaturing gradient gel

electrophoresisDialysis 278Diazobenzyloxymethyl 72Diazophenylthioether 72Diethylaminoethyl 73Differential display reverse transcription 32,

299, 331Diffusion 5, 13, 17, 52, 56, 346, 375

immunodiffusion 19, 158Diffusion blotting 68DIGE 93, 98, 104 ff, 105 ffDigestion, tryptic 110Dimethylsulfoxide 209, 345Discontinuous buffer System 5, 22, 34 ff, 45,

175, 218, 228, 235, 238, 362blotting 74, 248 fDNA separations 300 ffproteins 34 ff, 218, 235, 284

Dissociation constant 3, 54, 59, 230, 260 f,281

Disulfide bond 17, 37, 96, 212, 277Disulfides 101, 380Dithioerythreitol 96, 268Dithiothreitol 37, 96, 214, 268, 273, 277,

363 ff, 380 ffDMSO s. dimethylsulfoxideDNA s. desoxyribonucleic acidDNA sequence 24 ff, 91 ff, 116DNA sequencing 24, 91, 116, 125 f

automated 12, 25Double Replica Blotting 71DPT s. diazophenylthioetherDrying of gels 298DSCP 30DTE s. dithioerythreitol

eEDTA s. ethylendinitrilotetracetic acidElectric field 3, 23, 55, 121

strength 25, 45, 158, 236Electrode buffer 7, 14, 22, 34 f, 70 ff, 131, 146

precipes for DNA 244, 300 f, 322, 333recipes for DNA 317recipes for proteins 152, 183, 215, 228,

233, 245, 248 f, 280Electrode solutions 57, 193, 208, 286, 347Electroendosmosis 7, 13, 15, 54, 175, 189,

280Electroendosmotic effect 290, 341, 348, 380

in capillaries 11Electro-osmotic movement 7, 19, 22Electrophoresis 4, 9, 122

2D electrophoresis 42, 92 ff, 275 ff, 379 ffautomated 12, 25, 128 fbidirectional 24continuous free flow zone 10discontinuous 34 fin pulsed electric field 23, 126

Electrophoretic blotting 70, 127, 247 ff, 371 ffElectrophoretic mobility 3, 15, 21, 23, 45Electrospray ionization 114, 119

Index

Elution, electrophoretic 63, 110Emission 26, 85, 104Enzymatic digestion 92, 110 ffEnzyme

enzyme blotting 76enzyme detection 13, 18, 20, 76enzyme detection System 76enzyme inhibitor 20enzyme substrate complex 54, 209enzyme substrate reaction 76separation 13, 18, 20, 209

Equilibration 280, 290, 381Equipment 123 fEquivalence point 19Escherichia coli 277 fESI 115Ethidium bromide 22, 27, 84, 132, 244, 299,

311Ethylendinitrilotetraacetic acid

buffer 24, 75, 225, 244, 277, 317, 333sample preparation 277, 316staining 225

Evaluation 81 ff, 107Excitation 26, 85, 104

fFar western blotting 70Ferguson plot 21Field strength 3, 25, 45, 158, 270Flatbed systems 14, 126 f, 145 ff, 292 fFluorescence

detection 85 flabeling 25, 104staining 104

Fluorography 13, 25Focusing, isoelectric s. isoelectric focusingFree flow electrophoresis 10, 99 f

gGC s. group specific componentsGC clamps 322Gel 3, 13

non-restrictive 17rehydrated 41

for DNA 300 ff, 314 ff, 323 f, 333for proteins 42, 53, 61, 62, 163, 175,

197 ffrestrictive 21, 32

Glassfiber membrane 63, 73, 78Glycoproteins 41, 72, 73, 78, 367Gold, colloidal 75, 77Gradient

additive 269, 321

pH 4 f, 45 ff, 163, 189 ff, 197, 265, 281porosity 219, 236 f, 240, 284 f

Gradient gel 36, 129preparation 219 f, 236 f, 240, 265, 269, 281,

284 f, 323Group specific component 268Guanine 25

hHenderson-Hasselbalch equation 60HEPES s. N-(2-hydroxyethyl)piperazine-N¢-2-

ethanesulfonic acidHeteroduplex 30, 299High Molecular Weight 79, 214High-resolution 2D electrophoresis 42, 92 ff,

