Current Protocols in Immunology

Table of Contents

• Foreword
• Preface
• Chapter 1 Care and Handling of Laboratory Animals
  • Introduction
  • Section I Animal Health and Husbandry
    • UNIT 1.1 Animal Health Assurance
    • UNIT 1.2 Managing Immunocompromised Animals
    • UNIT 1.3 Handling and Restraint
    • UNIT 1.4 Anesthesia
    • UNIT 1.5 Animal Identification
  • Section II Use of Mouse, Rat, Hamster, and Rabbit
    • UNIT 1.6 Parenteral Injections
    • UNIT 1.7 Blood Collection
    • UNIT 1.8 Euthanasia
  • Section III Selected Surgical Procedures
    • UNIT 1.9 Removal of Lymphoid Organs
    • UNIT 1.10 Survival Surgery: Removal of the Spleen or Thymus
    • UNIT 1.11 Surgical Removal of Mouse Salivary Glands
    • UNIT 1.12 Techniques in Aseptic Rodent Surgery
• Chapter 2 Induction of Immune Responses
  • Introduction
  • Section I Assays for Antibody Production
    • UNIT 2.1 Enzyme‐Linked Immunosorbent Assays
    • UNIT 2.2 Isotype Determination of Antibodies
    • UNIT 2.3 Double‐Immunodiffusion Assay for Detecting Specific Antibodies
  • Section II Production of Antibodies
    • UNIT 2.4 Production of Polyclonal Antisera
    • UNIT 2.5 Production of Monoclonal Antibodies
    • UNIT 2.6 Monoclonal Antibody Supernatant and Ascites Fluid Production
    • Production of Antibodies
  • Section III Purification and Fragmentation of Antibodies
    • UNIT 2.7 Purification of Immunoglobulin G
    • UNIT 2.8 Fragmentation of Immunoglobulin G
    • UNIT 2.9 Isolation of Murine and Human Immunoglobulin M and Murine Immunoglobulin D
    • UNIT 2.9B Measurement of Human Serum IgD Levels
    • UNIT 2.10A Fragmentation of Immunoglobulin M
    • UNIT 2.10B Purification of Human IgA
  • Section IV Specialized Techniques
    • UNIT 2.11 Preparation of Immune Stimulating Complexes (ISCOMs) as Adjuvants
    • UNIT 2.12 Cloning, Expression, and Modification of Antibody V Regions
    • UNIT 2.13 Production of Bispecific Antibodies
    • UNIT 2.14 Nucleic Acid Immunizations
    • UNIT 2.15 Vaccination of Mice with Baculovirus‐Infected Insect Cells Expressing Antigenic Proteins
• Chapter 3 In Vitro Assays for Mouse Lymphocyte Function
  • Introduction
  • Section I Isolation and Fractionation of Mononuclear Cell Populations
    • UNIT 3.1 Isolation of Mouse Mononuclear Cells
    • UNIT 3.2 T Cell Enrichment By Nonadherence to Nylon
    • UNIT 3.3 T Cell Enrichment by Cytotoxic Elimination of B Cells and Accessory Cells
    • UNIT 3.4 T Cell Depletion by Cytotoxic Elimination
    • UNIT 3.5A Fractionation of T and B Cells Using Magnetic Beads
    • UNIT 3.5B Isolation of Ly‐1+/CD5+ B Cells by Cell Sorting
    • UNIT 3.6 Depletion of Accessory Cells by Adherence to Sephadex G‐10
    • UNIT 3.7 Isolation of Dendritic Cells
  • Section II Assays for B Cell Function
    • UNIT 3.8 Assays for B Lymphocyte Function
    • UNIT 3.9 Early Events in B Lymphocyte Activation
    • UNIT 3.10 Proliferative Assays for B Cell Function
  • Section III Assays for T Cell Function
    • UNIT 3.11 Induction and Measurement of Cytotoxic T Lymphocyte Activity
    • UNIT 3.12 Proliferative Assays for T Cell Function
    • UNIT 3.13 Production of TH1 and TH2 Cell Lines and Clones
    • UNIT 3.14 Production of Mouse T Cell Hybridomas
    • UNIT 3.15 Quantitation of Functional T Cells by Limiting Dilution
    • UNIT 3.16 Granule Enzyme Exocytosis Assay for Cytotoxic T Lymphocyte Activation
  • Section IV Related Isolation Procedures and Functional Assays
    • UNIT 3.17 Measurement of Apoptosis and Other Forms of Cell Death
    • UNIT 3.18 In Vitro Systems for the Study of T Cell Development: Fetal Thymus Organ Culture and OP9‐DL1 Cell Coculture
