CONTENTS - Contributors xx Preface xxi Acknowledgements xxiv Foreword xxv

PART 1 PRINCIPLES

1 Introduction to lmmunoassay Principles Chris Dauies Immunoassay for beginners Kinetics of antibody-antigen interactions lmmunoassay design

Competitive (reagent limited) assays Single-site irnmunometric assays Two-site immunometric assays (reagent excess) Determinants of assay sensitivity Detection and quantification of antibodies Special considerations for solid-phase imrnunoassays Comparison of experimental and theoretical immunoassay performance

2 Non-Competitive lmmunoassays for Small Molecules - The Anti-Complex, Selective Antibody and Apposltlon Systems Colin H. Self, Stephen Thompson and Larry A. Winger lntroduction The anti-complex assay

--=*miry------ Multiple binding assay Selective antibody immunomeMc assay Apposition assay Conclusions

3 Ambient Analyte Assay Roger Ekins Introduction Basic theoretical principles Applications of the ambient analyte assay principle - -

Microspot assays Free (non-protein bound) hormone immunoassays Other applications of the principle

Summary and conclusion

4 Free Analyte lmmunoassay Nic D. Christofides Basic principles governing the free hormone concentration Calculation of free analyte concentration Spreadsheet for calculation of free analyte concentration Effect of serum proteins on free analyte concentration in uftm measurement of free analyte concentration

Direct equilibrium dialysis Immunoassays for free analytes Tests of validity (accuracy)

Concluding remarks

The lmmunoassay Handbook

5 The Foundations of Immunochemistry Robert F. Ritchie Introduction The immunological reaction (1895-1935)

Summary Qualification by diffusion in gel (1861-1977)

Summay Qualitative analysis - by immunoelectrophoresis (1953-1978)

Summary Quantification of antigens by in-gel immunochemistry (1963-present)

Summay Quantification of antigens by in-liquid immunochemistry (1935-present)

Changes in immunization practice Summay

Quantification of antigens by particle-enhanced immunochemistry (1972-present) Summary

Labeled immunoassay (1959-present) Summay

Comparative studies (1907-present) Summay

6 Concepts Chris Dauies Assay concepts

Assay sensitivity Accuracy and bias Precision and imprecision Cross-reactivity Interference Recovey Dilution Correlation and method comparison Assay drift

Clinical concepts Diagnosis Reference interval Bias Clinical sensitivity and specificity Positive and negative predictive values Prevalence and clinical utility Likelihood ratio Continuous measurement, ROC curves Confidence interval analysis of clinical performance Probabilistic interpretation of test results Analytical goals

Summary

7 lmmunoassay Development in the In Vitro Diagnostic Industry Doug Brandt and Steve Figard Assay design overview

The needs of the customer The capability of the technology The requirements for consistent manufaduring

Experimental design in immunoassay development Preamble The process in overview The problems assuaged by DOE The particulars

Four key assay design principles

Contents

Plan first Think from a theoretical perspective Simplify the approach Establish specifications before starting

Concluding comments

8 Antibodies E y l Liddell Introduction Antibody structure The antibody response in vfuo Antibody diversity Immunization

Immunogen Polyclonal antisera

Purification of antisera Monoclonal antibodies

Immune lymphocytes Myeloma cell lines Cell fusion Hybrid selection Antibody screening tests Cloning Storage and propagation Human monoclonal antibodies Purification of monoclonal antibodies

Antibody fragments Bispecific antibodies Chimeric and humanized antibodies Recombinant antibodies

Recombinant phage antibody library construction Selection of specific phage antibodies Altering antibody characteristics Synthetic recombinant antibodies Bivalent and bipecific recombinant antibodies

Antibody conjugates and fusion proteins in diagnostics Polyclonal, monoclonal or recombinant?

