Ab Structure and Function

What are antibodies (Ab), and what type of molecule are they?

Antibodies are immunoglobulins (Ig) — glycoproteins found in the serum; most are circulating, but some can be bound to cells.

What is the composition of antibodies by percentage?

82–96% polypeptide and 2–14% carbohydrate.

What are the five major classes of immunoglobulins?

IgG, IgM, IgA, IgD, and IgE.

What is the role of antibodies in immunity, and what produces them?

They are the key element of the humoral immune response; they result from stimulated B cells differentiating into plasma cells.

Where do antibodies primarily appear on serum electrophoresis, and what are they commonly called?

They appear primarily in the gamma (γ) band at pH 8.6 and are called "gammaglobulins."

Tetrapeptide Structure

What is the basic structural unit of all immunoglobulins?

A basic four-chain polypeptide: two large heavy (H) chains and two small light (L) chains, held together by noncovalent forces and disulfide interchain bonds.

What regions does each immunoglobulin chain have?

A single, unique variable region (at the amino-terminal end) and one or more constant regions (at the carboxy-terminal end).

Fc Fragment

What is the Fc fragment, and does it bind antigen?

The Fc fragment has NO antigen-binding ability; it represents the carboxy-terminal halves of two H chains held together by S–S bonding.

What are the effector functions of the Fc fragment?

Opsonization and complement fixation.

Fab Fragment

What is the Fab fragment composed of, and how is it obtained?

One L chain and one-half of an H chain, held together by disulfide bonding; obtained by papain digestion of an immunoglobulin.

What is the F(ab')2 fragment, how is it obtained, and what does it consist of?

Obtained by pepsin digestion; it consists of two antigen-binding sites together, with the Fc' portion in pieces.

Light Chains (Bence Jones Proteins)

Who discovered light chains, when, and what disease are they associated with?

Discovered by Dr. Henry Bence Jones in 1845; L chains secreted by malignant plasma cells are associated with Bence Jones disease.

What are the two types of light chains, and in which Ig classes are they found?

Kappa (κ) and Lambda (λ); found in all 5 classes of Ig.

What are the structural features of light chains?

Contain 200–220 amino acids, have a constant region and a variable region, and are disulfide bonded to heavy chains (with intrachain bonds too).

Heavy Chains

What determines the class (isotype) of an immunoglobulin?

The unique, stable sequences of the constant region of the heavy chain.

What are the constant region designations of heavy chains, and how many regions are there?

Three or more regions with similar sequences designated CH1, CH2, and CH3.

What heavy chain name corresponds to each immunoglobulin class?

IgG → γ (gamma); IgM → μ (mu); IgA → α (alpha); IgD → δ (delta); IgE → ε (epsilon).

What is an isotype?

A unique amino acid sequence common to all immunoglobulin molecules of a given class in a given species.

What are allotypes?

Minor variations of isotype sequences present in some individuals but not others.

What is an idiotype?

Variations in variable regions that give individual antibody molecules their specificity.

Hinge Region

Where is the hinge region located, and which Ig classes have it?

Between the CH1 and CH2 regions of the H chain; found in IgG, IgD, and IgA only.

What is the composition of the hinge region, and what does it allow?

High content of proline (allowing flexibility) and hydrophobic residues; it allows the two antigen-binding sites to operate independently and assists initiation of the complement cascade.

Carbohydrate Portion

Where is the carbohydrate portion of immunoglobulins located, and what are its functions?

Found between the CH2 domains of the two H chains; it increases solubility, provides protection against degradation, and enhances functional activity of the Fc domains.

Three-Dimensional Structure

How are antibodies folded, and what stabilizes them?

Folded into compact globular subunits stabilized by intrachain disulfide bonds.

What is the "immunoglobulin fold," and what is the hypervariable region?

Made up of the folded domains of H and L chains; the hypervariable region (~30 AA long) captures antigen and is called the complementarity-determining region (CDR1, 2, 3).

