Handbook of affinity chromatography

Table of Contents......Page 0
Handbook of Affinity Chromatography, Second Edition......Page 6
Preface......Page 8
Contents......Page 9
Editor......Page 12
Contributors......Page 13
Section I: Introduction and Basic Concepts......Page 16
1.1 INTRODUCTION......Page 17
1.2.1 Origins of Affinity Chromatography......Page 19
1.2.2 Early Immobilization Methods......Page 20
1.2.3 Modern Era of Affinity Chromatography......Page 21
REFERENCES......Page 25
CONTENTS......Page 28
2.2.1 Chemical Inertness......Page 29
2.2.2 Chemical Stability......Page 33
2.2.4 Pore Size......Page 34
2.2.5 Particle Size......Page 35
2.2.5.1 Zonal Elution Chromatography......Page 36
2.2.5.3 Partial Loading in Adsorption-Desorption Affinity Purification......Page 37
2.2.6 Standard Commercially Available Affinity Supports......Page 38
2.3.2 Membranes......Page 39
2.3.3 Flow-Through Beads......Page 41
2.3.4 Continuous Beds......Page 42
2.4 SUMMARY AND CONCLUSIONS......Page 43
SYMBOLS AND ABBREVIATIONS......Page 44
REFERENCES......Page 45
CONTENTS......Page 47
3.1 INTRODUCTION......Page 48
3.2.1.1 Multisite Attachment......Page 49
3.2.1.2 Improper Orientation......Page 50
3.2.2.1 Choice of Support......Page 51
3.2.2.4 Other Considerations......Page 53
3.2.3.1 Amount of Immobilized Ligand......Page 55
3.2.3.3 Ligand Affinity and Adsorption Rate......Page 56
3.3.1 Nonspecific Adsorption......Page 57
3.3.2.1 Avidin and Streptavidin......Page 58
3.3.2.2 Protein A and Protein G......Page 60
3.4 COVALENT IMMOBILIZATION METHODS......Page 61
3.4.1 Amine-Reactive Methods......Page 63
3.4.1.2 Reductive Amination......Page 64
3.4.1.3 N-Hydroxysuccinimide Method......Page 66
3.4.1.4 Carbonyldiimidazole Method......Page 67
3.4.2 Sulfhydryl-Reactive Methods......Page 69
3.4.2.2 Maleimide Method......Page 70
3.4.4 Carbonyl-Reactive Methods......Page 72
3.5 OTHER IMMOBILIZATION TECHNIQUES......Page 75
3.5.1.1 General Techniques for Protein Entrapment......Page 76
3.5.1.3 Cell Entrapment......Page 77
3.5.2 Molecular Imprinting......Page 78
SYMBOLS AND ABBREVIATIONS......Page 79
REFERENCES......Page 80
CONTENTS......Page 91
4.1 INTRODUCTION......Page 92
4.2.1 General Factors Affecting Retention......Page 93
4.2.2 Relationship of Elution Method to the Affinity Ligand......Page 94
4.2.3 Affinity Chromatography versus Other Chromatographic Methods......Page 95
4.3.1 Solvent Considerations......Page 96
4.3.2 Factors Related to the Column and Sample......Page 97
4.3.3 Description of Sample Retention......Page 98
4.4.2 Selection of Elution Buffers......Page 100
4.4.3 Description of Sample Elution......Page 102
4.5.2 Description of Column Regeneration......Page 105
SYMBOLS AND ABBREVIATIONS......Page 106
REFERENCES......Page 107
Section II: General Affinity Ligands and Methods......Page 110
CONTENTS......Page 111
5.2.1 General Properties......Page 112
5.2.2 Applications in Affinity Chromatography......Page 114
5.2.4 Application and Elution Conditions......Page 115
5.3.2 Applications in Affinity Chromatography......Page 116
5.4 LECTINS......Page 118
5.4.2 Applications in Affinity Chromatography......Page 119