9 Standardization and Calibratlon Dauid Wild Standardization

The role of external quality assessment (proficiency testing) schemes International standards Definitive and reference methods Other reference materials Heterogeneity of standard material Method-related causes of standardization diierences Special considerations for assay of antibodies

Calibration Analyte International standards Secondaw standards Calibrators Standard and calibrator matrices Reduced and stored calibration curves Recovery and dilution

10 Separation Systems David Wild and WIad Kusnezow Liquid-phase separations

Electrophoresis

iii

The lmmunoassay Handbook

Gel filtration Dextran-coated charcoal Precipitation by salts, organic solvents and polyethylene glycol (PEG) Double (second) antibody precipitation PEG-assisted second antibody precipitation Aspiration and decantation methods

Surface-coated solid phases General principles of protein binding to plastic surfaces Covalent attachment of proteins to solid phases Glass and plastic particles Magnetizable particles Tubes, wells and microtitration plates Beads Common solid phase Membrane tiltration Immunochromatography Western blot

Washing Competitive assays Immunometric assays Removal of interfering substances The mechanics of washing

Microarray Immunoassay Separation Basic principles of receptor ligand interaction on microarray Solid-phase medium Protein attachment on microarrays

11 Signal Generation and Detection Systems (Excluding Homogeneous Assays) Larry J. Kricka and David Wild Radioactive labels

Radioactivity measurement Preparation of radioactive tracers

Enzyme labels Colorimetry Fluorometty Chemiluminescence Enhanced chemiluminescence

Direct fluorescence Long wavelength fluorescence

Time-resolved fluorescence Direct chemiluminescence Bioluminescence Phosphorescence Microparticle labels StreptavididAvidin-Blotin Protein A Amplification strategies Multiple analytes and miniaturization

Microarrays

12 Homogeneous lmmunoassays Edwin F. Ullman Introduction Particle agglutination

Erythrocytes and latex Gold sols

Lysis immunoassays Spin immunoassays

Contents

Fluorescent labels Fluorescence polarization immunoassay Fluorescence resonance energy transfer Fluorescence protection assays Fluorescence fluctuation

Enzyme immunoassays Enzyme-multiplied immunoassay technique (EMITm) Charge induced enzyme activation Enzyme channeling

Enzyme effector immunoassays Subsirate linked fluorescence immunoassay (SLFIA) Enzyme cofactor immunoassay Enzyme inhibitor immunoassay Enzyme complementation immunoassay

Isotopic labels Scintillation proximity assay

Electroactive labels Electrochemical detection Electrochemiluminescence

Oxygen channeling immunoassays Luminescent oxygen channeling immunoassay

Conclusion

13 Calibration Curve-fitting Bany Nix and David Wild Introduction

Dose-response metameter Response-error relationship Homo- and heteroscedasticity Linearizing transformations Normalizing transformations Outliers Empirical and theoretical curve-fitting methods Least-squares fitting procedure Weighted least-squares procedure Precision profiles

Examples of calibration curve-fitting methods Hand-plots Linear interpolation Spline fits Polynomial regression Logit-log and four-parameter log-logistic methods Five-parameter log-logistic Four-parameter law of mass action

Monitoring the quality of curve-fitting Stored calibration curves, factory master c w e s , and adjusters

Master calibration curve Adjusters Modeling calibration curve changes over shelf life Use of electronic data transmission Summary

Suitable calibration curve-fit software

14 Conjugation Methods Alastair H. Dent Categories of conjugates employed in immunoassay Protein-protein coupling

Functional chemistry of proteins Categories of protein-protein coupling reaction Common protein-protein coupling methods

The lmmunoassay Handbook

Genetic engineering approaches to protein conjugation Protein-small molecule coupling

Introduction Common protein-small molecule coupling methods

Purification of conjugates Chromatographic approaches to conjugate purification Other approaches to conjugate purification

Characterization of conjugates Conclusion

15 immunological Biosensors James K. Gimzewski, Jason Reed, Michael A. Teitell and P. Gordon Malan Overview Electrochemical sensors

Arnperometric sensors Potentiometric sensors

Nanomechanical sensors Piezoelectric mass sensors Microcantilever sensors Micromagnetic sensors

Optical sensors Fluorescent evanescent wave sensors Integrated optical sensors Quantum dots

Conclusions and future directions

16 Surface Plasmon Resonance in Kinetic, Concentration and Binding Site Analyses --

~TKarlsson~erfmWaan~a-rgFPn*----------- lntroduction

Objectives of protein studies in basic research Protein studies and therapeutics Technology demands

