THE ESSENTIAL OF IMMUNOLOGY

lgG3 - Gamma 3 (a3) hea\y chains lgG4 - Gamma 4 (a4) heavy chains 2. IgA Subclasses

a)IgAl - Alpha 1 (al) heavy chains

b)lgA2 - Alpha 2 (a2) heavy chains

E.Nomenclature - Immunoglobulins are named based on the class, or subclass of the heavy chain and type or subtype of light chain. Unless it is stated precisely you arc to assume that all subclass, types and subtypes are present. IgG means that all subclasses and types are present.

F.Heterogeneity - Immunoglobulins considered as a population of molecules are normally very heterogeneous because they are composed of different classes and subclasses each of which has different types and subtypes of light chains. In addition, different immunoglobulin molecules can have different antigen binding properties because of different vh and VL regions.

VII. STRUCTURE AND SOME PROPERTIES OF IG CLASSES AND SUBCLASSES

A IgG

1 Structure - The structures of the IgG subclasses are presented in Figure 7. All IgG's are monomers (7S immunoglobulin). The subclasses differ in the number of disulfide bonds and length of the hinge region.

2. Properties - Most versatile immunoglobulin because it is apable of earning out all of the functions of immunoglobulin molecules.

a)IgG is the major Ig in serum - 75% of serum Ig is IgG

b)IgG is the major Ig in extra vascular spaces

c)Placental transfer - IgG is the only class of Ig that crosses the placenta. Transfer ismediated by receptor on placental cells for the Fc region of IgG. Not all subclasses cross equally; lgG2 does not cross well.

d)Fixes complement - Not all subclasses fix equally well: lgG4 does not fix complement

e)Binding to cells - Macrophages. monocytcs, PMN's and some lymphocytes have Fc receptors for the Fc region of IgG. Not all subclasses bind equally well; lgG2 and lgG4 do not bind to Fc receptors. A consequence of binding to the Fc receptors on PMN's. monocytes and macrophages is that the cell can now internalize the antigen

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better. The antibody has prepared the antigen for eating by the phagocytic cells. The term opsonin is used to describe substances that enhance phagocytosis. IgG is a good opsonin. Binding of IgG to Fc receptors on other types of cells results in

the activation of other functions.

B.IgM

1.Structure - The structure of IgM is presented in Figure 8. IgM normally exists as a pentamer (I9S immunoglobulin) but it can also exist as a monomer. In the pentameric form all heavy chains are identical and all light chains are identical. Thus, the valence is theoretically 10. IgM has an extra domain on the i chain (CH4) and it has another protein covalently bound via a S-S bond called the J chain. This chain functions in polymerization of the molecule into a pentamer.

2.Properties

a)IgM is the 3rd most common serum Ig.

b)IgM is the first Ig to be made by the fetus and thejirsUgto be made by a virgin B cellswhen it is stimulated by antigen.

c)As a consequence of its pentameric structure, IgM is a good complement fixing Ig. Thus. IgM antibodies are very efficient in leading to the lysis of microorganisms.

d)As a consequence of its structure. IgM is also a good gglutinating Ig . Thus. IgM antibodies are very good in clumping icroorganisms for eventual elimination from the body.

e)IgM binds to some cells via Fc receptors.

f)B cell surface Ig - Surface IgM exists as a monomer and lacks J chain but it has an extra 20 amino acids at the C-terminal end to nchor it into the membrane. Cell surface IgM functions as a receptor for antigen on B cells. Surface IgM is noncov alently

associated with two additional proteins in the membrane of the B cell called Ig-a and Ig-a. These additional proteins act as signal transducing molecules since the cytoplasmic tail of the Ig molecule itself is too short to transduce a signal. Contact between surface immunoglobulin and an antigen is required before a signal can be transduced by the Ig-a and Ig-a chains. In the case of T-independent antigens, contact between the antigen and surface immunoglobulin is sufficient to activate B cells to differentiate into antibody secreting plasma cells. However, for T-dependent antigens, a second signal provided by helper T cells is required before B cells are activated.

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a)IgD is found in low levels in serum: its role in serum isuncertain.

b)IgD is primarily found on B cell surfaces where it functions as a receptor

for antigen. IgD on the surface of B cells has extra amino acids at C-terminal end for anchoring to the membrane. It also associates with the Ig-a and Ig-a chains.

c)IgD does not bind complement.

E. IgE

1.Structure - IgE exists as a monomer and has an extra domain in the constant region.