    • UNIT 3.19 Isolation of Mouse Small Intestinal Intraepithelial Lymphocytes, Peyer's Patch, and Lamina Propria Cells
    • UNIT 3.20 Isolation of Mouse Neutrophils
    • UNIT 3.21 Isolation of Mouse Intrahepatic Lymphocytes
    • UNIT 3.22 Isolation of Murine Natural Killer Cells
    • UNIT 3.23 Generation, Isolation, and Maintenance of Rodent Mast Cells and Mast Cell Lines
• Chapter 4 In Vivo Assays for Lymphocyte Function
  • Introduction
  • UNIT 4.1 In Vivo Depletion of CD4‐ and CD8‐Specific T Cells
  • UNIT 4.2 Contact Hypersensitivity
  • UNIT 4.3 Animal Models of Acute and Chronic Graft‐Versus‐Host Disease
  • UNIT 4.4 Skin Allograft Rejection
  • UNIT 4.5 Delayed‐Type Hypersensitivity
  • UNIT 4.6 Assessment of Lymphocyte Development in Radiation Bone Marrow Chimeras
  • UNIT 4.7 Measurement of T and B Cell Turnover with Bromodeoxyuridine
  • UNIT 4.8 Construction of Human‐SCID Chimeric Mice
  • UNIT 4.9 Use of the Intracellular Fluorescent Dye CFSE to Monitor Lymphocyte Migration and Proliferation
• Chapter 5 Immunofluorescence and Cell Sorting
  • Introduction
  • UNIT 5.1 Overview of Flow Cytometry
  • UNIT 5.2 Analysis of Flow Cytometry Data
  • UNIT 5.3 Preparation of Cells and Reagents for Flow Cytometry
  • UNIT 5.4 Flow Cytometry Analysis Using the Becton Dickinson FACS Calibur
  • UNIT 5.5 Measurement of Intracellular Ions by Flow Cytometry
  • UNIT 5.6 Measurement of Intercellular Conjugates by Flow Cytometry
  • UNIT 5.7 Analysis of Cellular DNA Content by Flow Cytometry
  • UNIT 5.8 Multiparameter FACS Analysis
  • UNIT 5.9 Visualization of Multiprotein Complexes by Flow Cytometry
• Chapter 6 Cytokines and Their Cellular Receptors
  • Introduction
  • UNIT 6.1 Measurement of Lymphokine Receptors
  • UNIT 6.2 Measurement of Soluble and Membrane‐Bound Interleukin 1 Using a Fibroblast Bioassay
  • UNIT 6.3 Measurement of Human and Murine Interleukin 2 and Interleukin 4
  • UNIT 6.4 Measurement of Interleukin 3 and Other Hematopoietic Cytokines, such as GM‐CSF, G‐CSF, M‐CSF, Erythropoietin, Steel Factor, and Flt‐3 Ligand
  • UNIT 6.5 Measurement of Mouse and Human Interleukin 5
  • UNIT 6.6 Measurement of Interleukin 6
  • UNIT 6.7 Measurement of Interleukin 7
  • UNIT 6.8 Measurement of Mouse and Human Interferon γ
  • UNIT 6.9 Measurement of Antiviral Activity Induced by Interferons α, β, and γ
  • UNIT 6.10 Measurement of Tumor Necrosis Factor α and β
  • UNIT 6.11 Measurement of TGF‐β in Biological Fluids
  • UNIT 6.12 Biological Responses to Chemokine Superfamily Members
  • UNIT 6.13 Measurement of Mouse and Human Interleukin 9
  • UNIT 6.14 Measurement of Mouse and Human Interleukin 10
  • UNIT 6.15 Measurement of Human Interleukin 11
  • UNIT 6.16 Measurement of Human and Mouse Interleukin‐12
  • UNIT 6.17 Measurement of Human and Murine Stem Cell Factor (c‐kit Ligand)
  • UNIT 6.18 Measurement of Interleukin‐13
  • UNIT 6.19 ELISPOT Assay to Detect Cytokine‐Secreting Murine and Human Cells
  • UNIT 6.20 Immunoenzymetric Assay of Mouse and Human Cytokines Using NIP‐Labeled Anti‐Cytokine Antibodies
  • UNIT 6.21 Detection of Cytokine Receptors by Flow Cytometry
  • UNIT 6.22 Measurement of Interleukin 15
  • UNIT 6.23 Measurement of Interleukin 16
  • UNIT 6.24 Detection of Intracellular Cytokines by Flow Cytometry
  • UNIT 6.25 Measurement of Interleukin‐17
  • UNIT 6.26 Measurement of Human and Mouse Interleukin 18
  • UNIT 6.27 Detection and Isolation of Cytokine Secreting Cells Using the Cytometric Cytokine Secretion Assay
  • UNIT 6.28 The In Vivo Cytokine Capture Assay for Measurement of Cytokine Production in the Mouse