What is an epitope?

An antigenic determinant — the specific site on an antigen recognized by the antibody.

IgG — General Properties

What percentage of total serum immunoglobulins does IgG represent, and what is its half-life?

75–80% of total serum immunoglobulins; longest half-life of any immunoglobulin class at 23 days.

How many subclasses does IgG have, and what are their percentages?

Four subclasses: IgG1 (66%), IgG2 (23%), IgG3 (7%), IgG4 (4%).

What is the molecular weight of IgG?

~150,000 Da; can span about 14 nm.

IgG — Subclass Properties

What are the distinguishing features of IgG3?

Has the largest hinge region, the largest number of interchain disulfide bonds, is the most efficient at binding complement, and is induced in response to protein antigens.

What are the distinguishing features of IgG2 and IgG4?

Have shorter hinge segments, are poor mediators of complement activation, and are involved in responses to polysaccharide antigens.

IgG — Functions

What are the major functions of IgG?

Providing immunity for the newborn (can cross the placenta), fixing complement, coating antigen for enhanced phagocytosis (opsonization), neutralizing toxins and viruses, and participating in agglutination and precipitation reactions.

Which cells have Fc receptors specific to IgG, and what does this enhance?

Macrophages, monocytes, and neutrophils; enhances efficiency of phagocytosis.

What property allows IgG to enter extravascular spaces more readily than other Ig types?

A high diffusion coefficient.

IgM — General Properties

Why is IgM known as a macroglobulin, and what are its size and abundance?

It has a molecular weight of ~900,000 Da; it accounts for 5–10% of all serum immunoglobulins.

What is the half-life of IgM?

6 days (1–2 weeks); much shorter than IgG.

In what two forms can IgM exist?

As a monomer (on the surface of B cells — mIgM) or as a pentamer (in secretions and plasma/serum; can span ~30 nm).

What holds the IgM pentamer together, and how many antigen-binding sites does it have?

J chains link adjacent monomers via disulfide bonds; the pentamer has 10 antigen-binding sites and a star-like shape.

IgM — Clinical Significance

Why is IgM known as the "primary response antibody"?

It appears first after antigenic stimulation and in the maturing infant; synthesized only as long as antigen remains present; indicates a primary immune response.

Can IgM cross the placenta, and what is its pool location?

No, IgM cannot cross the placenta; found mainly in the intravascular pool.

Does IgM produce memory cells?

No — IgM produces no memory cells.

What are the functions of IgM?

Complement fixation, agglutination, opsonization, and toxin neutralization.

IgA — General Properties

What percentage of circulating immunoglobulins does IgA account for, and what is its structure in serum?

10–15% of circulating immunoglobulins; serum IgA appears as a monomer with MW ~160,000.

What are the two subclasses of IgA and where is each primarily found?

IgA1 — mainly found in serum; IgA2 — predominantly found in secretions at mucosal surfaces.

What is the function of IgA1?

Acts as an anti-inflammatory agent; downregulates IgG-mediated phagocytosis, chemotaxis, bactericidal activity, and cytokine release.

What is the structure and advantage of IgA2 over IgA1?

IgA2 is a dimer along mucosal surfaces; it is more resistant to some bacterial proteinases that can cleave IgA1; it keeps antigen from penetrating farther into the body.

Secretory IgA

Where is secretory IgA synthesized, and how is it released?

Synthesized in plasma cells found mainly in mucosal-associated lymphoid tissue (MALT); released in dimeric form and captured by secretory component (SC) on epithelial cells.

What are the functions of secretory IgA?

Patrols mucosal surfaces as a first line of defense, neutralizes toxins from microorganisms, prevents bacterial adherence to mucosal surfaces, and passively transfers immunity to newborns during breastfeeding.

How does IgA contribute to phagocytosis?

Neutrophils, monocytes, and macrophages have specific receptors for serum and secretory IgA; binding triggers a respiratory burst and degranulation, making IgA an opsonin.