SPR biosensors Technology principles Sensorgrams and report points lmmobiition Surface activity and immobilization levels

Binding site analysis - epitope mapping Pair-wise epitope mapping

Kinetic analysis SPR in kinetic analysis

Concentration analysis Direct binding assays Inhibition assays

Summary

17 Lab-on-a-Chip, Micro-, and Nanoscale lmmunoassay Systems Larry J. Kricka and David Wild Fundamental Uerences due to miniaturization

Viscosity and surface tension Capillary flow Electroosmosis The effects of reduced volume on low concentration samples Effect of reduced volumes on kinetics

Immunoassay design at micro- and nanoscale Assay format Antibodies

Contents

Standardization and calibration Separation Signal generation Homogeneous immunoassay On-chip detection methods Bioelectronic chips and irnmunoassay Microarrays and irnmunoassay Atomic force microscopy

ManufacMng and business considerations Conclusions

PART 2 PRODUCT TECHNOLOGY

18 Introduction to Product Technology in Clinical Diagnostic Testing David Wild Immunoassay product technologies

Use of fluorescence-labeled antibodies Agglutination assays Radioirnmunoassay and immunoradiometric assay Heterogeneous enzyme immunoassay Homogeneous enzyme immunoassay Automated homogeneous batch analyzer Automated heterogeneous batch analyzer Non-radioactive systems with increased sensitivity Semi-automated systems Automated dual technology batch analyzer Automated, multianalyte batch analyzers Random-access analyzers with bulk reagent packs Unitized random-access systems Clinical chemistry analyzers Near-patient tests Home-use tests Other applications Future developments

19 Market Trends David Huckle and David Wild lmmunoassay market status Established trends

Market drivers Advances in technology New analytes

The future of the immunodiagnostics business Application of marketing theory to immunodiagnostics Future market requirements Changes in the customer base Potential impact of new technologies New analytes

Conclusion Summary of likely trends In immunodiagnostics and related products

20 Choosing an Automated lmmunoassay System Lori J. Sokoll and Daniel W. Chan Defining automation goals Sources of information Considerations and criteria

Laboratoty environment considerations Test menu considerations

The lmmunoassay Handbobk

Technical and clinical considerations Financial considerations Operational considerations

Summary

21 Automated System Features Alain Truchaud, Tanguy Le Neel, Murielle Cazaubiel, Bernard capolaghi and Jean-Pierre Yuert Introduction Specific requirements of immunoassays Concepts in immunoassay automation: workstations vs. integrated systems Reagent features

Infrequent calibration Traceability Multiselective testing and continuous operation

Sample features Sample integrity Biosafety Dilution and reflex testing Stat testing

Incubation and washing Measurement Total quality managemenffbaceability Maintenance Ergonomics Integrated automation Conclusion

22 Over-the-Counter Pregnancy Test Kits Michael J. Wheeler -

Introduction Evaluation of OTC kits User preferences Factors affecting the measurement of hCG in urine

Summary

23 Fluorescence Microscopy: MicroTrakm Tom Houts

24 Automated Batch Analyzers: IMX@ Kent Ford

25 Bulk Reagent Random-Access Analyzers: U ~ ~ C A P @ 100 Gareth Evans and Mats Rilven

26 Automated Panel Analyzers PRISM^" Dinesh Shah and Jim Stewart

27 Unitized Reagent Random-Access Analyzers: IMMULITE~ and IMMULITE 1000 Arthur L. Babson

28 Bulk Reagent Random-Access Analyzers: ~cS:180@' SE Ehio Gramignano

29 Bulk Reagent Random-Access Analyzers: AxSYM" Theresa Donahoe

30 Bulk Reagent Random-Access Analyzers: ~lecsys@' lmmunoassay Systems May Beth Myers

31 Bulk Reagent Random-Access Analyzers: ~itros'@ ECi David Wild

32 Bulk Reagent Random-Access Analyzers: IMMULITE@ 2000 and IMMULITE 2500 Arthur L. Babson

Contents

33 Bulk Reagent Random-Access Analyzers: ADVlA centauraP Laura Taylor and Elvio Gmmignano

34 ~ r c h i t e c t ~ R000@ and D000& Analyzers Frank A. Quinn

35 CEDIA~, a Homogeneous Enzyme lmmunoassay System William A. Coty and Rueyming Loor

36 Clinical Chemlstry Analyzers: v ITRos~~@ Immuno-Rate and ~ i c r o ~ i p ~ ~ Assays Susan J. Danielson and David A. Hilborn 418

