Immunoassay

Front Cover......Page 1
Immunoassay......Page 4
Copyright Page......Page 5
Contents......Page 8
Contributors......Page 28
Preface......Page 32
CHAPTER 1. PAST, PRESENT AND FUTURE OF IMMUNOASSAYS......Page 34
1. Cellular and Molecular Basis of the Humoral Immune Response......Page 38
2. Antibody Structure......Page 44
3. Antigen Binding......Page 51
4. Antibody Structure and Immunologic Diversity......Page 53
5. Conclusion......Page 54
1. Binding Theory......Page 58
2. Equilibria Involving Many Ligands and a Heterogeneous Population of Binding Sites......Page 61
3. Principles of Saturation Analysis......Page 62
4. Kinetics of Immunoassay......Page 67
5. The Theory of Two-Site Immunoassays......Page 71
6. Graphical Presentation of Immunoassay Data......Page 73
7. Curve Fitting of Immunoassay Data......Page 75
8. Detection Limit of Immunoassays......Page 80
References......Page 82
1. Introduction......Page 84
2. Assay Validation......Page 86
3. Determining Immunoassay Sensitivity......Page 90
4. Tests of Similarity (Parallelism)......Page 95
5. Alternative Methods for Defining Cross Reactivity......Page 115
6. Quality Control......Page 116
References......Page 125
CHAPTER 5. PRODUCTION AND PURIFICATION OF ANTIBODIES......Page 128
2. Polyclonal versus Monoclonal Antibodies......Page 129
3. Choice of Animal......Page 130
4. Immunization Procedures......Page 134
5. Obtaining and Processing Blood and Lymphocytes......Page 137
6. Monoclonal Antibody Production......Page 139
7. Antibody Purification......Page 145
References......Page 148
1. Introduction......Page 150
2. Cloning Antibody Variable Region Genes......Page 151
3. Expression of Recombinant Antibodies......Page 157
4. Antibody Structure-Function Studies......Page 162
5. Therapeutic Antibodies......Page 170
6. Designer Antibodies......Page 183
7. Future Directions......Page 188
References......Page 190
1. Introduction......Page 198
2. Detecting Interferences......Page 199
3. Cross-Reactivity and Heterogeneous Analytes......Page 201
4. Reducing Cross-Reactive Interferences......Page 204
5. Heterophile and Anti-Animal Antibodies......Page 207
6. Recognizing and Reducing Interference from Endogenous Antibodies......Page 215
7. Interferences Due to Masking of Antigens......Page 218
9. Matrix Effects......Page 219
References......Page 220
1. Introduction......Page 224
2. Requirements for Quality of Labeled Reactants......Page 225
3. Chemical Reactions for Conjugation......Page 226
5. Conjugation of Antibody IgG to Enzymes......Page 227
6. Conjugation of Antibody Fragments to Enzymes......Page 229
7. Conjugation of Antigens, Avidin, and Streptavidin to Enzymes......Page 234
8. Conjugation of Small Molecules to Antibodies and Antigens......Page 235
References......Page 237
1. Interfacial Immunochemistry......Page 238
2. Diversity of Solid Phases......Page 246
3. Immobilization Procedures......Page 249
4. SPI in the Future......Page 255
References......Page 256
1. Introduction......Page 260
2. Competitive Immunoassays......Page 261
3. Noncompetitive Immunoassays......Page 262
4. Indirect Detection of the Immunocomplexes......Page 266
References......Page 269
1. Introduction......Page 270
2. Components of the System......Page 271
3. Amplified Detection Systems......Page 285
5. Auxiliary Enhancement Systems......Page 288
6. Selected Protocols......Page 292
References......Page 298
CHAPTER 12. RADIOIMMUNOASSAY......Page 302
2. Counting of Radioactive Isotopes......Page 303
6. Iodination Methods......Page 306
7. Iodination Damage......Page 311
9. Purification of Iodinated Tracer......Page 312
10. Chemical Evaluation of Tracer......Page 313