5.5 CARBOHYDRATES......Page 120
5.5.1 GENERAL PROPERTIES......Page 121
5.5.2 Applications in Affinity Chromatography......Page 122
5.6.1 General Properties......Page 123
5.6.2 Applications in Affinity Chromatography......Page 125
5.7.1 General Properties......Page 126
5.7.2 Applications in Affinity Chromatography......Page 127
5.8 SUMMARY AND CONCLUSIONS......Page 128
REFERENCES......Page 129
CONTENTS......Page 137
6.1.1 Antibody Structure......Page 138
6.1.2 Antibody-Antigen Interactions......Page 140
6.2.1 Antibody Production......Page 141
6.2.1.1 Polyclonal Antibodies......Page 142
6.2.1.2 Monoclonal Antibodies......Page 144
6.2.1.3 Autoantibodies......Page 145
6.2.1.5 Related Ligands......Page 146
6.2.2 Supports for Immunoaffinity Chromatography......Page 147
6.2.3.1 Random Immobilization Methods......Page 149
6.2.3.3 Adsorption to Secondary Ligands......Page 150
6.2.4.1 pH Elution......Page 152
6.2.4.2 Elution with Chaotropic Agents......Page 153
6.3.2 Off-Line Immunoextraction......Page 154
6.3.3 On-Line Immunoextraction......Page 157
6.4.1 Direct Analyte Detection......Page 160
6.4.2 Indirect Analyte Detection......Page 163
6.4.3 Postcolumn Immunodetection......Page 166
6.5 NEW DEVELOPMENTS IN IMMUNOAFFINITY CHROMATOGRAPHY......Page 168
SYMBOLS AND ABBREVIATIONS......Page 170
REFERENCES......Page 171
CONTENTS......Page 183
7.1.2 General Applications of DNA Affinity Chromatography......Page 184
7.2.1 Stationary Phases and Supports......Page 185
7.3.1 Stationary Phases and Supports......Page 186
7.3.2 Mobile Phases and Elution Methods......Page 187
7.4 PURIFICATION OF DNA REPAIR PROTEINS......Page 190
7.4.1 Proteins in Base Excision Repair......Page 191
7.4.2 Proteins in Nucleotide Excision Repair......Page 194
7.4.3 Proteins in Mismatch Repair......Page 195
7.4.4 Proteins in Recombination Repair......Page 196
7.4.5 Alternative Methods for Separating DNA Repair Proteins......Page 197
7.5 PURIFICATION OF TRANSCRIPTION FACTORS......Page 198
7.6.1 Histones......Page 199
7.6.2 Helicases......Page 200
7.6.4 Primases......Page 201
7.6.5 DNA Polymerases......Page 202
7.7 MECHANISM-BASED AFFINITY CHROMATOGRAPHY......Page 204
7.8 PURIFICATION OF RNA POLYMERASES......Page 205
7.10 PURIFICATION OF RESTRICTION ENZYMES......Page 206
7.10.2 Catalytic Chromatography......Page 207
7.11.1 The Bi-Column Method......Page 208
7.11.2 Oligonucleotide Trapping Method......Page 209
7.11.4 Catalytic Chromatography......Page 210
7.11.5 Aptamers......Page 214
7.12 SUMMARY AND CONCLUSIONS......Page 215
SYMBOLS AND ABBREVIATIONS......Page 216
REFERENCES......Page 217
CONTENTS......Page 225
8.2.1 Primary Interactions......Page 226
8.2.2.2 Ionic Interactions......Page 227
8.3.1 Boronate Ligands......Page 228
8.3.2 Solid Supports......Page 230
8.4.1 Carbohydrates......Page 231
8.4.2 Nucleosides, Nucleotides, and Nucleic Acids......Page 232
8.4.3 Glycoproteins and Enzymes......Page 233
8.4.4 Miscellaneous Small Molecules......Page 234
SYMBOLS AND ABBREVIATIONS......Page 235
REFERENCES......Page 236
CONTENTS......Page 240
9.1 INTRODUCTION......Page 241
9.2 DYE-LIGAND AFFINITY CHROMATOGRAPHY......Page 242