  • UNIT 6.29 Cytokine, Growth Factor, and Chemokine Ligand Database
  • UNIT 6.30 Measurement of Interleukin‐21
• Chapter 7 Immunologic Studies in Humans
  • Introduction
  • Section I Preparation of Human Mononuclear Cell Populations and Subpopulations
    • UNIT 7.1 Isolation of Whole Mononuclear Cells from Peripheral Blood and Cord Blood
    • UNIT 7.2 Isolation of T Cells Using Rosetting Procedures
    • UNIT 7.3 Purification of T Cell Subpopulations
    • UNIT 7.4 Immunomagnetic Purification of T Cell Subpopulations
    • UNIT 7.4B The Purification and Functional Analysis of Human CD4+CD25high Regulatory T Cells
    • UNIT 7.5 Isolation of Human B Cell Populations
    • UNIT 7.6A Isolation of Human Monocyte Populations
    • UNIT 7.6B Isolation and Purification of Human Intestinal Macrophages
    • UNIT 7.7 Isolation of Human NK Cells and Generation of LAK Activity
    • UNIT 7.8 Isolation of Mononuclear Cells from Tonsillar Tissue
    • UNIT 7.9 Detection of Unseparated Human Lymphocytes by Flow Cytometry
    • Preparation of Human Mononuclear Cell Populations and Subpopulations
  • Section II Measurement of Basic Immunologic Characteristics of Human Mononuclear Cells
    • UNIT 7.10 Measurement of Proliferative Responses of Cultured Lymphocytes
    • UNIT 7.11 Induction of Polyclonal Immunoglobulin Synthesis
    • UNIT 7.12 Measurement of Polyclonal Immunoglobulin Synthesis Using ELISA
    • UNIT 7.13 Measurement of Polyclonal Immunoglobulin Synthesis Using the Reverse Plaque Technique
    • UNIT 7.14 Measurement of Immunoglobulin Synthesis Using the ELISPOT Assay
    • UNIT 7.15 Measurement of In Vitro Specific‐Antibody Synthesis
    • UNIT 7.16 Induction and Measurement of In Vivo Antibody Responses
    • UNIT 7.17 Measurement of Polyclonal and Antigen‐Specific Cytotoxic T Cell Function
    • UNIT 7.18 Measurement of Cytotoxic Activity of NK/LAK Cells
    • UNIT 7.18B Measurement of Cytokine Production Using Whole Blood
    • Measurement of Basic Immunologic Characteristics of Human Mononuclear Cells
  • Section III Long‐Term and Immortalized Human Cell Lines and Clones
    • UNIT 7.19 Generation and Maintenance of Cloned Human T Cell Lines
    • UNIT 7.20 Generation of HTLV‐I‐Transformed T Cell Lines
    • UNIT 7.21 Herpesvirus saimiri Transformation of Human T Lymphocytes
    • UNIT 7.22 Generation of Epstein‐Barr Virus (EBV)–Immortalized B Cell Lines
  • Section IV Preparation and Functional Analysis of Other Human Lymphoid and Nonlymphoid Cells
    • UNIT 7.23 Isolation and Functional Analysis of Neutrophils
    • UNIT 7.24 Isolation of Human Basophils
    • UNIT 7.25 Isolation of Tissue Mast Cells
    • UNIT 7.26 Measuring Degranulation of Mast Cells
    • UNIT 7.27 51Cr Release Assay of Antibody‐Dependent Cell‐Mediated Cytotoxicity (ADCC)
    • UNIT 7.28 Measurement of Cellular Adhesion Under Static Conditions
    • UNIT 7.29 Cell Adhesion Under Hydrodynamic Flow Conditions
    • UNIT 7.30 Isolation of Human Intestinal Mucosal Mononuclear Cells
    • UNIT 7.31 Isolation of Human Eosinophils
    • UNIT 7.32 Isolation and Generation of Human Dendritic Cells
    • UNIT 7.33 Measurement of Prostaglandins and Other Eicosanoids
    • UNIT 7.34 Isolation and Characterization of Human Natural Killer Cell Subsets
    • UNIT 7.35 Assays for B Cell and Germinal Center Development
    • UNIT 7.36 In Vitro Models of Human T Cell Anergy
• Chapter 8 Isolation and Analysis of Proteins
  • Introduction
  • Section I Solubilization of Cellular Proteins
    • UNIT 8.1A Solubilization of Lymphocytes
    • UNIT 8.1B Purification of Organelles from Mammalian Cells
    • UNIT 8.1C Subcellular Fractionation of Tissue Culture Cells