11. Immunological Evaluation of Tracer......Page 315
12. Variations on the Use of Radiolabeled Tracers......Page 316
Reference......Page 317
1. Introduction......Page 320
2. Enzymes as Labeling Substances......Page 321
3. Conjugation Procedures......Page 323
4. Determining Enzyme Activity......Page 327
5. The Variety of Enzyme Immunoassays......Page 332
6. Prospects......Page 337
References......Page 338
1. Introduction......Page 342
2. Principles of Fluorescence Spectroscopy......Page 343
4. Fluorescent Labels......Page 347
5. Homogeneous Fluorescence Immunoassays......Page 349
6. Heterogeneous Fluorescence Immunoassays......Page 355
References......Page 366
1. Introduction......Page 370
2. Chemiluminescent Labels and Labeling......Page 372
4. Types of Chemiluminescent Immunoassay......Page 378
5. Chemiluminescent Detection of Enzyme Labels......Page 380
References......Page 384
1. Introduction......Page 388
2. Principles of Bioluminescent Light Emission......Page 389
5. Bioluminescence Enzyme Immunoassays......Page 390
6. Other Bioluminescence Immunoassay Systems......Page 392
References......Page 393
1. Introduction......Page 396
2. Historical Perspective......Page 397
3. Characteristics of the Immunoprecipitin Reaction......Page 399
4. Principles of Light Scattering......Page 401
5. Definition and Principles of Nephelometry and Turbidimetry......Page 403
6. Instrumentation......Page 414
References......Page 418
1. Introduction......Page 422
2. Labels and Immunoassay Formats......Page 423
3. Labels......Page 424
4. Discrete Test Zones......Page 427
6. Conclusions......Page 432
References......Page 434
1. Introduction......Page 438
2. Anti-idiotypic Antibodies as Novel Probes in Immunodiagnostics......Page 439
3. Idiometric Assays for Estradiol......Page 445
4. Idiometric Assays for Progesterone......Page 449
5. Conclusions......Page 452
References......Page 453
CHAPTER 20. FREE HORMONE MEASUREMENTS......Page 456
1. Introduction......Page 457
2. Overview of Thyroid, Steroid, and Secosteroid Hormones......Page 459
3. Transport of Thyroid, Steroid, and Secosteroid Hormones in the Blood......Page 461
4. The Theory of Free Hormones and Alternative Concepts......Page 471
6. Classification of Free Hormone Assays......Page 476
7. Principles of Free Hormone Assays......Page 484
8. Total Hormone/Total Hormone-Binding Globulin Ratio......Page 493
9. Tests Based on the Measurement of "Thyroid Hormone Binding"......Page 494
10. Calculated Free Hormone Concentrations......Page 496
12. Assay Parameters of Free Hormone Assays......Page 497
13. Specimens for Free Hormone Assays......Page 502
14. Interferences with Free Hormone Measurements......Page 503
15. Selected Issues Regarding Interpretation of Free Hormone Results......Page 507
17. Quality Control of Free Hormone Assays......Page 508
References......Page 509
1. Introduction......Page 516
2. Automation of Immunoassay......Page 517
3. Automated Immunoassay Systems......Page 521
4. Performance of Immunoassay Systems......Page 524
5. Future Trends......Page 534
References......Page 535
1. Introduction......Page 538
2. Basic Features of Dry Reagent Assays......Page 540
3. Examples of Thin-Film Immunoassays......Page 551
References......Page 566
1. Introduction......Page 570
2. Principles......Page 571
3. The Basic Protocol......Page 572
4. Detection Methods......Page 576
5. Specialized Uses and Applications......Page 582
Reference......Page 585
1. Introduction......Page 588
2. Development of an Immunoassay Method......Page 589
3. A Practical Example--An ELISA for p53......Page 596
References......Page 600
INDEX......Page 602