9.2.2 Selective Interactions of Dye-Ligands with Proteins......Page 243
9.2.3 Development of a Dye-Ligand Purification Method......Page 244
9.2.4 Recent Progress in Dye-Ligand Affinity Chromatography......Page 245
9.3.1 Methods Based on Protein Structure......Page 247
9.3.1.1.1 Biomimetic Dye-Ligands for (Keto)Carboxyl-Group-Recognizing Enzymes......Page 248
9.3.1.1.2 Biomimetic Dye-Ligands for Glutathione-Recognizing Enzymes......Page 249
9.3.1.1.3 Biomimetic Dye-Ligands for Galactose-Recognizing Enzymes......Page 250
9.3.1.2 Chlorotriazine-Linked Synthetic Biomimetic Ligands......Page 251
9.3.2 Combinatorial Approaches......Page 253
9.3.2.1 Affinity Chromatographic Screening of Combinatorial Peptide Libraries......Page 254
9.3.2.3 Ribosome Display Technology......Page 256
9.3.2.4 SELEX......Page 258
SYMBOLS AND ABBREVIATIONS......Page 260
REFERENCES......Page 261
CONTENTS......Page 265
10.1.1 Basic Principles of IMAC......Page 266
10.1.2 Recent Developments in the Use of Immobilized-Metal Affinity......Page 267
10.2.1 Support Selection......Page 268
10.2.2 Choice of Chelating Agents and Adsorption Center......Page 269
10.2.3 Selection of Metal Ions......Page 270
10.2.4 Mobile-Phase Selection......Page 273
10.2.6.1 pH Elution Schemes......Page 274
10.3 APPLICATIONS......Page 275
10.3.1.1 Proteins with Surface Histidines......Page 276
10.3.1.3 Plant Proteins......Page 277
10.3.2 Studies of Protein Surface Topography and Posttranslational Modifications......Page 278
10.3.3 Microanalytical Methods......Page 279
10.3.3.1 Immobilized Metal-Ion Affinity Gel Electrophoresis......Page 280
10.3.3.2 Immobilized Metal-Ion Affinity Capillary Electrophoresis......Page 281
10.3.4 Applications of IMAC in Genomics and Postgenomic Studies......Page 283
SYMBOLS AND ABBREVIATIONS......Page 285
REFERENCES......Page 286
Section III: Preparative Applications......Page 292
CONTENTS......Page 293
11.1.1 Challenges in Protein Purification......Page 294
11.2 GOALS FOR PROTEIN PURIFICATION......Page 295
11.3 GENERAL CONSIDERATIONS IN PROTEIN PURIFICATION......Page 296
11.3.3 Immobilization Conditions......Page 297
11.3.4 Adsorption and Elution Conditions......Page 298
11.4.1 Adsorption Isotherms......Page 299
11.4.2 Rate Equations......Page 301
11.4.3.1 General Description of Breakthrough Curves......Page 302
11.4.3.2 Use of Breakthrough Curves in Characterizing Separations......Page 304
11.4.4.1 General Description of NTUs......Page 308
11.4.4.2 Use of NTUs in Characterizing Separations......Page 310
11.5.1 Time Required for an Adsorption Process......Page 311
11.5.3 Effect of Flow Rate on Column Performance......Page 312
11.6 SUMMARY AND CONCLUSIONS......Page 313
SYMBOLS AND ABBREVIATIONS......Page 314
REFERENCES......Page 315
CONTENTS......Page 318
12.1 INTRODUCTION......Page 319
12.2.1 Cibacron Blue F3GA......Page 320
12.2.4 Lectins......Page 321
12.2.5 Thiopropyl Supports......Page 323
12.2.7 Hydrophobic Interaction Chromatography......Page 324
12.2.8 Immobilized Metal-Ion Affinity Chromatography......Page 325
12.3.1.1 Diazonium......Page 326
12.3.1.2 Glutathione......Page 327