  • Section II Isolation of Proteins Using Antibodies
    • UNIT 8.2 Immunoaffinity Chromatography
    • UNIT 8.3 Immunoprecipitation
  • Section III Electrophoretic Separation of Proteins
    • UNIT 8.4 One‐Dimensional SDS Gel Electrophoresis of Proteins
    • UNIT 8.5 Two‐Dimensional Gel Electrophoresis
    • UNIT 8.6 Diagonal Gel Electrophoresis
    • UNIT 8.7 Detection of Protein‐Protein Interactions by Coprecipitation
    • UNIT 8.8 Electroelution of Proteins from Stained Gels
  • Section IV Detection of Proteins
    • UNIT 8.9 Staining Proteins in Gels
    • UNIT 8.10 Immunoblotting and Immunodetection
    • UNIT 8.11 Iodination of Soluble and Membrane‐Bound Proteins
    • UNIT 8.12 Biosynthetic Labeling of Proteins
    • UNIT 8.13 Metabolic Radiolabeling of Animal Cell Glycoconjugates
    • UNIT 8.14 Inhibition of N‐Linked Glycosylation
    • UNIT 8.15 Endoglycosidase and Glycoamidase Release of N‐Linked Glycans
    • UNIT 8.16 Biotinylation and Analysis of Membrane‐Bound and Soluble Proteins
    • UNIT 8.17 Single‐Cell Phospho‐Protein Analysis by Flow Cytometry
• Chapter 9 Peptides
  • Introduction
  • UNIT 9.1 Introduction to Peptide Synthesis
  • UNIT 9.2 Synthetic Peptides for Production of Antibodies that Recognize Intact Proteins
  • UNIT 9.3 Production of Anti‐Peptide Antisera
  • UNIT 9.4 B Cell Epitope Mapping Using Synthetic Peptides
  • UNIT 9.5 Identification of B Cell and T Cell Epitopes Using Synthetic Peptide Combinatorial Libraries
  • UNIT 9.6 Synthesis and Application of Peptide Dendrimers As Protein Mimetics
  • UNIT 9.8 Phage Display Selection, Analysis, and Prediction of B Cell Epitopes
• Chapter 10 Molecular Biology
  • Introduction
  • Section I Isolation and Purification of DNA
    • UNIT 10.1 Purification and Concentration of DNA from Aqueous Solutions
    • UNIT 10.2 Preparation of Genomic DNA from Mammalian Tissue
    • UNIT 10.3 Preparation of Bacterial Plasmid DNA
  • Section II Resolution and Recovery of DNA Fragments
    • UNIT 10.4 Agarose Gel Electrophoresis
    • UNIT 10.5 Isolation and Purification of Large DNA Restriction Fragments from Agarose Gels
  • Section III Analysis of Genomic DNA by Blotting and Hybridization
    • UNIT 10.6A Southern Blotting
    • UNIT 10.6B Hybridization Analysis of DNA Blots
    • UNIT 10.7 Deprotection of Oligonucleotides and Purification Using Denaturing PAGE
  • Section IV Enzymatic Cleavage and Labeling of DNA
    • UNIT 10.8 Digestion of DNA with Restriction Endonucleases
    • UNIT 10.9 Reagents and Radioisotopes Used to Manipulate Nucleic Acids
    • UNIT 10.10 Enzymatic Labeling of Nucleic Acids
  • Section V Preparation and Analysis of RNA
    • UNIT 10.11 Preparation of RNA from Tissues and Cells
    • UNIT 10.12 Analysis of RNA by Northern and Slot‐Blot Hybridization
  • Section VI Transfection and Expression of Cloned DNA
    • UNIT 10.13 Calcium Phosphate Transfection
    • UNIT 10.14 Transfection Using DEAE‐Dextran
    • UNIT 10.15 Transfection by Electroporation
    • UNIT 10.16 Liposome‐Mediated Transfection
    • UNIT 10.17A Stable Transfer of Genes into Mammalian Cells
    • UNIT 10.17B Transfection of Lymphoid Cells
    • UNIT 10.17C Rapid Production of Retroviruses for Efficient Gene Delivery to Mammalian Cells Using 293T Cell–Based Systems
    • UNIT 10.18 Use of Monoclonal Antibodies for Expression Cloning
    • UNIT 10.19A Construction of Immunoglobulin Fusion Proteins
    • UNIT 10.19B Phage Display of Single‐Chain Antibody Constructs
  • Section VII The Polymerase Chain Reaction
    • UNIT 10.20 Enzymatic Amplification of DNA by PCR: Standard Procedures and Optimization
    • UNIT 10.21 Quantitation of Rare DNAs by PCR
