Bacterial Immunoglobulin–Binding Proteins

Bacterial Immunoglobulin–Binding Proteins

Applications in Immunotechnology

1st Edition - October 28, 1990

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  • Editor: Michael D. P. Boyle
  • eBook ISBN: 9781483216539

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Bacterial Immunoglobulin-Binding Proteins: Applications in Immunotechnology, Volume 2 covers the state of knowledge of bacterial immunoglobulin-binding proteins. The book focuses on practical approaches to isolation, characterization, and use of bacterial immunoglobulin-binding proteins. The majority of these studies involve the type I Fc-binding protein (staphylococcal protein A) and the type III Fc-binding protein (streptococcal protein G). Physiological chemists, pediatricians, and microbiologists will find the book invaluable.

Table of Contents

  • Contributors


    1 Introduction to bacterial immunoglobulin-binding proteins

    I. Introduction

    II. The Distribution and Functional Reactivity of Bacterial IgG Fc-Binding Proteins

    III. Bacterial IgG-Binding Proteins

    IV. Second Generation Immunoglobulin-Binding Proteins

    V. Summary


    2 Detection and enhancement of expression of bacterial cell surface immunoglobulin-binding proteins

    I. Introduction

    II. Preparation of Bacteria

    III. Standardization of Bacteria

    IV. Assays for the Detection of Immunoglobulin-Binding Proteins

    V. Direct Binding of Immunoglobulins to Bacteria

    VI. Direct Binding Assay

    VII. Absorption of IgG by Bacteria

    VIII. Dot Blot Procedure

    IX. Methods to Enhance Immunoglobulin-Binding Proteins

    X. Mouse Passage

    XI. Colony Blot Selection

    XII. Storage of Strains

    XIII. Conclusions


    3 Extraction and monitoring of soluble immunoglobulin-binding proteins

    I. Introduction

    II. Extraction Procedures

    III. Secreted IgG-Binding Proteins

    IV. Summary


    4 Isolation and functional characterization of bacterial immunoglobulin-binding proteins

    I. Introduction

    II. Selection of Immunoglobulin Source to Prepare Affinity Columns

    III. Nature of the Sample to Be Purified

    IV. Selection of Eluting Agent

    V. Affinity Purification of a Type III Fc-Binding Protein Solubilized by Bacteriophage Lysis of the Group C Streptococcus 26RP66

    VI. Characterization of Affinity-Purified Immunoglobulin-Binding Proteins

    VII. Comparison of Functional Activities of Fc-Binding Proteins

    VIII. Summary


    5 Determination of protein-binding affinities among bacterial cell surface proteins

    I. Introduction

    II. Binding of Mammalian Proteins to Bacterial Surfaces: Screening Procedures

    III. Solubilization of Immunoglobulin-Binding Bacterial Surface Proteins

    IV. Determination of Immunoglobulin-Binding Protein in Low Concentrations

    V. Analysis of the Binding Immunoglobulins and Other Host Proteins to Purified, Bacterial Surface Proteins

    VI. Determination of the Affinity Constant for Binding between a Bacterial Cell Wall Protein and Its Ligand


    6. Production of polyclonal antibodies to immunoglobulin-binding proteins

    I. Introduction

    II. Selection of the Species of Animal to Immunize

    III. Preparation of Immunogen and Immunization

    IV. Kinetics of Antibody Production

    V. Detection of Antibody to Immunoglobulin-Binding Proteins

    VI. Conclusion


    7. Use of radiolabeled bacterial Fc-binding proteins as tracers for soluble antigens

    I. Introduction

    II. Competitive Inhibition Radioimmunoassay

    III. Summary


    8. Application of enzyme-labeled IgG-binding proteins in immunoassay

    I. Introduction

    II. Types of Assays

    III. Preparation of Immunoglobulin Fc-Binding Protein-Enzyme Conjugate Tracers

    IV. Development of an ELISA to Quantify Human IgA Using a Type III Fc-Binding Protein-Alkaline Phosphatase Conjugate as Tracer

    V. Summary


    9. Use of Fc-binding proteins to identify cell surface and secreted antigens associated with group B streptococci

    I. Introduction

    II. Group B Streptococcal Typing Nomenclature

    III. Two-Stage Radioimmunoassay for Detection of Group B Streptococcal Type-Specific Antigens

    IV. Adaptation of the Two-Stage RIA to a Dot Blot Assay

    V. Adaptation of the Two-Stage RIA to an ELISA Typing Procedure

    VI. Summary


    10. Application of Fc-binding proteins for the detection of specific antibodies

    I. Introduction

    II. Development of an Assay for Antibodies to a Soluble Antigen

    III. Detection of Antibodies to Tumor-Associated Antigens

    IV. Detection of Rabbit IgM Antibodies to Sheep Erythrocytes

    V. Summary


    11. Use of fluorescent-conjugated bacterial immunoglobulin-binding proteins

    I. Standard Methods

    II. Conjugation Method Using GMBS

    III. Comparison of Recombinant Protein G to Wild Type Protein G Isolated from Streptococcus Cell Membranes


    12. Biotinylated IgG binding proteins—doubly versatile

    I. Introduction

    II. Biotinylation of IgG-Binding Protein

    III. Immunohistochemical Staining Using Biotinylated IgG-Binding Proteins


    13. Use of IgG-binding proteins in immunoelectronmicroscopy

    I. Introduction

    II. Preparation of Colloidal Gold

    III. Coupling of Proteins to Colloidal Gold

    IV. Use of Gold-Labeling for Localization of Immunoglobulin-Binding Sites and Antigen-Antibody Complexes in Bacteria

    V. Double Labeling Techniques with Different Sizes of Colloidal Gold to Localize Two Different Antigens on Thin Sections

    VI. Streptavidin/Avidin-Biotin Labeling for Detection of Immunoglobulin-Binding Proteins

    VII. Replica Method with Plasma Polymerization Film by Glow Discharge for Three-Dimensional Demonstration of Colloidal Gold Particles