2.Properties

a)IgE is the least common serum Ig since it binds very tightly to Fc receptors on basophils and mast cells even before interacting with antigen.

b)Involved in allergic reactions - As a consequence of its binding to

basophils an mast cells. IgE is involved in allergic reactions Binding of the allergen to the IgE on the cells results in the release of various pharmacological mediators that result inallergic symptoms.

c) IgE also plays a role in parasitic helminth diseases. Since serum IgE lev els rise in parasitic diseases, measuring IgE levels is helpful in diagnosing parasitic infections. Eosinophils have Fc receptors for IgE and binding of eosinophils to IgEcoated helminths results in killing of the parasite.

d)IgE does not fix complement.

Clinical Implications of Human Immunoglobulin Classes IgC

1. Increases in:

a)Chronic granulomatous infections

b)Infections of all types

c)Hyperimmunization

d)Liver disease

e)Malnutrition (severe)

f)Dysproteinemia

g)Disease associated with hypersensitivity granulomas. dermatologic disorders, and IgG myeloma

h)Rlieumatoid arthritis 2.

Decreases in:

a)Agammaglobulinemia

b)Lymphoid aplasia

c)Selective IgG, IgA deficiency

d)IgA myeloma

e)Bence Jones proteinemia

f)Chronic lymphoblastic leukemia

IgM

1.Increases (in adults) in:

a)WaldenstromA macroglobulinemia

b)Trypanosomiasis

c)Actinomycosis

d)Carrion=s disease (bartonellosis)

e)Malaria

f)Infectious mononucleosis

g)Lupus erythematosus

h)Rheumatoid arthritis

I) Dysgammaglobulinemia (certain cases)

Note. In the newborn, a level of IgM above 20 ng./dl is an indication of in utero stimulation of the

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immune system and stimulation by the rubella virus, the cytomegalovirus, syphilis, or toxoplasmosis.

2.Decreases in:

a)Agammaglobulinemia

b)Lymphoproliferative disorders (certain cases)

c)Lymphoid aplasia

d)IgG and IgA myeloma

e)Dysgammaglobulinemia

f)Chronic lymphoblastic leukemia

IgA

I. Increases in:

a)Wiskotl-Aldrich syndrome

b)Cirrhosis of the liver (most cases)

c)Certain stages of collagen and other autoimmune disorders such as rheumatoid arthritis and lupus erythematosus

d)Chronic infections not based on immunologic deficiencies

e)IgA myeloma

2. Decreases in:

a)Hereditary ataxia telangiectasia

b)Immunologic deficiency states (e.g.. dysgammaglobulinemia, congenital and acquired agammaglobulinemia, and hypogammaglobulinemia)

c)Malabsorption syndromes

d)Lymphoid aplasia

e)IgG myeloma

f)Acute lymphoblastic leukemia

g)Chronic lymphoblastic leukemia

IgD

1. Increases in:

a)Chronic infections

b)IgD myelomas

IgE

1. Increases in:

a)Atopic skin diseases such as eczema

b)Hay fever

c)Asthma

d)Anaphylactic shock

e)IgE-myeloma

2.Decreases in:

a)Congenital agammaglobulinemia

b)Hypogammaglobulinemia due to faulty metabolism or synthesis of immunoglobulins

IMMUNOGLOBULINS: ISOTYPES, ALLOTYPES AND IDIOTYPES

TEACHING OBJECTIVES:

1.To explain the structural basis for immunoglobulin isotypes. allotypes and idiotypes

2.To describe some of the uses of isotypes, allotypes and idiotypes

I. ISOTYPES

A. Definition - fsotypcs are antigenic determinants that characterize classes and subclasses of heavy

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chains and types and subtypes of light chains. If human IgM is injected into a rabbit the rabbit will recognize antigenic determinants on the heavy

chain and light chain and make antibodies to them. If that antiserum is absorbed with human IgG the antibodies to the light chain determinants and any determinants in common between human IgM and IgG will be removed and the resulting antiserum will be react only with human IgM. Indeed, the antibodies will only react with the constant region of the chain. Antibodies to the variable region are rare perhaps because only a few copies of each different variable region are represented in the IgM and thus effective immunization does not occur. The determinants that are recognized by such antibodies are called isotvpic determinants and the antibodies to those determinants are called antiisotvpic antibodies. Each class, subclass, type and subtype of immunoglobulin has its unique set of

II.ALLOTYPES

A. Definition - Allotypes are antigenic determinants specified by allelic forms of the Ig genes.

Allotypes represent slight differences in the amino acid sequences in the heavy or light chains of different indiv iduals. Even a single amino acid difference can give rise to an allotypic determinant, although in many cases the several amino acid substitutions have occurred.