37 Near-Patient Tests: The TRIAGE@ System Kenneth F. Buechler

38 Near-Patient Tests: stratusm CS Acute careTM Diagnostic System W.N. McLellan 43 1

39 Over-the-counter Tests: Clearblue Pregnancy TestTM, Clearblue Ovulation TestTM and ~ l e a r v i e w ~ Keith May

40 Over-the-counter Tests: Persona Keith May

PART 3 LABORATORY MANAGEMENT

41 Subject Preparation, Sample Collection and Handling CoHn Wilde Subject state and preparation

Stress Exercise Food and drink Posture Medical procedures Drugs Pregnancy Age Race

Timing Biological rhythms Dynamic tests Pathological changes Nonthyroidal illness Therapeutic drug monitoring

Blood collection by venepuncture Precautions relating to the patient Phlebotomy techniques Blood collection Preparation of serum Preparation of plasma Whole blood

Interference by tube and stopper components Interference Precautions

The use of serum separators Characteristics of separators Interference Other additives Hemolysis

Collection of blood by skin puncture

The lmmunoassay Handbook

Skin puncture sites Collection into capillary tubes

Urine collection Containers Preservative Types of collection Instructions Sample handling

Saliva Procedures for sampling saliva Special precautions

Cerebrospinal fluid Formation of cerebrospinal fluid Collection of CSF

Amniotic fluid Collection of amniotic fluid Amniocentesis Special precautions

Sweat Semen Hair Milk Storage and transportation Conclusions

42 Laboratory Quality Assurance Pierre Block and Manuella Martin Staff selection and training

F i e d volume pipettes Repeating pipettes Semi-automated instruments Manual dilutions

Equlpment maintenance and testing Laboratory water

Water-purification methods Water purity measurement Recommended water quality

Sample control Sample collection Centrifugation Storage

Reagent and control preparation Assay quality control

Within-assay precision Between-assay precision Internal QC programs QC software on automated analyzers New generation QC software programs QC parameters other than controls QC in infectious disease tests

Point-of-care tests

43 Point-of-Care Testing James H. Nichols Delivery options

Terminology Sites Operators Menu Economics

Contents

Clinical outcome Quality assurance

Components of good laboratory practice Regulations Data management

Practical management Technical validation Quality control Administration

Summary

44 lmmunoassay Troubleshooting Guide David Wild I n s ~ c t i o n s Troubleshooting guide

Control bias - consistent change in values from one period of time to another (or from one reagent lot to another) Gradual change in control values (not consistent change at one time or with change in reagent lot) Commercial controls consistently biased from target mean (not change in values across same lot of controls) Bias in proficiency testing (external quality assessment) scheme from other users of the same assay Bias in proficiency testing (external quality assessment) scheme of one method from all-laboratoy mean or reference method Poor within-assay precision Poor between-assay precision or individual control failure Assay drift Low signal level Low binding in a competitive immunoassay Low binding in an immunometric assay Increase in binding in a competitive immunoassay Reduction in EDso (estimated dose at 50% of binding at zero concentration) - competitive immunoassays only Increase in EDm (estimated dose at 50% of binding at zero concentration) competitive immunoassays only High non-specific binding Poor linearity of dilution Unexpected or inconsistent clinical classification Apparent shift in reference interval (normal range) Negative patient sample concentrations Poor recovery Poor assay sensitivity Poor correlation between two irnmunoassays

PART 4 APPLICATIONS

45 Thyroid Rhys John, Carole A. Spencer, Nic Christofides and David Wild Normal thyroid function Clinical disorders

Hypothyroidism (thyroid hormone deficiency) Hyperthyroidism Nontoxic goiter Nonthyroidal illnesdsick euthyroid syndrome L-thyroxine replacement therapy Endogenous antibodies to T3 and T4 Familial dysalbuminemic hyperthyroxinemia