12.3.1.4 FMN......Page 329
12.3.1.5 CDP and GM3......Page 330
12.3.3.2 Fibrin......Page 331
12.3.3.5 Cytochrome C......Page 332
12.3.3.6 Ubiquitin......Page 333
12.3.3.7 Enzyme Subunits......Page 334
12.3.4 Cofactors as Ligands......Page 335
12.3.5.2 CP-dMM......Page 336
12.3.5.4 2-[3-(2-Ammonioethoxy)benzoyl]ethyltrimethylammonium Bromide......Page 337
12.3.5.7 dATP......Page 338
12.3.5.10 Cilastatin......Page 339
12.3.5.13 t-Butyloxycarbonyl-gly-phe-NHCH2CN......Page 340
12.3.7 Immobilized Transition-State Analogs as Ligands......Page 341
12.4 TANDEM METHODS......Page 342
12.5.1 Isolation of Inhibitory Peptides......Page 343
SYMBOLS AND ABBREVIATIONS......Page 344
REFERENCES......Page 345
13.1 INTRODUCTION......Page 352
13.1.1 General Requirements for the Purification of Recombinant Proteins......Page 353
13.2 IMMUNOAFFINITY CHROMATOGRAPHY......Page 354
13.2.2 Optimizing Immunosorbent Performance......Page 355
13.2.3 Development of an Immunoaffinity Purification......Page 356
13.2.3.1 Importance of Protein Source and Subpopulations......Page 357
13.2.3.2 Sample Pretreatment......Page 359
13.2.3.3 Immunoaffinity Conditions......Page 361
13.3 PSEUDOBIOAFFINITY CHROMATOGRAPHY......Page 362
13.3.1 Histidine PAC......Page 363
13.3.2 Biomimetic Affinity Chromatography......Page 364
13.3.3 Anionic Ligand Affinity Chromatography......Page 365
SYMBOLS AND ABBREVIATIONS......Page 368
REFERENCES......Page 369
CONTENTS......Page 372
14.1.1 Antibodies......Page 373
14.2 ANTIBODY PURIFICATION......Page 374
14.2.2 Physicochemical Techniques for Antibody Purification......Page 375
14.2.2.1 Antibody Precipitation......Page 376
14.2.2.2 General Chromatographic Methods......Page 378
14.2.3.2 Thiophilic Chromatography......Page 381
14.2.3.3 Affinity Separations with Bacterial Coat Proteins......Page 382
14.2.3.3.2 Protein G......Page 384
14.2.3.4 Affinity Separations with Lectins......Page 386
14.2.3.5 Immunoaffinity Purification of Antibodies......Page 387
14.3.1 Physicochemical Approaches for Antigen Purification......Page 389
14.3.1.1 General Chromatographic Methods......Page 390
14.3.1.2 Electrophoretic Techniques......Page 391
14.3.2 Affinity Methods for Antigen Isolation......Page 392
14.4 SUMMARY AND CONCLUSIONS......Page 393
REFERENCES......Page 394
CONTENTS......Page 403
15.1.2 General Immobilization and Elution Methods......Page 404
15.2.1.2 Applications of CaM-Affinity Chromatography......Page 405
15.2.1.4 Preparation of CaM-Sepharose......Page 408
15.2.1.5 Preliminary Purification of CaM Target Proteins......Page 409
15.2.2 Purification of Other Calcium-Dependent Proteins......Page 410
15.3.1 Adenylyl Cyclase......Page 413
15.3.2 GTP-Binding Proteins......Page 414
15.3.4 Cyclic 3´,5´-Nucleotide Phosphodiesterase......Page 416
15.3.5 Cyclic Nucleotide-Dependent Protein Kinase......Page 418
15.3.6 Other Protein Kinases......Page 419
15.3.7 Protein Phosphatases......Page 420
15.4.1 Phosphatidylinositol and Inositol Metabolism......Page 422
15.4.2 Protein Kinase C......Page 425
15.5 OTHER SIGNAL-TRANSDUCTION SYSTEMS......Page 426
SYMBOLS AND ABBREVIATIONS......Page 427