    • UNIT 10.22 Detection of Isotype Switch Rearrangement in Bulk Culture by PCR
    • UNIT 10.23 Detection of Cytokine mRNA Expression by PCR
    • UNIT 10.24 cDNA Amplification Using One‐Sided (Anchored) PCR
    • UNIT 10.25 Double‐Stranded DNA Sequence Analysis
    • UNIT 10.26 Evaluating Human T Cell Receptor Gene Expression by PCR
    • UNIT 10.27 Molecular Analysis of Mouse T Cell Receptor Expression Using PCR
    • UNIT 10.28 Spectratype/Immunoscope Analysis of the Expressed TCR Repertoire
    • UNIT 10.29 Multiprobe Ribonuclease Protection Assay for Simultaneous Measurement of mRNA Expression
    • UNIT 10.30 Analysis of Telomere Length and Telomerase Activity
    • UNIT 10.31 Measurement of Mouse T Cell Receptor Excision Circles
    • UNIT 10.32 Use of Bacterial Artificial Chromosomes in Generating Targeted Mutations in Human and Mouse Cytomegaloviruses
• Chapter 11 Biochemistry of Cell Activation
  • Introduction
  • UNIT 11.1 Phosphoinositide and Inositol Phosphate Analysis in Lymphocyte Activation
  • UNIT 11.2 Preparation and Analysis of Phosphorylated Proteins
  • UNIT 11.3 Antiphosphotyrosine Blotting
  • UNIT 11.4 Immune‐Complex Assays for Tyrosine Protein Kinases
  • UNIT 11.5 Assessment of the Effects of Tyrosine Protein Kinase Inhibitors
  • UNIT 11.6 Production of Antibodies that Recognize Specific Tyrosine‐Phosphorylated Peptides
  • UNIT 11.7 Using Protein Kinase and Protein Phosphatase Inhibitors to Dissect Protein Phosphorylation Pathways
  • UNIT 11.8 Analyzing Mitogen‐Activated Protein Kinase (MAPK) Activities in T Cells
  • UNIT 11.9A Key Molecules Involved in Receptor‐Mediated Lymphocyte Activation
  • UNIT 11.9B Signal Transduction During Natural Killer Cell Activation
  • UNIT 11.9C Apoptosis Signaling Pathways
  • UNIT 11.9D TRAF‐Mediated TNFR‐Family Signaling
  • UNIT 11.10 Isolation and Use of Rafts
• Chapter 12 Detection and Analysis of HIV
  • Introduction
  • UNIT 12.1 General Guidelines for Experimenting with HIV
  • UNIT 12.2 Isolation and Quantitation of HIV in Peripheral Blood
  • UNIT 12.3 Infection of CD4+ Primary T Cells and Cell Lines, Generation of Chronically Infected Cell Lines, and Induction of HIV Expression
  • UNIT 12.4 Culture of HIV in Monocytes and Macrophages
  • UNIT 12.5 Detection Assays for HIV Proteins
  • UNIT 12.6 Detection of HIV DNA and RNA Using PCR
  • UNIT 12.7 Regulation of the HIV Promoter/Enhancer
  • UNIT 12.8 In Situ Hybridization for Detection of HIV RNA
  • UNIT 12.9 In Vitro Evaluation of Experimental Agents for Anti‐HIV Activity
  • UNIT 12.10 Vaccinia‐Based Reporter Gene Cell‐Fusion Assays to Quantitate Functional Interactions of HIV Envelope Glycoprotein with Receptors
  • UNIT 12.11 Evaluating Neutralizing Antibodies Against HIV, SIV, and SHIV in Luciferase Reporter Gene Assays
  • UNIT 12.12 Immunophenotyping of T Cell Subpopulations in HIV Disease
• Chapter 13 Complement
  • Introduction
  • UNIT 13.1 Classical Pathway Evaluation
  • UNIT 13.2 Alternative Pathway Evaluation
  • UNIT 13.3 Isolation and Purification of C3 from Human Plasma
  • UNIT 13.4 Assays for Membrane Complement Receptors
  • UNIT 13.5 Isolation and Functional Assay of the Membrane Complement Inhibitors CD55 (DAF) and CD59 (MIRL)
  • UNIT 13.6 Assays for the Mannan‐Binding Lectin Pathway
  • UNIT 13.7 Assays for Complement Proteins Encoded in the Class III Region of Human MHC
  • UNIT 13.8 Human Complement Components C4A and C4B Genetic Diversities: Complex Genotypes and Phenotypes
• Chapter 14 Innate Immunity
  • Introduction
  • UNIT 14.1 The Isolation and Characterization of Murine Macrophages
  • UNIT 14.2 Activation of Murine Macrophages