Analytes Thyrotropin

The lmmunoassay Handbook

Thyroxine T3 or T uptake and calculation of free thyroxine index Free T4 Triiodothyronine Free T3 Thyroxine-binding globulin Thyrofxopin receptor antibodies Thyroid peroxidase antibodies Thyroglobulin antibodies Thyroglobulin

Thyroid testing strategies

46 The Adrenal Cortex Sami Medbak Normal adrenocortical function Clinical disorders

Hypercortisolemia Hypocortisolemia

Analytes Cortisol Adrenocorticotropic hormone

Management of patients 4 t h Cushing's syndrome Pre-admission Admission to endocrine ward (e.g. on Monday)

47 Bone Metabolism Kay W. Colston and John C. Stevenson Normal calcium metabolism Clinical disorders

Hypercalcernia Parathyroid disorders Hypocalcemia Vitamin D disorders Medullary carcinoma of the thyroid Paget's disease Osteoporosis

Analytes Vitamin D metabolites: 25-hydroxyvitamin D,1,25-dihydroxyvitamin D Parathyroid hormone Parathyroid hormone-related protein Calcitonin

Biochemical markers of bone turnover Markers of bone formation Markers of bone resorption Serum tartrate resistant acid phosphatase (TRAP)

Conditions associated with changes in bone markers

48 lnfertllity Michael J. Wheeler Clinical disorders

Primary hypogonadism in the female Secondary hypogonadism in the female Infertility and normal menstrual function Primary hypogonadiim in the male Secondary hypogonadism in the male Impaired sperm transport and sperm viability

Analytes Luteinizing hormone (lutropin) Follicle stimulating hormone (follitropin) Proladin

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Inhibin Estradiol Progesterone Testosterone Dihydrotestosterone

Test strategy for infertility in women

49 In Vitro Fertiilzation and Embryo Transfer (IVF-ET) August C. Oliuar Stage I: induction of ovulation Stage 11: oocyte retrieval Stage 111: oocyte insemination and incubation Stage IV: embryo transfer

Other assisted reproductive technology procedures Gamete intra-Fallopian tube transfer Zygote inh-Fallopian transfer

Micromanipulation of gametes Assisted hatching Intracytoplasmic sperm injection

Analytes Estradiol

50 Hirsutism and Vlriiization in the Female Michael J. Wheeler Clinical disorders

Polycystic ovarian syndrome Androgen-secreting tumors of the ovary Congenital adrenal hyperplasia Cushing's syndrome

Analytes Luteinizing hormone and follicle-stimulating hormone Testosterone Sex hormone-binding globulin Free testosterone Androstenedione Dehydroepiandrosterone sulfate 17a-Hydroxyprogesterone Dihydrotestosterone Androstanediols and their glucuronides

51 Pregnancy Tim Chard Clinical disorders

Detection of early pregnancy Threatened abortion Ectopic pregnancy Chromosome defects of the fetus Neural tube defects of the fetus Premature labor Placental insufficiency Pre-eclampsia Miscellaneous disorders

Analytes Alphafetoprotein Human chorionic gonadotropin Estriol Pregnancy-associated plasma protein-A Other analytes Screening for Down's syndrome

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52 Growth and Growth Hormone Deficiency Michael Preece and Jane Pringle Normal childhood growth Clinical disorders

Growth hormone deficiency or insufficiency Growth hormone resistance Excessive growth hormone secretion

Anal ytes Growth hormone Growth hormone binding protein Insulin-like growth factors: IGF-I (somatomedin C) and IGF-I1 Insulin-like growth factor binding proteins

General strategy

53 Diabetes Mellltus Penny Clark Analytes

Insulin, proinsulin(s) and C-peptide Glycohemoglobin (GHb) Other glycated proteins Microalbumin Autoimmune and other a s ~ ~ y s

54 Hematology Derek Dawson, Hany Waters and John Ardem Normal blood function Clinical disorders

Anemia Polycythemia Iron overload and hemochromatosis Hemostatic disorders Leukemia and lymphoma

Analytes Anemia Intrinsic factor antibody Ferritin Transferrin Transferrin receptors Erythropoietin

