ANTIBODIES

A LABORATORY MANUAL • SECOND EDITION

Edited by

Edward A. Greenfield
Dana-Farber Cancer Institute

PRESS
COLD SPRING HARBOR LABORATORY
Cold Spring Hsrbor, New York • www cshlprGss.org
Contents

Preface xxi

CHAPTER 1

Antibody Production by the Immune System

Stefanie Sarantopoulos

INTRODUCTION 1

Immune Responses Combat Microbial Invasion 1

Innate and Adaptive Immunity Are Linked Together by Cell-Cell Interactions 2

B Cells Differentiate into Antibody-Secreting Cells 3

Memory B Cells Are Produced after an Initial Encounter with Foreign Antigens 4

Adaptive Immunity Relies on Clonal Selection 4

B Cells and Antibodies Can Distinguish Foreign Organisms and Molecules from Self 6

Antibody Cross-Reactivity 6

Antibodies Mediate Effector Functions 6

CHAPTER 2

The Antibody Molecule

Stefanie Sarantopoulos

INTRODUCTION 9

Antibody Structure 9

Antibodies from the IgC Class Have Two Identical Antigen-Binding Sites 10

In Addition to the IgC Molecules, Serum Contains Other Classes of

Antibody Molecules 11

Comparison of the Primary Amino Acid Sequences of Light Chains Reveals

a Constant and a Variable Region 12

Comparison of the Sequences of Heavy Chains Also Reveals Variable and

Constant Regions 12

The Variable Regions of the Heavy and Light Chains Form the Antigen-Binding Site 14

Antibody Diversity 14

DNA Rearrangements Are Required for the Formation of a Functional k Cene 14

DNA Rearrangements Arc1 Also Required to Form a Functional A Cene IS

Forming a Functional Heavy-Chain Cene Requires Two DNA Rearrangements IS

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These Rearrangements and Additional Special Mechanisms Create

a Vast Number ofAntigen-Binding Sites 17

Allelic Exclusion Ensures That Only One Rearranged Light-Chain Gene

and One Rearranged Heavy-Chain Gene Aie Expiessed in Any One B Cell 17

Somatic Mutation and Affinity Maturation The Hypervariable CDRs 18

Additional Recombination Events Are Used to Generate the Different Classes

and Subclasses of Antibodies 19

CHAPTER 3

Antibody-Antigen Interactions 21

Stefanie Sarantopoulos

INTRODUCTION 21

Structure of the Antibody-Antigen Complex 21

Affinity 24

Avidity 24

CHAPTER 4

Antibody Responses 31

Stefanie Sarantopoulos

INTRODUCTION 31

B-Cell Receptor (BCR) and BAFF Receptor Signaling Is Required for B-Cell Survival 31

Primary and
Secondary Antibody Responses to Vaccination Occur with

Stereotypical Kinetics 32

At the End of the Primary Response, the Antigen Is Cleared, Leaving

Primed Memory Cells 32

Subsequent Injections of Antigen Induce a More Potent "Secondary"

Antibody Response 33

The Type of Antigen and Its Method of Delivery Can Determine the Type of
Immune Response 33

Antigen Is Presented to T Cells after Phagocytosis and Antigen Processing by APCs 34

Characteristics of the T-Cell Receptor Genes and Protein Account

for Their Capacity to Recognize Antigens 35

Properties of the Interactions between Class II and Peptide and between

MHC-Peptide Complex and the TCR Explain Why Only Some Compounds
Make Good Antigens 35

High-Affinity Antibody Production Requires Both TCR Binding to an Antigen-

Fragment-Class II Protein Complex on an APC and BCR Recognition
of That Antigen 36

B-Cell Activation Requires Second Signal Delivery That Occurs with T-Cell-B-Cell
Synapse 36

Additional Signals and Cytokines Lead to B-Cell Activation and Differentiation 37

Immune Tolerance Can Result in Difficult-to-Produce Antibodies 37

High-Affinity, Functional Antibody Production Occurs in the Germinal

Center Reaction 39

Binding of TFH Cells to B Cells Leads to Proliferation and Differentiation of B Cells 39