How does IgA interact with the complement system?

Aggregation of IgA immune complexes may trigger the alternate complement pathway.

IgD — Properties and Function

How scarce is IgD, and what is its molecular weight and heavy chain?

Less than 0.001% of total immunoglobulins; MW ~180,000; has a δ heavy chain (MW ~62,000) with an extended hinge region of 58 amino acids.

What makes IgD susceptible to degradation, and what is its half-life?

More susceptible to proteolysis than other immunoglobulins; half-life of only 1–3 days.

Where is IgD found, when does it appear, and what is its proposed role?

Found on the surface of immunocompetent but unstimulated B lymphocytes; appears second after IgM; may play a role in B-cell activation, maturation, and differentiation.

What does the secreted form of IgD NOT do?

Does not bind complement, does not bind to neutrophils or macrophages, and does not cross the placenta.

IgE — Properties

How abundant is IgE, and what is its molecular weight and heavy chain?

0.0005% of total serum immunoglobulins; MW ~190,000; has an ε H chain of ~550 amino acids distributed over one variable and four constant domains.

Where are IgE-producing plasma cells primarily located?

Primarily in the lungs and skin.

What does IgE NOT do, and what does it attach to?

Does not participate in complement fixation, agglutination, or opsonization; does not cross the placenta; attaches to basophils, eosinophils, and tissue mast cells through high-affinity FcεRI receptors.

IgE — Functions

How does IgE trigger an allergic reaction?

Two adjacent IgE molecules on a mast cell bind a specific antigen, causing a cascade of cellular events resulting in degranulation of mast cells and release of vasoactive amines (such as histamine and heparin).

What type of hypersensitivity results from IgE-mediated mast cell degranulation, and what are the clinical manifestations?

Type I immediate hypersensitivity — hay fever, asthma, vomiting and diarrhea, hives, and life-threatening anaphylactic shock.

What role does IgE play in parasitic infections?

Eosinophils (activated by IgE) play a major part in the destruction of large antigens, such as parasitic worms, that cannot be easily phagocytized.

Antibody Diversity Theories

What was Ehrlich's side-chain theory (early 1900s)?

Certain cells had specific surface receptors for antigen present before contact with antigen occurred; when antigen is introduced, it combines with the proper receptors, which break off and enter the circulation as antibody molecules; the process can be repeated with further antigen contact.

What was the clonal selection hypothesis (Jerne and Burnet, 1950s)?

Lymphocytes are genetically preprogrammed to produce one type of immunoglobulin; a specific antigen finds the particular cells capable of responding, causing them to proliferate; this would require a large number of genes.

What did Dryer and Bennett propose in 1965?

That the constant and variable portions of immunoglobulin chains are coded for by separate genes.

Genes Coding for Immunoglobulins

On which chromosomes are the human immunoglobulin gene clusters located?

H chain genes on chromosome 14, κ chain genes on chromosome 2, and λ chain genes on chromosome 22.

What gene segments code for the H chain variable region, and what do L chains lack?

VH, D, and J segments code for the H chain variable region; L chains lack a D region.

What is the result of the random selection and joining of gene segments?

Each lymphocyte is committed to making antibody of a single specificity.

What mnemonic is used to remember class switching order?

"MDs Gives Everyone Apples" — IgM, IgD, IgG, IgE, IgA.

Monoclonal Antibodies

What are monoclonal antibodies primarily used for?

In vitro diagnostic testing and delivery of therapeutic agents in diseases.

From what are monoclonal antibodies derived?

A single parent antibody-producing cell that has reproduced many times.

What is a hybridoma, and how is it made?

A fusion of an activated B cell with a laboratory-grown myeloma cell that cannot make its own DNA (due to deficiency of HGPRT); made by immunizing a mouse with a certain antigen, harvesting spleen cells, combining spleen cells with myeloma cells in the presence of PEG, then selecting fused cells and screening for the desired ant