  • UNIT 14.3 Characterization of Human Monocytes/Macrophages
  • UNIT 14.5 Oxidative Metabolism of Murine Macrophages
  • UNIT 14.6 Measurement of Bacterial Ingestion and Killing by Macrophages
  • UNIT 14.7 Assay for Macrophage‐Mediated Anti‐Tumor Cytotoxicity
  • UNIT 14.8 Measurement of Fcγ Receptor–Mediated Binding and Phagocytosis
  • UNIT 14.9 Measurement of Interferon‐Mediated Antiviral Activity of Macrophages
  • UNIT 14.10 Human Natural Killer Receptors, Co‐Receptors, and Their Ligands
  • UNIT 14.11 Isolation and Functional Use of Human NK T Cells
  • UNIT 14.12 Toll‐Like Receptors
  • UNIT 14.13 Characterization and Functional Analysis of Mouse Invariant Natural T (iNKT) Cells
  • UNIT 14.14 Measuring Autophagy in Macrophages
  • UNIT 14.15 Monocyte and Neutrophil Isolation and Migration Assays
• Chapter 15 Animal Models for Autoimmune and Inflammatory Disease
  • Introduction
  • UNIT 15.1 Experimental Autoimmune Encephalomyelitis in the Mouse
  • UNIT 15.2 Experimental Autoimmune Encephalomyelitis in the Rat
  • UNIT 15.3 The BB Rat as a Model of Human Insulin‐Dependent Diabetes Mellitus
  • UNIT 15.4 Adjuvant Arthritis in the Rat
  • UNIT 15.5 Collagen‐Induced Arthritis
  • UNIT 15.6 Experimental Autoimmune Uveoretinitis in the Rat and Mouse
  • UNIT 15.7 Experimental Autoimmune Thyroiditis in the Mouse
  • UNIT 15.8 Experimental Autoimmune Myasthenia Gravis in the Mouse
  • UNIT 15.9 The NOD Mouse: A Model for Insulin‐Dependent Diabetes Mellitus
  • UNIT 15.10 Streptococcal Cell Wall Arthritis
  • UNIT 15.11 Animal Models of IgA Nephropathy
  • UNIT 15.12 Induction of Autoimmune Disease by Depletion of Regulatory T Cells
  • UNIT 15.13 Induction of Inflammatory Bowel Disease in Immunodeficient Mice by Depletion of Regulatory T Cells
  • UNIT 15.14 Autoimmune Myocarditis
  • UNIT 15.15 Mercuric Chloride–Induced Autoimmunity
  • UNIT 15.16 The Day‐3 Thymectomy Model for Induction of Multiple Organ‐Specific Autoimmune Diseases
  • UNIT 15.17 Autoimmune Ovarian Disease Induced by Immunization with Zona Pellucida (ZP3) Peptide
  • UNIT 15.18 Animal Models of Airway Sensitization
  • UNIT 15.19 Induction of TNBS Colitis in Mice
  • UNIT 15.20 Animal Models for SLE
  • UNIT 15.21 Creation of “Humanized” Mice to Study Human Immunity
  • UNIT 15.22 The K/BxN Arthritis Model
• Chapter 16 Antigen Processing and Presentation
  • Introduction
  • UNIT 16.1 Choosing and Preparing Antigen‐Presenting Cells
  • UNIT 16.2 Presenting Exogenous Antigen to T Cells
  • UNIT 16.3 Targeted Identification of Infection‐Related HLA Class I–Presented Epitopes by Stable Isotope Tagging of Epitopes (SITE)
• Chapter 17 Engineering Immune Molecules and Receptors
  • Introduction
  • UNIT 17.1 Bacteriophage Library Construction and Selection of Recombinant Antibodies
  • UNIT 17.2 Monitoring Antigen‐Specific T Cells Using MHC‐Ig Dimers
  • UNIT 17.3 MHC‐Peptide Tetramers to Visualize Antigen‐Specific T Cells
  • UNIT 17.4 In Situ Staining Using MHC Class I Tetramers
  • UNIT 17.5 Preparation of Stable Single‐Chain Trimers Engineered with Peptide, β2 Microglobulin, and MHC Heavy Chain
• Chapter 18 Ligand‐Receptor Interactions in the Immune System
  • Introduction
  • UNIT 18.1 Binding of Biotinylated Peptides to MHC Class II Proteins on Cell Surfaces
  • UNIT 18.2 Detection of Low‐Affinity Ligand‐Receptor Interactions at the Cell Surface with Fluorescent Microspheres
  • UNIT 18.3 Measurement of MHC/Peptide Interactions by Gel Filtration
  • UNIT 18.4 Measurement of Peptide‐MHC Interactions in Solution Using the Spin Column Filtration Assay
  • UNIT 18.5 Measurement of the Kinetics of Biomolecular Interactions Using the IAsys Resonant Mirror Biosensor
