Immunity is brought about by a variety of leukocytes (white blood cells), including macrophages and lymphocytes, all of which develop from undifferentiated stem cells in the bone marrow. Leukocytes can leave the bloodstream and patrol the tissues, each cell producing one or more proteins capable of recognizing and binding to molecules that might signal an infection.

The immune response consists of two complementary systems, the humoral and cellular immune systems. The humoral immune system (Latin humor, “fluid”) is directed at bacterial infections and extracellular viruses (those found in the body fluids), but it can also respond to individual foreign proteins. The cellular immune system destroys host cells infected by viruses and also destroys some parasites and foreign tissues.

At the heart of the humoral immune response are soluble proteins called antibodies or immunoglobulins, often abbreviated Ig. Immunoglobulins bind bacteria, viruses, or large molecules identified as foreign and target them for destruction. Making up 20% of blood protein, the immunoglobulins are produced by B lymphocytes, or B cells, so named because they complete their development in the bone marrow.

The agents at the heart of the cellular immune response are a class of T lymphocytes, or T cells (so called because the latter stages of their development occur in the thymus), known as cytotoxic T cells ( cells, also called killer T cells). Recognition of infected cells or parasites involves proteins called T-cell receptors on the surface of cells. Receptors are proteins, usually found on the outer surface of cells and extending through the plasma membrane, that recognize and bind extracellular ligands, thus triggering changes inside the cell.

In addition to cytotoxic T cells, there are helper T cells ( cells), whose function it is to produce soluble signaling proteins called cytokines, which include the interleukins. cells interact with macrophages. The cells participate only indirectly in the destruction of infected cells and pathogens, stimulating the selective proliferation of those and B cells that can bind to a particular antigen. This process, called clonal selection, increases the number of immune system cells that can respond to a particular pathogen. A host organism needs time, often days, to mount an immune response against a new antigen, but memory cells permit a rapid response to pathogens previously encountered. A vaccine to protect against a particular viral infection often consists of weakened or killed virus or isolated proteins from a viral or bacterial protein coat. When injected into a person, the vaccine generally does not cause an infection and illness, but it effectively “teaches” the immune system what the viral particles look like, stimulating the production of memory cells. On subsequent infection, these cells can bind to the virus and trigger a rapid immune response.

The importance of cells is dramatically illustrated by the epidemic produced by HIV (human immunodeficiency virus), the virus that causes AIDS (acquired immune deficiency syndrome). cells are the primary targets of HIV infection; elimination of these cells progressively incapacitates the entire immune system.

Each recognition protein of the immune system, either a T-cell receptor or an antibody produced by a B cell, specifically binds some particular chemical structure, distinguishing it from virtually all others. Humans are capable of producing more than different antibodies with distinct binding specificities. Given this extraordinary diversity, any chemical structure on the surface of a virus or an invading cell will most likely be recognized and bound by one or more antibodies. Antibody diversity is derived from random reassembly of a set of immunoglobulin gene segments through genetic recombination mechanisms that are discussed in Chapter 25 (see Fig. 25-42).

A specialized lexicon is used to describe the unique interactions between antibodies or T-cell receptors and the molecules they bind. Any molecule or pathogen capable of eliciting an immune response is called an antigen. An antigen may be a virus, a bacterial cell wall, or an individual protein or other macromolecule. A complex antigen may be bound by several different antibodies. An individual antibody or T-cell receptor binds only a particular molecular structure within the antigen, called its antigenic determinant or epitope.

It would be unproductive for the immune system to respond to small molecules that are common intermediates and products of cellular metabolism. Molecules of are generally not antigenic. However, when small molecules are covalently attached to large proteins in the laboratory, they can be used to elicit an immune response. These small molecules are called haptens. The antibodies produced in response to protein-linked haptens will then bind to the same small molecules in their free form. Such antibodies are sometimes used in the development of analytical tests described later in this chapter or as a specific ligand in affinity chromatography (see Fig. 3-17c). We now turn to a more detailed description of antibodies and their binding properties.

Immunoglobulin G (IgG) is the major class of antibody molecule and one of the most abundant proteins in the blood serum. IgG has four polypeptide chains: two large ones, called heavy chains, and two smaller ones, called light chains, linked by noncovalent and disulfide bonds into a complex of 150,000. The heavy chains of an IgG molecule interact at one end, then branch to interact separately with the light chains, forming a Y-shaped molecule (Fig. 5-20). At the “hinges” separating the base of an IgG molecule from its branches, the immunoglobulin can be cleaved with proteases. Cleavage with the protease papain liberates the basal fragment, called Fc because it usually crystallizes readily, and the two branches, called Fab, the antigen-binding fragments. Each branch has a single antigen-binding site.