Thrombosis and hemostasis Thrombophilia

Proteins C and S Antithrombin

Coagulation Coagulation factors Fibrinogen

Evidence of disseminated intravascular coagulation FibrinogenIFibrin degradation products D-Dimer test

Infrequently used and research assays P-Thromboglobulin Platelet factor 4 Heparin cofactor I1 Plasminogen Tissue plasminogen activator

lmmunodetection methods Leukemia and lymphoma analytes Malarial parasites Hemoglobinopathies

Contents

55 Cardiac Markers Alan H.B. Wu Normal heart function Clinical disorders

Coronary artery disease Heart failure Hypertension

Analytes Creatine kinase and the MB isoenzyme Myoglobin Cardiac troponin (T and I) Free btty acid binding proteins and carbonic anhydrase III isoenzyme Apolipoproteins AI, AII, and B Lipoprotein (a) Homocysteine C-reactive protein Brain natriuretic peptide and NT-PRO-BNP Plasma renin Plasminogen activators and inhibitors

56 Cancer Markers Mavanur R. Suresh Introduction History and classification

Nomenclature and idiosyncrasies of glycoprotein tumor antigens New developments

Analytes Carcino-embryonic antigen a-fetoprotein Carbohydrate antigen 19-9 (sialyl Lewisa) Cancer antigen 125 (Muc-16) Cancer antigen 15-3 Estrogen receptor and progesterone receptor Fecal occult blood Prostate-specific antigen Prostatic acid phosphatase Pz-Microglobulin Neuron-specific enolase Carbohydrate antigen 50 Squatnous cell carcinoma antigen Tissue polypeptide antigen, tissue polypeptide specific antigen and Cyfra 21-1 Placental alkaline phosphatase Human chorionic gonadotropin Cathepsin D Interleukin-2 receptor C-Erb B-2 (Her-2/Neu) oncoprotein P53 Bladder tumor antigen Immunochromatography assays for tumor markers Free light chain (FLC) assays

Novel experimental and other minor markers Cancer antigen 195 (CA 195) Tumor-associated glycoprotein 72.4 (TAG 72.4)

sialyl s-lOO Lewi (SLX-I) Mucin-like carcinoma-associated antigen (MCA) Cancer-associated antigen 549 (CA 549) Bone alkaline phosphatase B/T gene rearrangement test

The lmmunoassay Handbook

BRCAl and BRCA2 P21 or RAS DR-70 9 0 W - 2 BP Ekl-2 NMP-22 MMP assays TAG-12 CA 1-18 Angiogenic factors Chromogranin A Telomerase Urinaiy bladder cancer (UBC) antigen HHV-8 antibody Nor-IMetanephrine RIA HER2/neu PML protein

57 Allergy Lars Yman Allergic disease

Diagnosis and therapy Allergens

Analytes Total serum IgE Allergenspecific IgE antibody IgG antibody assays IgA antibody assay Markers of cell activation

Standardization and evaluation Quantitative measurement of allergen-specific IgE antibodies Reference values Quality control

Clinical applications Diagnosis of atopic disease and identification of offending allergen Prediction and monitoring of the development of allergic disease

58 Autoimmune Disease David F. Keren Analytes

Anti-nuclear antibodies Anti-double-stranded (ds) DNA Anti-Sm Anti-RNF' Anti-SSA/Ro Anti-SSB/La Anti-histone Anti-DNP Anti-centromere Anti-scl-70 Rheumatoid factor Anti-cyclic citrullinated (anti-CCP) peptide Anti-neutrophil cytoplasmic antibodies (C-ANCA, P-ANCA) Anti-glomerular basement membrane (GBM) AntiJo-1 Anti-microsomal (thyroid peroxidase - TPO) Islet cell autoantibodies (ICA) Anti-adrenal cortical antibodies

Contents

Anti-parietal cell antibodies (PCA) Anti-mitochondrial antibodies (AMA, or M2) Anti-smooth muscle antibodies (SMA) Anti-liver - kidney microsomal (LKM) IgA anti-endomysium IgA anti-tissue transglutaminase (anti-tTG) IgG and IgA anti-gliadin Anti-acetylcholine receptor (ACHR) Striational antibodies Calcium channel antibodies Anti-cardiolipin antibodies Anti-myelin oligodendrocyte glycoprotein (MOG) and anti-myelin basic protein (MBP) antibodies