  • UNIT 18.6 Affinity‐Based Biosensors for Biomolecular Interaction Analysis
  • UNIT 18.7 Biotin Labeling and Quantitation of Cell‐Surface Proteins
  • UNIT 18.8 Measuring Protein‐Protein Interactions by Equilibrium Sedimentation
  • UNIT 18.9 Binding of Small Peptides to Immobilized Antibodies: Kinetic Analysis by Surface Plasmon Resonance
  • UNIT 18.10 Fluorescence Resonance Energy Transfer: Techniques for Measuring Molecular Conformation and Molecular Proximity
  • UNIT 18.11 Peptide Induction of Surface Expression of Class I MHC
  • UNIT 18.12 Competition‐Based Cellular Peptide Binding Assay for HLA Class I
  • UNIT 18.13 Supported Planar Bilayers for Study of the Immunological Synapse
  • UNIT 18.14 Measurement of Peptide Dissociation from MHC Class II Molecules
  • UNIT 18.15 Characterizing Protein‐Protein Interactions by Sedimentation Velocity Analytical Ultracentrifugation
  • UNIT 18.16 Generation of Peptide MHC Class I Monomers and Multimers Through Ligand Exchange
  • UNIT 18.17 The MHC Motif Viewer: A Visualization Tool for MHC Binding Motifs
• Chapter 19 Animal Models for Infectious Diseases
  • Introduction
  • UNIT 19.1 Schistosomiasis
  • UNIT 19.2 Animal Models for the Analysis of Immune Responses to Leishmaniasis
  • UNIT 19.3 Animal Models for Toxoplasma gondii Infection
  • UNIT 19.4 Experimental Asexual Blood Stage Malaria Immunity
  • UNIT 19.5 Animal Models of Mycobacteria Infection
  • UNIT 19.6 Animal Models for Candidiasis
  • UNIT 19.7 A Mouse Model for Cytomegalovirus Infection
  • UNIT 19.8 Helicobacter Animal Models
  • UNIT 19.9 Animal Model for Infection with Listeria monocytogenes
  • UNIT 19.10 Animal Models Using Lymphocytic Choriomeningitis Virus
  • UNIT 19.11 Influenza Virus
  • UNIT 19.12 Animal Model of Nippostrongylus brasiliensis and Heligmosomoides polygyrus
• Chapter 20 Animal Models for Tumor Immunology
  • Introduction
  • UNIT 20.1 B16 as a Mouse Model for Human Melanoma
  • UNIT 20.2 Mouse 4T1 Breast Tumor Model
  • UNIT 20.3 Mouse SaI Sarcoma Tumor Model
  • UNIT 20.4 The P815 Mastocytoma Tumor Model
  • UNIT 20.5 The TRAMP Mouse as a Model for Prostate Cancer
  • UNIT 20.6 Models for Lymphoma
  • UNIT 20.7 Identification of Human Tumor Antigens by Serological Analysis of Recombinant cDNA Expression Libraries (SEREX)
  • UNIT 20.8 Carcinoembryonic Antigen Transgenic Mouse Models for Immunotherapy and Development of Cancer Vaccines
  • UNIT 20.9 ErbB2 Transgenic Mice: A Tool for Investigation of the Immune Prevention and Treatment of Mammary Carcinomas
  • UNIT 20.10 Molecular Cloning and Characterization of MHC Class I– and II–Restricted Tumor Antigens Recognized by T Cells
  • UNIT 20.11 The Immune Response to Tumors
• Chapter 21 Microscopy
  • Introduction
  • UNIT 21.1 Proper Alignment and Adjustment of the Light Microscope
  • UNIT 21.2 Fluorescence Microscopy
  • UNIT 21.3 Immunofluorescence Staining
  • UNIT 21.4 Immunohistochemistry
• Chapter 22 Isolation and Differentiation of Stem and Progenitor Cells
  • Introduction
  • Section A Sources and Methods to Obtain Stem and Progenitor Cells
    • UNIT 22A.1 Large Animal Models for Stem and Progenitor Cell Analysis
    • UNIT 22A.2 Sources of Human and Murine Hematopoietic Stem Cells
  • Section B Isolation of Murine Stem and Progenitor Cells
    • UNIT 22B.1 Isolation of Murine Hematopoietic Stem Cells and Progenitor Cells
    • UNIT 22B.2 Characterization of Mouse Hematopoietic Stem and Progenitor Cells
  • Section F Differentiation of Special Populations of Stem and Progenitor Cells
    • UNIT 22F.1 Isolation of Prolymphocytes from Bone Marrow and Fetal Liver