REFERENCES......Page 428
CONTENTS......Page 439
16.2.1 General Principles......Page 440
16.2.2.1 Mass Transfer......Page 441
16.2.2.3 Desorption Kinetics......Page 442
16.3.2 Selection of a Support for RAC......Page 443
16.3.4 Receptor-Affinity Chromatography......Page 444
16.3.5 Procedures for Receptor-Affinity Purification......Page 445
16.3.5.3 Elution, Polishing, and Storage......Page 446
16.3.5.4 Method Validation, Scale-Up, and Automation......Page 447
16.4.1 Recombinant Human rIL-2......Page 448
16.4.2 Human rIL-2 Analogs......Page 449
16.5 MEMBRANE-BASED RECEPTOR-AFFINITY CHROMATOGRAPHY......Page 450
16.5.2 Construction of an IL-2R Affinity Membrane......Page 451
16.5.5 Process-Scale MRAC......Page 453
16.6.1 Immobilization of a Receptor for FB-RAC......Page 454
16.6.2 Determination of Binding Capacities for FB-RAC......Page 455
16.6.3 Example of an FB-RAC Purification......Page 456
SYMBOLS AND ABBREVIATIONS......Page 459
REFERENCES......Page 460
Section IV: Analytical and Semipreparative Applications......Page 462
17.1 INTRODUCTION......Page 463
17.2 ENZYMOLOGY......Page 464
17.3 IMMUNOLOGY......Page 465
17.4 ENDOCRINOLOGY......Page 468
17.5 TOXICOLOGY AND THERAPEUTIC DRUG MONITORING......Page 471
17.6 LIPIDS AND LIPOPROTEINS......Page 473
17.7 CANCER AND TUMOR MARKERS......Page 474
17.8 PROTEINS AND PROTEOMICS......Page 475
17.9 SUMMARY AND CONCLUSIONS......Page 478
REFERENCES......Page 479
CONTENTS......Page 489
18.2.1 Antibodies and Antigens......Page 490
18.2.3 Phosphoryl Groups......Page 491
18.2.5 Glycosyl Groups and Sugar Residues......Page 493
18.2.6 Immunoglobulins......Page 496
18.2.8 Proteins......Page 497
18.2.10 Affinity Tags......Page 498
18.3.1.2 Methods for Downstream Processing......Page 501
18.3.2 Process Monitoring......Page 504
18.4.1.1 Analysis of Complex Mixtures......Page 505
18.4.1.2.1 Primary-Structure Variants......Page 506
18.4.1.2.2 Posttranslational Modifications......Page 508
18.4.1.2.3 Metabolic Studies......Page 509
18.4.3 Diagnostics......Page 511
18.5 SUMMARY AND CONCLUSIONS......Page 513
REFERENCES......Page 514
CONTENTS......Page 519
19.1 INTRODUCTION......Page 520
19.2.2 Examples of Applications......Page 521
19.2.2.2 Analysis of Metal Ions and Metal Ligands by IMAC......Page 522
19.3.1 General Principles......Page 523
19.3.2.2 Herbicide Detection by Sequential-Injection Assays......Page 524
19.3.2.3 Environmental Testing by Flow-Injection Liposome Immunoanalysis......Page 525
19.3.3 Homogeneous Immunoassays......Page 526
19.4.1 General Principles......Page 528
19.4.3 Environmental Testing by On-Line Affinity Extraction and HPLC......Page 529
19.4.4 Analysis of Degradation Products by On-Line Affinity Extraction......Page 531
19.4.5 On-Line Affinity Extraction and Field-Portable Systems......Page 533
19.4.6 On-Line Affinity Extraction with Other Methods......Page 534
19.5 MOLECULARLY IMPRINTED POLYMERS......Page 536
19.6 STUDIES OF MOLECULAR INTERACTIONS......Page 537
19.7 AFFINITY BIOSENSORS......Page 539
19.8 AFFINITY CAPILLARY ELECTROPHORESIS......Page 540
19.9 SUMMARY AND CONCLUSIONS......Page 541
REFERENCES......Page 542