The fundamental structure of immunoglobulins was first established by Gerald Edelman and Rodney Porter in the 1960s. Each chain is made up of identifiable domains. Some are constant in sequence and structure from one IgG to the next; others are variable. The constant domains have a characteristic structure known as the immunoglobulin fold, a well-conserved structural motif in the all-β class of proteins (Chapter 4). There are three of these constant domains in each heavy chain and one in each light chain. The heavy and light chains also have one variable domain each, in which most of the variability in amino acid sequence is found. The variable domains associate to create the antigen-binding site (Fig. 5-20), allowing formation of an antigen-antibody complex (Fig. 5-21).

In many vertebrates, IgG is but one of five classes of immunoglobulins. Each class has a characteristic type of heavy chain, denoted α, δ, ε, γ, and μ for IgA, IgD, IgE, IgG, and IgM, respectively. Two types of light chain, κ and λ, occur in all classes of immunoglobulins. The overall structures of IgD and IgE are similar to that of IgG. IgM occurs either in a monomeric, membrane-bound form or in a secreted form that is a cross-linked pentamer of this basic structure (Fig. 5-22). IgA, found principally in secretions such as saliva, tears, and milk, can be a monomer, a dimer, or a trimer. IgM is the first antibody to be made by B lymphocytes and the major antibody in the early stages of a primary immune response. Some B cells soon begin to produce IgD (with the same antigen-binding site as the IgM produced by the same cell), but the particular function of IgD is less clear.

The IgG described above is the major antibody in secondary immune responses, which are initiated by a class of B cells called memory B cells. As part of the organism’s ongoing immunity to antigens already encountered and dealt with, IgG is the most abundant immunoglobulin in the blood. When IgG binds to an invading bacterium or virus, it activates certain leukocytes such as macrophages to engulf and destroy the invader, and it also activates some other parts of the immune response. Receptors on the macrophage surface recognize and bind the Fc region of IgG. When these Fc receptors bind an antibody-pathogen complex, the macrophage engulfs the complex by phagocytosis (Fig. 5-23).

IgE plays an important role in the allergic response, interacting with basophils (phagocytic leukocytes) in the blood and with histamine-secreting cells called mast cells, which are widely distributed in tissues. This immunoglobulin binds, through its Fc region, to special Fc receptors on the basophils or mast cells. In this form, IgE serves as a receptor for antigen. If antigen is bound, the cells are induced to secrete histamine and other biologically active amines that cause the dilation and increased permeability of blood vessels. These effects on the blood vessels are thought to facilitate the movement of immune system cells and proteins to sites of inflammation. They also produce the symptoms normally associated with allergies. Pollen and other allergens are recognized as foreign, triggering an immune response normally reserved for pathogens.

The binding specificity of an antibody is determined by the amino acid residues in the variable domains of its heavy and light chains. Some of those residues, particularly those lining the antigen-binding site, are hypervariable — especially likely to differ. Specificity is conferred by chemical complementarity between the antigen and its specific binding site. For example, a binding site with a negatively charged group may bind an antigen with a positive charge in the complementary position. In many instances, complementarity is achieved interactively as the structures of antigen and binding site influence each other as they come closer together. Conformational changes in the antibody and/or the antigen then allow the complementary groups to interact fully. This is an example of induced fit. The complex of a peptide derived from HIV (a model antigen) and an Fab molecule, shown in Figure 5-24, illustrates some of these properties. The changes in structure observed on antigen binding are particularly striking in this example.

A typical antibody-antigen interaction is quite strong, characterized by values as low as (recall that a lower corresponds to a stronger binding interaction; see Table 5-1). The reflects the energy derived from the hydrophobic effect and the various ionic, hydrogen-bonding, and van der Waals interactions that stabilize the binding. The binding energy required to produce a of is about 65 kJ/mol.

The extraordinary binding affinity and specificity of antibodies make them valuable analytical reagents. Two types of antibody preparations are in use: polyclonal and monoclonal. Polyclonal antibodies are those produced by many different B lymphocytes responding to one antigen, such as a protein injected into an animal. Cells in the population of B lymphocytes produce antibodies that bind specific, different epitopes within the antigen. Thus, polyclonal preparations contain a mixture of antibodies that recognize different parts of the protein. Monoclonal antibodies, in contrast, are synthesized by a population of identical B cells (a clone) grown in cell culture. These antibodies are homogeneous, all recognizing the same epitope.

The specificity of antibodies has practical uses. In a versatile analytical technique, an antibody is attached to a reagent that makes it easy to detect (Fig. 5-25). When the antibody binds the target protein, the label reveals the presence of the protein in a solution or its location in a gel, or even in a living cell (Fig. 5-25a). In one application of this technique, an immunoblot or Western blot assay (Fig. 5-26b), proteins that have been separated by gel electrophoresis are transferred electrophoretically to a nitrocellulose membrane. After washing, the membrane is treated successively with primary antibody, secondary antibody linked to enzyme, and substrate. A colored precipitate forms only along the band containing the protein of interest. Immunoblotting allows the detection of a minor component in a sample and provides an approximation of its molecular weight.

We will encounter other aspects of antibodies in later chapters. They are extremely important in medicine and can tell us much about the structure of proteins and the action of genes.