    • UNIT 22F.2 In Vitro Differentiation and Measurement of B Cell Progenitor Activity in Culture
    • UNIT 22F.3 Measurement of Natural Killer Cell Progenitor Activity in Culture
    • UNIT 22F.4 Differentiation of Peripheral Blood Monocytes into Dendritic Cells
    • UNIT 22F.5 Differentiation and Characterization of Myeloid Cells
    • UNIT 22F.6 Culture, Expansion, and Differentiation of Murine Megakaryocytes
    • UNIT 22F.7 Differentiation of Human Erythroid Cells in Culture
    • UNIT 22F.8 Differentiation and Characterization of T Cells
    • UNIT 22F.9 Differentiation of Multipotent Adult Progenitor Cells into Functional Endothelial and Smooth Muscle Cells
• Appendix 1 Abbreviations and Useful Data
  • APPENDIX 1B Amino Acids
  • APPENDIX 1C Commonly Used Mouse Strains
  • APPENDIX 1D Commonly Used Detergents
  • APPENDIX 1E Murine Models of Immunodeficiency and Autoimmune Disease
  • APPENDIX 1F Mouse Endogenous Superantigens: Ms and Mls‐like Determinants Encoded by Mouse Retroviruses
  • APPENDIX 1G HLA Type of EBV‐Transformed Human B Cell Lines
  • APPENDIX 1H Commonly Used Rat Strains
  • APPENDIX 1I Peptide Binding Motifs for MHC Class I and II Molecules
  • APPENDIX 1K Human MHC Genes and Products
  • APPENDIX 1L Mouse MHC Genes and Products
  • APPENDIX 1M Listing, Location, Binding Motifs, and Expression of Nonclassical Class I and Related Genes and Molecules
  • APPENDIX 1N Overview of Protein Folds in the Immune System
  • APPENDIX 1O Nomenclature of the Human T Cell Receptor Genes
  • APPENDIX 1P Nomenclature of the Human Immunoglobulin Genes
  • APPENDIX 1Q Safe Use of Radioisotopes
  • APPENDIX 1R Receptors Involved in the Regulation of Murine NK Cells
  • APPENDIX 1S Nomenclature and Serology of HLA Class I and Class II Alleles
  • APPENDIX 1T Heat‐Shock Proteins
  • APPENDIX 1U Large‐Scale Cell Culture
  • APPENDIX 1V Biosafety: Guidelines for Working with Pathogenic and Infectious Microorganisms
  • APPENDIX 1W Using Bioinformatics Tools for the Sequence Analysis of Immunoglobulins and T Cell Receptors
  • APPENDIX 1X Paired Receptor Systems of the Innate Immune System
• Appendix 2 Commonly Used Reagents and Equipment
  • APPENDIX 2A Common Media, Buffers, and Stock Solutions
  • APPENDIX 2B Equipment
• Appendix 3 Commonly Used Immunological Techniques
  • APPENDIX 3A Monitoring Cell Growth
  • APPENDIX 3B Trypan Blue Exclusion Test of Cell Viability
  • APPENDIX 3C Wright‐Giemsa and Nonspecific Esterase Staining of Cells
  • APPENDIX 3D Labeling Cells in Microtiter Plates for Determination of [3H]Thymidine Uptake
  • APPENDIX 3E Diagnosis and Treatment of Mycoplasma‐Contaminated Cell Cultures
  • APPENDIX 3F Obtaining Human Peripheral Blood Cells
  • APPENDIX 3G Cryopreservation of Cells
  • APPENDIX 3H Dialysis and Concentration of Protein Solutions
  • APPENDIX 3I Column Chromatography of Antibodies and Fragments
  • APPENDIX 3J Detection and Quantitation of Radiolabeled Proteins and DNA in Gels and Blots
  • APPENDIX 3K Silanizing Glassware
  • APPENDIX 3L Quantitation of DNA and RNA with Absorption and Fluorescence Spectroscopy
  • APPENDIX 3M Preparing Frozen Bacterial Stocks
  • APPENDIX 3N Transformation of E. coli by Electroporation
  • APPENDIX 3O Gram Staining
• Appendix 4 The CD System of Leukocyte Surface Molecules
  • APPENDIX 4A Monoclonal Antibodies to Human Cell Surface Antigens
  • APPENDIX 4B Monoclonal Antibodies to Mouse Cell‐Surface Antigens
  • APPENDIX 4C Monoclonal Antibodies to Rat Leukocyte Surface Antigens, MHC Antigens, and Immunoglobulins
• Appendix 5 Suppliers

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