275 fftrouble shooting 379 ff

Histones 41, 61HMW s. High Molecular WeightHybridization 76Hydrolysis 40, 102, 259

iIdentification 13, 76, 76 f, 109IEF s. isoelectricfocusingIEF marker 54, 65IgG s. immunoglobulin GImage analysis 87 ff, 107 ffImmobilized pH gradients 59 ff, 63, 100,

257 ff, 279 ff, 352 ffrecipes 261 f, 281

Immobilizing membranes 67 ffImmunoblotting 76, 109, 253, 377

native 79Immunodiffusion 19, 158Immunoelectrophoresis 19 ff, 42, 82, 151 ffImmunofixation 18, 151, 160, 189,

196, 209Immunoglobulin G 19, 76, 151Immunoglobulin M 54Immunoglobulins 94Immunoprecipitation 13, 18 ffImmunoprinting 13, 18Indian Ink staining 75, 254Internal standard 93, 107Ion-exchange chromatography 172Ionic strength 6, 34, 39, 60, 228Ion trop 115IPG s. immobilized pH gradientsIsoDalt system 276Isoelectric focusing 4, 51 ff, 100 f, 173, 189 ff,

197 ff, 257 ffblotting of – 69, 73, 372

Index402

current during – 122equipment 127, 130in agarose gel 189 ffin immobilized pH gradients 257 ffin polyacrylamide gel 197 ffprinciples 51 ffsample application 82, 194, 204, 269 f,

286 fftrouble-shooting 339 ff

Isoelectric membranes 63, 100Isoelectric point 5, 42, 65, 87 f, 93Isotachophoresis 5, 35, 45 ffIsotachophoresis effect 35, 71ITP s. isotachophoresis

jJoule heat 6, 10, 122, 288

lLabelling

of nucleic acids 25 ffof proteins 98, 105 ff

Laboratory equipment 133 ffLactate dehydrogenase 18, 21Laser 26, 78, 85 f, 107, 111Laser capture universdissection 94LDAO, Lauryldimethylamine-N-oxide 209Leading ion 4, 45, 248Lectin 20, 67Lectin blotting 78Linear polyacrylamide 12, 27, 54Lipopolysaccharide 13, 73Low molecular Weight 56, 116, 196

peptides 40, 81, 210, 230, 242 ffsubstances 20, 145, 182

Lysis buffer 96, 277

mMALDI 78, 110 ff, 111Marker

DNA 299protein 39, 54, 82, 90

Mass spectrometry 78, 91 ff, 92, 102, 297 fMedium, stabilizing 6, 7, 12 ffMEKC 12

s. a. Micellar electrokinetic chromatographyMembrane

Membranecellulose acetate 13immobilizing 67 ffisoelectric 63 f

Membrane proteins 41, 268, 277

MES, 2-(N-Morpholino)ethanesulfonic acid42

Micellar electrokinetic chromatography 12Microsatellites 31Migration, electrophoretic 7, 19, 121Minimum labelling 98, 105Mixed disulfides 101Mobility 30, 45, 52

relative electrophoretic 4, 12, 45Molecular diameter, radius 21, 36Molecular weight 12, 37, 41, 42, 87, 214MOPS, 3-(N-morpholino)propanesulfonic

acid 42Mouse liver 95Moving boundary electrophoresis 5, 9MuDPIT 92Mutation detection 28 ff

nN-(2-Hydroxyethyl)piperazine-N¢-

ethanesulfonic acid 42, 175NAP s. Nucleic Acid PurifierNative electrophoresis DNA 27, 76

electrophoresis proteins 22, 175immunoblotting 79

Negative spots 295Negative staining 103, 225, 295NEPHGE 100Net charge 5, 17, 35, 39

isoelectric focusing 51speed of migration 21, 121

Net charge curve 51, 64, 172Neutral blotting 74Nitrocellulose 72, 253NN¢-Methylenebisacrylamide

formula 17gel recipe, DNA gels 303

IEF gel 200, 259Immobiline gel 258, 279native PAGE 177SDS gradient gel 215, 233titration curve gel 164ultrathin layer gel 145

