• Provides step-by-step procedures
• Presents PCR techniques and covers both human and murine models
• Devotes an entire chapter to techniques related to Apoptosis
• Includes list of manufacturers of all chemicals and materials used in the procedures with addresses, phone numbers, and Web sites

Immunologists as well investigators in other disciplines may often use protocols involving the isolation, cultures and characterization of different types of leukocytes. Advanced Methods in Cellular Immunology is a collection of techniques in an easy-to-use format.

Each chapter provides readers with related program information, a step-by-step description of the methodology, alternative techniques, pertinent references, and information about commercial sources for materials and regents. In addition to leukocytes, the authors guide readers through the processes of cell culture as well as inflammation and autoimmunity in a variety of animal models.

Covering topics such as PCR and Apoptosis, this book will serve a guide to commonly used procedures in cellular immunology while utilizing both human and murine models.

Contents

1. Sources
   • Preparation of Cell Suspensions
   • Isolation of Cells
   • Mitomycin C Treatment
   • Preparation of Mycobacteria Suspension
   • Preparation of Listeria Suspension

2. ELISA
   • Incorporation of TritiatedThymidine into DNA
   • MTT Assay
   • Cr Labeling

3. Isolation and Characterization of Neutrophils
   • Percoll Sedimentation and Ficoll-Paque Centrifugation
   • Magnetic Cell Sorting with Anti-CD15 Beads
   • Functional Assessment

4. Isolation and Characterization of Eosinophils
   • Discontinuous Density Percoll Gradients
   • Magnetic Cell Sorting with Anti-CD16 Beads
   • Functional Assessment

5. Enrichment of Human Basophils Macrophages
   • Isolation of Macrophages
   • Activation of Macrophages
   • Production of Oxidative Metabolism Intermediates
   • Phagocytosis
   • Killing of Bacteria
   • Killing of Bacteria - Colorimetric Evaluation
   • Microbicidal Assay

6. B Lymphocyte Cloning
   • Murine B Lymphocyte Cloning Assay
   • Proliferation of Pre-B Cell Colonies in Methycellulose
   • Clonal Assay for Murine Lymphohematopoietic Progenitors
   • Clonal Assay for Fetal Cells
   • Long-Term Bone Marrow Cultures

7. Cell-Cultivation In-Vitro
   • Tissue Culture Laboratory
   • Aseptic Techniques
   • Safety Concerns
   • Substrate
   • Basic Principles
   • Primary Cultures
   • Freezing of Cell Lines

8. Analysis of Cytokine or Cytokine Receptor mRNA
   • Introduction
   • Total RNA Isolation
   • Reverse Transcription-Polymerase Chain Reaction (RT-PCR)

9. Intracellular Cytokine Staining
   • Cell Activation
   • Staining

10. Cell Cytotoxicity
    • Preparation of Mouse Cytotoxic T Cells
    • Whole Blood Assay
    • DNA Fragmentation
    • Apoptosis
    • Quantitation of Apoptosis Using Fluorescent Dyes
    • Quantitation of DNA Fragmentation
    • Target Cell Detachment Assay
    • Cytolysin Hemolytic Activity
    • Isolation of Cytoplasmic Granules
    • Esterase Assay
    • Activation of Antibodies
    • Activation by Phorbol Myristate Acetate and Ionophore

11. Mixed Lymphocyte Reaction
    • Unidirectional MLR

12. Animal Models of Autoimmune Diseases
    • Diabetes Mellitus
    • Nonobese Mice
    • Chemically Induced Diabetes
    • Experimental Autoimmune Encephalomyelitis
    • Rheumatoid Arthritis

13. Buffers and Media
14. Commercial Sources

Index

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