    VIII. General Applications of Colloidal Gold Labeling


    14. The use of bacterial Fc-binding proteins as probes for antigen-antibody complexes immobilized on nitrocellulose membranes

    I. Dot Blot Assay

    II. Colony and Plaque Blotting of Antigens Expressed by Bacteria

    III. Western Blot Analysis

    IV. Summary


    15. Application of bacteria expressing immunoglobulin-binding proteins to immunoprecipitation reactions

    I. Introduction

    II. General Background

    III. Preparation of Bacterial Immunosorbent Reagents

    IV. Practical Applications Using Bacterial Immunosorbents

    V. Summary


    16. Use of bacteria expressing immunoglobulin-binding proteins in coagglutination assays

    I. Introduction

    II. Detection of Cell-Bound Antigens

    III. Detection of Specific Antibody

    IV. Procedure for Establishing a Coagglutination Assay to Measure a Polyvalent Soluble Antigen

    V. Summary


    17 Utilization of whole bacteria expressing IgG-binding proteins to detect cell surface antigens

    I. Introduction

    II. Reagents and Equipment

    III. Preparation of Antibodies

    IV. Preparation of Anti-Immunoglobulin-Coated Bacteria

    V. Preparation of Hybridoma Antibody-Coated Bacteria

    VI. Binding Assay

    VII. Staining and Quantitation

    VIII. Variations on the Theme

    IX. Comments


    18 Use of immobilized protein A to purify immunoglobulins

    I. Introduction

    II. Overview of Purification Procedure

    III. Choice of Ligand

    IV. Choice of Matrix

    V. Immobilization Procedure

    VI. Available Binding Capacity

    VII. Column Preparation

    VIII.Sample Preparation

    IX. Sample Application

    X. Elution Procedures

    XI. Collection and Detection Methods

    XII. Column Reequilibration, Reuse, and Storage

    XIII. Limitations of Method

    XIV. General Methods Using Protein A Sepharose CL-4B and Protein A Sepharose 4 Fast Flow for Mouse and Human IgG Purification


    19. Purification and quantitation of monoclonal antibodies by affinity chromatography with immobilized protein A

    I. Purification of IgGi Monoclonal Antibodies

    II. Purification of IgM Monoclonal Antibodies

    III. Quantitation of Monoclonal Antibodies

    IV. Purification of Injectable-Grade Monoclonal Antibodies

    V. Conclusions


    20. Use of immobilized protein G to isolate IgG

    I. Introduction

    II. Determination of Optimal Conditions for Protein G Affinity Chromatography

    III. Examples of Affinity Chromatography Using Protein G Agarose

    IV. Use of Protein G Agarose to Make an Antigen-Binding Column

    V. Summary


    21. Bacterial immunoglobulin-binding proteins and complement

    I. Introduction

    II. Measurement of Functional Complement Activity

    III. Detection of Classical Pathway Complement Activity

    IV. Measurement of the Functional Activity of the Alternate Complement Pathway

    V. Application of Functional Complement Titrations to Measurement of Activity of Immunoglobulin-Binding Proteins

    VI. Measurement of the Generation of Complement Split Products

    VII. Measurement of Complement Split Products Generated as a Consequence of Complement Activation Mediated by Bacterial Immunoglobulin-Binding Proteins

    VIII. Studies of Binding of the First Component of Complement

    IX. Antigenic Determination of Complement Activation

    X. Analysis of Complexes Formed between Bacterial Immunoglobulin-Binding Proteins and IgG

    XI. Summary


    22. Activation and differentiation of human lymphocytes by bacterial Fc-binding proteins

    I. Introduction

    II. Lymphocyte Isolation and Purification

    III. Assays for Lymphocyte Proliferation

    IV. Assays of Lymphocyte Differentiation

    V. Conclusion


    23. Measurement of in vivo leucocyte chemotaxis mediated by Fc-binding proteins

    I. Introduction

    II. Air Sac Procedure

    III. Use of Fc-Binding Proteins in the Air Sac Procedure

    IV. Advantages and Limitations of the Air Sac Procedure


    24. The cloning of streptococcal protein G genes

    I. Colony Immunoassay

    II. Streptococcal Clinical Isolates

    III. Preparation of Streptococcal DNA

    IV. Initial Gene Cloning

    V. Cloning of Protein G Genes from Other Isolates


    25. Bacterial immunoglobulin-binding proteins—future trends

    I. Introduction

    II. Role of Bacterial Immunoglobulin-Binding Proteins in Pathogenicity

    III. Structure-Function Relationships of Bacterial Fc-Binding Proteins

    IV. Applications Involving Immunoglobulin-Binding Proteins—Future Trends

    V. Summary



    I. General Buffers

    II. Iodination Buffers and Related Solutions

    III. ELISA Buffers and Related Solutions

    IV. Electrophoresis Buffers

    V. Buffers for Use in Applications Involving Nitrocellulose

    VI. General Buffers and Reagents


Product details

  • No. of pages: 494
  • Language: English
  • Copyright: © Academic Press 1990
  • Published: October 28, 1990
  • Imprint: Academic Press
  • eBook ISBN: 9781483216539

About the Editor

Michael D. P. Boyle

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