Nonidet s. non-ionic detergentNon-ionic detergent 42, 53, 79, 96, 197, 268,

340Non-restrictive medium 17Northern blotting 70Nucleic Acid Purifier 151, 163, 176, 190, 198,

258Nucleoprotein 41, 52Nylon membrane 72, 75

Index 403

oO.D. s. optical densityOligoclonal IgG 206Oligo nucleotides 310Oligo peptides 185, 210, 230, 236, 242 ff

peptides 204Optical density 84Oxidation 96, 116, 209, 343, 363

pPaper electrophoresis 12Paper replica 62PBS s. Phosphate Buffered SalinePCR� s. Polymerase Chain ReactionPefabloc 97, 277Peptide gels 40, 229 ff, 242 ffPeptide massfingerprinting 116Peptides 40, 63, 92, 102, 229 ff, 242 ffPeptides detection 170, 204, 210, 245, 271Peroxidase, horseradish 76, 77PFG s. Pulsed-Field Gel ElectrophoresisPhenylmethyl-sulfonylfluoride 97, 258pH gradient 5, 52 ff, 55, 65, 122, 339 ff

2D electrophoresis 42, 96, 379 ffIEF method 202 ff, 259 ff, 281

Phosphatase, alkaline 20, 76, 77Phosphate Buffered Saline 254, 276Phosphoglucomutase 268Phosphor imager 85Photographing 83PI s. Protease InhibitorpK value s. dissociation constantPlasma, human 94Plasma proteins 94Plateau phenomenon 58, 100, 341PMSF s. phenylmethyl-sulfonylfluoridePolyacrylamide gel 16 ff, 20, 24 ff, 227

DNA 24 ff, 244, 300 ff, 385 ffproteins 175

electrophoresis 145isoelectric focusing 53, 197 ff, 257 ff, 279 fflinear 12, 27rehydratable 41, 163, 175, 197 ff, 257 ff,

281 f, 299 ffPolymerase Chain Reaction 3, 27 ff, 244,

299 ff, 385 ffPolymerization 17Polyphenols 145Polysaccharide 15Polysaccharide separation 13Polyvinyledendifluoride 72, 376Pore gradient gels 36, 129, 219, 236 f, 240,

284 f

preparation 219 f, 236 f, 240, 284 fPost-source decay 113Post-translational modifications 91 ff, 113,

116, 118Power, electric 122Precipitation 18, 34, 97, 277, 278

line 18, 82Prefractionation 98Pressure blotting 68Protease Inhibitor 61, 97, 258, 277Protein A, radioiodinated 76Protein sequencing 78, 109Proteome analysis 5 f, 43, 92 ff, 275Pulsed-Field Gel Electrophoresis 23Purity control 81Purity test 5PVDF s. polyvinylidenfluoride

qQuantification 5, 81 ff, 89 ffQuantitative analysis 5, 48

rr s. molecular radiusRadioactive labeling 13, 24, 32, 76, 85, 103,

377Radioiodinated protein A 76RAPD, Random amplified polymorphic

DNA 28, 299Reflectron 112Rehydratable agarose gel 54Rehydrated polyacrylamide gels 300 ff

proteins 41, 175, 197 ff, 230, 257 ff, 281 fRelative mobility 12, 21Resolution 13, 52, 60, 92, 112, 228, 242, 257Resolving gel 34 ff, 180, 218, 235 ff,

243, 284, 303Restriction Fragment Length Polymorphism

22Restrictive medium 21, 34Retardation coefficient 21Reversible staining 13, 75, 103, 225, 253, 295RFLP s. Restriction Fragment Length