CONTENTS......Page 548
20.2.1 Homopolymer Ligands......Page 549
20.2.2 Heteropolymer Ligands and Subtractive Hybridization......Page 550
20.3.1 Histidine Tags......Page 552
20.3.3 Strep-Tag......Page 554
20.3.5 Glutathione S-Transferase......Page 555
20.4.1 Immunoprecipitation......Page 556
20.4.2 Antibody Binding to Immobilized Bacterial Proteins......Page 557
20.4.3 Immobilized Antibodies and Antigens......Page 559
20.4.4 Cell Isolation......Page 560
20.5.1 General Principles of the ChIP Assay......Page 561
20.5.2.2 Chromatin Solubilization......Page 563
20.5.2.5 Analysis of Immunoprecipitated DNA and Identification of Binding Sites......Page 564
20.6 MULTIPLEXED PARTICLE-BASED FLOW CYTOMETRIC ASSAYS......Page 565
20.6.1 Nucleic Acid Hybridization......Page 566
20.7 SUMMARY AND CONCLUSIONS......Page 567
SYMBOLS AND ABBREVIATIONS......Page 568
REFERENCES......Page 569
CONTENTS......Page 572
21.2 GENERAL EXPERIMENTAL APPROACHES......Page 573
21.3.1 alpha-Chymotrypsin (ACHT)......Page 574
21.3.1.1 Preparation of an ACHT-CSP......Page 575
21.3.1.2 Chiral Separations Based on ACHT-CSPs......Page 576
21.3.2.2 Chiral Separations Based on TRYP-CSPs......Page 578
21.3.2.3 Chromatographic Conditions for TRYP-CSPs......Page 579
21.3.3.2 Chiral Separations Based on CBH I-CSPs......Page 580
21.3.3.3 Chromatographic Conditions for CBH I-CSPs......Page 581
21.4.1.2 Chiral Separations Based on SA-CSPs......Page 582
21.4.1.3 Chromatographic Conditions for SA-CSPs......Page 583
21.4.2.1 Preparation of AGP-CSPs......Page 584
21.4.2.3 Bioanalytical Applications of AGP-CSPs......Page 585
21.4.2.4 Chromatographic Conditions for AGP-CSPs......Page 586
SYMBOLS AND ABBREVIATIONS......Page 589
REFERENCES......Page 590
Section V: Biophysical Applications......Page 594
CONTENTS......Page 595
22.1 INTRODUCTION......Page 596
22.2 ZONAL ELUTION......Page 597
22.2.1 General Principles......Page 598
22.2.2 Common Applications of Zonal Elution......Page 599
22.2.2.2 Competition and Displacement Studies......Page 600
22.2.2.3 Solvent and Temperature Studies......Page 603
22.2.2.4 Determining the Location and Structure of Binding Sites......Page 605
22.2.3 Alternative Applications......Page 606
22.2.3.2 Work with Low-Solubility Compounds......Page 607
22.2.3.4 Studies of Allosteric Interactions......Page 608
22.2.4.1 Choice of Affinity Column......Page 609
22.2.4.2 Choice of Additive Concentrations......Page 610
22.2.4.3 Selection of Sample Size......Page 611
22.2.4.4 Determination of Analyte Retention......Page 612
22.3 FRONTAL ANALYSIS......Page 613
22.3.2.1 Measurement of Affinity and Number of Binding Sites......Page 614
22.3.2.2 Competition and Displacement Studies......Page 616
22.3.2.3 Solvent and Temperature Studies......Page 617
22.3.3.2 Choice of Analyte Concentrations......Page 618
22.3.3.3 Determination of Breakthrough Times......Page 619
22.4.1 Band-Broadening Studies......Page 620
22.4.2 Peak Decay Analysis......Page 621
22.5 SUMMARY AND CONCLUSIONS......Page 623
SYMBOLS AND ABBREVIATIONS......Page 624
REFERENCES......Page 625
CONTENTS......Page 629
23.2 SUMMARY OF BASIC EXPRESSIONS......Page 630