59 Sexually Transmitted Diseases Bruce J. Dille, Alan S. Armstrong and lsa K. Mushahwar Neisseria gonorrhoeae Chlamydia trachomatis Trichomonas uaginalis Candida albicans - vaginitis Gardnerella vaginalis Herpes simplex virus type 2 Syphilis (Treponema pallidum)

60 Congenital Diseases of Microbiological Origin Bruce J. Dille, John W. Saford Jr. and lsa K. Mushahwar Cytomegalovirus Rubella virus Toxoplasma gondli IgG avidity tests Parvovirus B19 Group B streptococci

61 Hepatitis Isa K. Mushahwar Hepatitis A virus, anti-HAV IgM, anti-HAV IgG Hepatitis B virus, HBsAg, HBcAg, HBeAg, anti-HBs, anti-HBc, anti-H& IgM, anti-HBe Hepatitis C virus Hepatitis D virus Hepatitis E virus GB viruses

GB virus C

62 Human Retroviruses George J. Dawson and lsa K. Mushahwar Etiologic agents HTLV-I/-I I HIV-11-2

63 Dengue Virus Infections Bruce J. Dille and Isa K. Mushahwar

64 Miscellaneous Diseases of Microbiological Origin Bruce J. Dille and Isa K. Mushahwar Herpes simplex virus type 1 Epstein-Barr virus Chagas' disease Candida albicans - invasive candidiasis Borrelfa burgdorferi Helicobacter pylori

The lmmunoassay Handbook

Group A streptococcus Human herpesvirus 6 Human herpesvirus 8 (Kaposi's sarcoma herpesvirus)

65 Therapeutic Drug Monitoring (TDM) Philip A. Routledge and Alun D. Hutchings lnttoduction

Assay technology Measurement of free drug concentration Practical aspects of TDM

Antiarrhythmic drugs Acecainide (N-acetylprocainamide) Disopyramide Lidocaine Procainarnide Propranolol Quinidine Arniodarone Flecainide

Antibiotics Arnikacin Kanamycin Gentamicin Tobramycin Netilmicin Chloramphenicol Vancomycin

Anticonvulsants -- -Phenm------

Phenobarbital (phenobarbitone) Primidone Carbamazepine Valproic acid Ethosuxirnide

Chemotherapeutic agents Methotrexate

Miscellaneous Acetaminophen (paracetamol) Tricyclic antidepressants Cy closporin Digoxin Digitoxin TheophyUine

66 Drugs of Abuse Brian Widdop Applications

Drug dependence treatment centers Psychiatric clinics Medical-legal applications Drug abuse in the workplace

lmmunoassays for drugs of abuse Amphetamine Methamphetamine Methylenedioxyamphetamine Methylenedioxy-methamphetamine Barbiturates Benzodiazepines Cannabis

xviii

Contents

Cocaine Fentanyl Lysergic add diethylamide Methadone Opiates Phencyclidine Propoxyphene

Legal addictive substances Alcohol (ethyl alcohol) and the use of carbohydrate-deficient transferrin Nicotine and the use of cotinine

Steroid abuse in sport Anabolic androgenic steroids

Rapid immunoassay tests for drugs of abuse

67 Assays for Drug-screening Applications and Research Jefiey K. Horton, Stephen J. Capper, Molly J. Price Jones and Kelvin T. Hughes Assay technology

Tritium radioimmunoassays Iodine-125 radioimmunoassays Scintillation proximity assay The application of SPA technology to the measurement of protein: protein interactions using an antibody capture format Imaging technologies and instrumentation for ultra high-throughput drug-screening Enzyme immunoassays

Analytes Prostaglandins Leukotrienes Rat hormone assays Cytokines Signal transduction assays Cell proliferation immunoassay Matrii metalloproteinases (MMPs) Cardiovascular peptides Neurodegeneration assays

68 lmmunoassay Appllcatlons In Veterinary Diagnostics Erwin Workman Infectious disease diagnosis/assessment of immune status

Feline Canine Porcine Equine Bovine Avian Murine

Assessment of reproductlve/metabolic status FelineICanine Equine Bovine

Conclusion

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