PolymorphismRibonucleic acid 20, 32 ff, 68, 76, 299RNA s. ribonucleic acidRNase protection assay 332Rocket technique 19, 155Routine, clinical 13, 84

sSample 6Sample preparation

Index404

DNA 305, 315 f, 322, 332proteins 82, 96 ff

denaturing 212 ff, 276 ffimmunoelectrophoresis 151Isoelectric focusing 189 f, 198, 258native electrophoresis 151, 176SDS treatment 212 fftitration curve analysis 163

vertical gels 232Saturation labelling 98, 105Scanner 85 f, 104Scanning 83 ffSDS s. sodium dodecyl sulfateSDS electrophoresis 37 ff, 101SDS treatment 212 ff, 290Secondary antibody 77, 377Semi dry blotting 70 f, 74, 127, 129, 247 ff,

371 ffSeparator IEF 58Sephadex 62, 100Sephadex IEF 62, 100Sequencing

DNA 24 ff, 116proteins 109, 113, 118

Silver amplifying 77Silver staining 16, 29, 27 ff, 103, 128, 129,

367recipe, protein PAGE 226recipes, agarose gels 162, 196

PAGIEF 206 ffprotein PAGE 186 f, 295

Single nucleotide polymorphism 31Single strand conformation polymorphism

29, 313 ff, 385Slotformer 146, 152 f, 178, 216, 301SNP Analysis 31Sodium dodecyl sulfate 35 ff, 73, 78, 101,

211, 231, 248, 250, 279, 284Software, image analysis 87 fSpot

identification 109picker 104, 107 f

SSCP 29s. a. Single strand conformation

polymorphismStabilizing medium 12Stable isotope labeling 103Stacking gel 34 ff, 101, 180, 218, 228, 235 ff,

284, 303Staining 13

2-D gels 103 f, 294 ffagarose gels 159, 195blotting membranes 75 ff, 253 f

DNA gels 22, 310IEF gels 195, 204 ff, 271 f, 290native gels 185 fSDS gels 224 ff, 294 fftitration curve gels 170 f

Starch gel 15Storage phosphor plates 85, 103Storage phosphor screen 85“Submarine” chamber, gel 22, 125SYPRO Ruby� 102, 104

tT s. total acrylamide concentration [%]Tank blotting 70, 73TBE s. Tris borate EDTATBP 101TCEP 101TEMED, N,N,N¢,N¢-

tetramethylethylendiamine 16, 42, 175,189, 303gel recipes 148

DNA gels 235 ff, 303Immobiline gel 264, 281native electrophoresis 180SDS gradient gel 218, 235 ff, 284titration curve gel 167

Temperature gradient gel electrophoresis 31,331

Terminating ion 4 f, 47, 248TGGE s. Temperature gradient gel

electrophoresisThermoblotting 68Thiourea 97, 279Thymine 25, 299Titration curve analysis 64, 163 ff, 175, 346TMPTMA 254Transferrin 268Transmission 107Trichloroacetic acid, fixing 159, 170, 185,

195, 205, 272Tris, tris(hydroxymethyl)-aminomethane 74

acetate 41, 228, 245, 300, 317ascorbic acid 273borate-EDTA 24, 41, 244, 301, 317, 322,

333, 367glycine 34 ff, 74, 101, 215, 233,

248, 280HCl 34 ff, 74, 101, 164, 177, 200, 215,

233, 280phosphate 301, 317, 322, 333tricine 41, 152, 228, 245, 300, 317tricine lactate 152

Triton X-100 s. detergent, non-ionic

Index 405

Trouble-shooting guide 339 ffTwo-dimensional electrophoresis 42, 60, 62,

92, 100 ff, 275 ff, 379 ffDNA 33

uUltraviolet light 11, 48, 75, 81, 254Urea 30 ff, 53, 57, 74, 79, 189, 208, 268

2D electrophoresis 42, 96 ff, 268, 277DNA 24, 322 f, 331 ff

UV s. ultraviolet light

vVacuum 111, 115

blotting 69

Vertical systems 14, 124 f, 231 ff, 291Viroids 33Viscoelastic relaxation time 23VLDL, very low density lipoproteins 269Voltage 122

wWestern blotting 70

zZone electrophoresis 17 ff, 19, 36,

52, 170Zone sharpening effect 5, 35, 36, 45

Index406