23.2.1 Equations for Univalent Partitioning Solutes......Page 632
23.2.2 Equations for Multivalent Solutes......Page 634
23.3.1 Column Chromatography......Page 635
23.3.3 Biosensor Technology......Page 637
23.4.1 Alternative Derivation of the Scatchard Expression for a Multivalent Solute......Page 640
23.4.2 Linearity of the Conventional Scatchard Plot for a Multivalent Solute......Page 641
23.4.3 The Rectangular Hyperbolic Relationship for a Multivalent Partitioning Solute......Page 642
23.5.1 Studies with Erythrocyte Membranes as the Affinity Matrix......Page 643
23.5.2 Erythrocyte Ghosts as an Affinity Matrix......Page 645
23.6.1 Theoretical Considerations......Page 646
23.6.2 Experimental Applications......Page 647
23.7 IMPACT OF BIOSENSOR TECHNOLOGY......Page 648
23.7.1.2 Allowance for Solute Depletion in Experiments with Cuvette-Based Biosensors......Page 649
23.7.1.3 Contribution of Biosensor Studies to an Understanding of RNA Trafficking......Page 650
23.7.2.1 Pseudo-First-Order Kinetics......Page 652
23.7.2.2 Second-Order Kinetics......Page 653
23.7.2.3 Biacore Analysis of a 1:1 Interaction......Page 654
23.8 SUMMARY AND CONCLUSIONS......Page 655
SYMBOLS AND ABBREVIATIONS......Page 656
REFERENCES......Page 657
CONTENTS......Page 663
24.2 CHIRAL RECOGNITION MECHANISMS......Page 664
24.3.1 alpha-Chymotrypsin......Page 665
24.4 CHIRAL RECOGNITION BY SERUM PROTEINS......Page 668
24.4.1 Human Serum Albumin......Page 669
24.4.2 alpha1-Acid Glycoprotein......Page 672
24.5.1 Determination of Percent Binding and Affinity......Page 674
24.5.2 Characterization of Binding Sites on HSA......Page 675
24.5.3 Characterizing Binding Sites Using Chemically Modified HSA......Page 676
24.5.4 Studies of Drug-Drug Interactions......Page 677
24.5.5 Application of HSA-CSPs in New Drug Approval......Page 678
24.5.6 Solute Binding to Other Serum Albumins......Page 679
SYMBOLS AND ABBREVIATIONS......Page 680
REFERENCES......Page 681
25.1 INTRODUCTION......Page 684
25.2.2 Experimental Design for Biosensors......Page 685
25.2.3 Ligand Immobilization for Biosensors......Page 686
25.2.4 Typical Biosensor Cycle......Page 687
25.3.2 Determination of Active Sample Concentrations......Page 688
25.3.3 Purification and Identification of Biochemicals......Page 690
25.3.4 Characterization of Membrane-Associated Systems......Page 692
25.4.2 Array Systems......Page 693
REFERENCES......Page 694
Section VI: Recent Developments......Page 696
CONTENTS......Page 697
26.1 INTRODUCTION......Page 698
26.2 GENERAL USE OF LIGANDS IN CE......Page 699
26.3 THEORY OF CE WITH AFFINITY LIGANDS......Page 700
26.4 GENERAL TYPES OF LIGANDS IN CE......Page 703
26.4.1 Free Ligands......Page 704
26.4.2 Soluble Modified Ligands and Ligand Constructs......Page 706
26.5.1 Selectivity Enhancement......Page 707
26.5.2 Quantitative Assays......Page 712
26.5.3 CE Binding Studies......Page 714
26.6 SPECIFIC LIGANDS USED IN CE......Page 716
26.7.1 CE-MS......Page 717
26.7.2 CE-NMR......Page 718
26.7.4 Other Detection Schemes......Page 719
SYMBOLS AND ABBREVIATIONS......Page 720
REFERENCES......Page 722
CONTENTS......Page 734