Allotypic differences are detected by using antibodies directed against allotypic determinants. These antibodies can be prepared by injecting the Ig from one person into another. In practice however we obtain anti-allotype antisera from women who have had multiple pregnancies or from people who

D.Human Ig Allotypes

Nomenclature - Human Ig allotypes are named on the basis of the heavy or light chain on which it is located. Thus, an allotype on a Gamma 1 heavy chain is given the name: Glm(3). An allotype on a Kappa light chain is given the name: Km(l).

E. Genetics

1. Codominant autosomal genes - Allotv pes that represent amino acid substitutions at the same position in a heavy or light chain (eg. Gl m(3) and Glm(17) or Km(l) and Km(3) are inherited as codominant autosomal genes.

2. Allelic Exclusion - Although in a helerozvgote both alleles are expressed, any individual Ig molecule will only have one allot) pe. This is because an individual B cell can only expresses one allele. This is called allelic exclusion. Allotypes that represent amino acid substitutions at different locations in a molecule (eg. Glm(l) and Glm(17)) can be found on the same molecule.

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and semen and are useful in forensic medicine.

Paternity testing - The immunoglobulin allotypes are one of the characteristics used in legal cases involving paternity.

III IDIOTYPES (Id)

A. Definition - Unique antigenic determinants present on individual antibody molecules or on molecules of identical specificity. Identical specificity means that all antibodies molecules have the exact same hvperv ariable regions. To understand what idiotypes are. it is helpful to understand how they are detected.

B.Location - Idiotypes are localized on the Fab fragment of the Ig molecules.

C.Importance

1.V region marker - Id's are a useful marker for a particular variable region.

IMMUNOGLOBULINS: AG-AB REACTIONS

TEACHING OBJECTIVES:

1.To describe the nature of Ag-Ab reactions

2.To compare and contrast antibody affinity and avidity

3.To delineate the basis for antibody specificity and cross reactivity

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antigenic determinant and a single combining site on the antibody. It is the sum of the

antibody and the valence of the antigen. Avidity is more tha n the sum of the individual affinities.

to one on the immunizing antigen(multispecificity).

IV. TESTS FOR ANTIGEN-ANTIBODY REACTIONS

A. Factors affecting measurement of Ag/Ab reactions - The only way that one knows that an antigen-antibody reaction has occurred is to have some means of directly or indirectly detecting the complexes formed between the antigen and antibody. The ease with which on can detect antigenantibody reactions will depend on a number of factors

1Affinity - The higher the affinity of the antibody for the antigen, the more stable will be the interaction. Thus, the ease with which one can detect the interaction is enhanced.

2Avidity - Reactions between multivalent antigens and multivalent antibodies are more stable and thus easier to detect.

3Ag:Ab ratio - The ratio between the antigen and antibody influences the detection of Ag/Ab complexes because the sizes of the complexes formed is related to the concentration of the antigen and antibody. This is depicted in Figure 1.

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4. Physical form of (he antigen- The physical form of the antigen influences how one detects its reaction with an antibody. If the antigen is a paniculate, one generally looks for agglutination of the antigen by the antibody. If the antigen is soluble one generally looks for the precipitation of the antigen after the production of large insoluble Ag/Ab complexes.

B. Agglutination Tests

1. Aqqlutmation/Hemaqqlutination - When the antigen is paniculate the reaction of an antibody with the antigen can be detected by agglutination (clumping) of the antigen. When the antigen is an erythrocyte the term hemagglutination is used. The term agglutinin is used to describe antibodies that agglutinate paniculate antigens. When the antigen is an erythrocyte the term hemagglutinin is often used. All antibodies can theoretically agglutinate paniculate antigens but IgM due to its high valence is particularly good agglutinin and one sometimes infers that an antibody may be of the IgM class if it is a good agglutinating antibody.

a) Qualitative agglutination test - Agglutination tests can be used in a qualitative manner to assay for the presence of an antigen or an antibody. The antibody is mixed with the paniculate antigen and a positive test is indicated by the agglutination of the paniculate antigen. (Figure 2).

e.g. A patients red blood cells mixed with antibody to a blood group antigen to determine a persons blood type.

e.g. A patients serum mixed with red blood cells of known blood type to assay for the presence of antibodies to that blood type in the patient's serum

b) Quantitative agglutination test - Agglutination tests can also be used to quantitate the level of antibodies to paniculate antigens. In this test one makes serial dilutions of a sample to be tested for antibody and then adds a fixed number of red blood cells or bacteria or other such paniculate antigen and determines the maximum dilution which gives agglutination. The maximum dilution that gives visible agglutination is called the titer. The results are reported as the reciprocal of the maximal dilution that gives visible agglutination.

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