27.1 INTRODUCTION......Page 735
27.2.1 Off-Line Methods......Page 736
27.2.2 Electrospray Ionization Mass Spectrometry......Page 737
27.2.3 Matrix Assisted Laser Desorption Ionization Mass Spectrometry......Page 738
27.2.5 Inductively Coupled Plasma Mass Spectrometry......Page 741
27.3.1 Mass Spectrometry with Immunoextraction......Page 742
27.3.2 Lab-on-Valve Systems......Page 743
27.3.4 On-Line Affinity MALDI......Page 744
27.4.1 Screening Combinatorial Libraries......Page 745
27.4.2 Proteomic Studies......Page 746
27.5.1 Gas-Phase Studies......Page 748
27.5.2 Solution-Phase Studies......Page 749
27.6.2 Mass Spectrometric Immunoassays......Page 750
27.7 SUMMARY AND CONCLUSIONS......Page 751
REFERENCES......Page 752
CONTENTS......Page 759
28.2 MANUFACTURING MICROFABRICATED DEVICES......Page 760
28.2.1 Micromachining, Embossing, and Imprinting......Page 762
28.2.2 Lithography......Page 763
28.3.1 Static Devices......Page 764
28.3.2 Devices Based on Microfluidics......Page 766
28.3.2.2 Chromatographic Devices......Page 767
28.3.2.2.2 Design of Components for Microchromatography......Page 768
28.3.2.2.3 Pumps......Page 769
28.3.2.2.5 Microcolumn Construction......Page 770
28.3.2.3 Lab-on-a-Chip......Page 771
28.4 DETECTION SYSTEMS......Page 774
28.4.2 Fluorescence......Page 775
28.4.5 Electrochemical Detection......Page 776
28.5 SUMMARY AND CONCLUSIONS......Page 777
SYMBOLS AND ABBREVIATIONS......Page 778
REFERENCES......Page 779
CONTENTS......Page 784
29.1 INTRODUCTION......Page 785
29.2.2 Fluorescence......Page 786
29.2.3 Enzyme Labels......Page 789
29.2.4 Chemiluminescence......Page 791
29.2.6 Radiolabels......Page 793
29.2.7 Electrochemical Detection......Page 795
29.2.9 Miscellaneous Methods......Page 796
29.3.1 General Principles......Page 797
29.3.2 Applications......Page 798
29.3.3 Theory......Page 800
29.3.4 Practical Considerations......Page 801
29.4.1.1 General Principles......Page 802
29.4.1.3 Theory......Page 804
29.4.1.4 Practical Considerations......Page 806
29.4.2.2 Applications......Page 807
29.4.2.3 Theory......Page 809
29.4.2.4 Practical Considerations......Page 810
29.4.3.1 General Principles......Page 811
29.4.3.2 Applications......Page 812
29.4.3.3 Theory......Page 813
29.5.1 Sandwich Immunoassays......Page 814
29.5.1.1 General Principles......Page 815
29.5.1.2 Applications......Page 816
29.5.1.3 Theory......Page 817
29.5.2.1 General Principles......Page 818
29.5.2.3 Theory......Page 820
29.5.2.4 Practical Considerations......Page 822
SYMBOLS AND ABBREVIATIONS......Page 823
ACKNOWLEDGMENT......Page 824
REFERENCES......Page 825
CONTENTS......Page 832
30.2.1 General Principle of Molecular Imprinting......Page 833
30.2.2.1 Acrylic and Vinyl Polymers......Page 835
30.2.3 Target Molecules......Page 837
30.2.4 Physical Forms and Preparation Methods for MIPs......Page 838
30.2.4.2 Thin-Imprinted Polymer Films......Page 839
30.3.1 Liquid Chromatography......Page 840
30.3.3 Thin-Layer Chromatography......Page 842
30.3.4 Solid-Phase Extraction......Page 843
30.3.5 Membrane-Based Separations......Page 844
30.4 SUMMARY AND CONCLUSIONS......Page 845
REFERENCES......Page 846