IMMUNOLOGY:BASIC CONCEPTS

Most of us are aware that the immune system is designed to protect us from thousands of predatory microorganisms that can invade and seriously damage virtually every part of the body. Our immune system is finely tuned; highly integrated complex that tracks, identifies and destroys a wide range of would be body crashers. This is the side of immune system which most of us are aware of the nurturing and protective side. The immune system provides a powerful defense against potential pathogens, but what is less obvious is it is also capable of bringing in too much power to bear during the course of clearing away foreign invaders. And as almost always happens in such situations, the most devastating damages of all may be done to innocent bystanders. The results could range from a mildly annoying allergy or be more deadly. Eg. People die of hepatitis, not because the virus destroys the liver the virus is actually quite harmless but because of the violence of the attack of the immune system on the infected liver. The same is true for lung damage seen in Tuberculosis. A great deal of degeneration of our bodies as we grow older may be due to subclinical autoimmune diseases. The immune system bedevils us in other ways. The immune system is the major barrier to organ transplantation. Healthy donor organs that could save the life of individuals suffering from end-stage heart or kidney disease are violently rejected by the immune system. Bone marrow transplants that could save the lives of leukaemia victims or children dying of immune deficiency fail due to immunological complications. Yet the immune system fails to protect us from most of the cancers that afflict us. In AIDS, the loss of immune function, that is the hallmark of this disease may be due as much to the immune system attacking itself as to the damage done by the AIDS virus. FUNCTION OF THE IMMUNE SYSTEM Our immune system is designed to protect us from any potential pathogen that may invade our bodies. The main function of immune system is to prevent or limit infections by microorganisms such as bacteria, viruses and parasites. At the heart of adaptive immune response, lie three important features: MEMORY: - We rarely suffer twice from diseases like measles, mumps, chicken pox, etc. The first contact with the infectious agent clearly imprints some information i.e.

imparts some memory, so that the body is effectively prepared to repel any invasion by a similar micro-organism later in life. PRIMARY RESPONSE When antigen is first encountered, antibodies are detectable in the serum after a longer lag period. The lag period is typically 7-10 days but can be longer depending on the nature & dose of the antigen. A small clone of B cells & plasma cells specific for the antigen is formed. The serum antibody concentration continues to rise for several weeks, then declines & may drop to very low level. The first antibodies to appear are IgM, followed by IgG or IgA. Ig M levels decline earlier than IgG level.

SECONDARY RESPONSE During 2nd encounter, there is rapid antibody response (lag period of 3-5 days). The reason is the persistence of antigen specific memory cells after the first contact. These memory cells proliferate to form a large clone of specific B cells & plasma cells which mediate the secondary immune response. During secondary response, IgM is the same. However, Ig G is secreted in large amount & for a longer period. Vaccination utilizes this principle of memory. SPECIFICITY: - The establishment of memory or immunity by one organism does not confer protection against another organism. The body differentiates specifically between the two organisms. The basis of this specificity was explained when haptens were made to induce antibody formation by injecting them after coupling with specific proteins that acted as carriers. This could help us relate the variations in the chemical structure of a hapten to its ability to bind a given antibody. Forces binding the antigens to antibodies include: Elecrostatic: - Attraction between oppositely charged ionic groups on protein side chains. Hydrogen Bonding: - Occurs by bridge formation within hydrophilic groups like OH, -COOH, etc. Hydrophobic: - The force that causes hydrophobic molecules or nonpolar portions of
molecules to aggregate together rather than to dissolve in water is called the hydrophobic bond. This is not a separate bonding force; rather, it is the result of the energy required to insert a nonpolar molecule into water. A nonpolar molecule cannot form hydrogen bonds with water molecules, so it distorts the usual water structure, forcing the water into a rigid cage of hydrogenbonded molecules around it.

Van der Waals forces: - forces between molecules depending on interaction between external electron clouds. Affinity refers to the strength of interaction of antibody with a monovalent hapten or an antigenic determinant. In most practical situations, we are concerned with interaction of antiserum with a multivalent antigen and the term employed to express this binding is AVIDITY. Multivalency of most antigens leads to a bonus effect in which binding of two antigen molecules by an antibody is always greater than the individual antibody links.

Biological Significance of Antibody Affinity and Multivalency

An antibody with high affinity for its antigen can function most effectively in the immune system (e.g. in biological reactions such as haemagglutination, virus neutralization, enzyme inactivation, haemolysis, immune elimination of foreign antigens, etc.). However, an antibody molecule with high affinity for target antigen does not usually exist in the primary naive antibody library, and antibody affinity usually increases during an immune response (called affinity maturation) in vivo. Nature provides us with numerous examples of molecules with low-intrinsic affinity binding sites that are capable of high-avidity interactions with their targets due to multivalent binding. For example, the lowintrinsic affinity of IgM produced during the primary immune response is compensated by its pentameric structure, resulting in a high avidity toward repetitive antigenic determinants present on the surface of bacteria or viruses. Thus, one of the most efficient ways to increase the binding activity of an antibody to a surface (e.g. cell surface) is to make use of the multivalency effect.Therefore, even if the intrinsic affinity of an antibody molecule toward various invaders (e.g. viruses, proteins) is relatively low, high avidity can overcome the low intrinsic affinity, leading to the production of antibody molecules with high intrinsic affinity for the target antigen through affinity maturation in the immune system. RECOGNITION OF SELF AND NON-SELF: - Failure to discriminate between self and non-self can synthesize antibodies directed against components of subjects own body (autoantibodies). Burnet and Fenner postulated that those circulating body components which were able to reach the developing lymphoid system in the perinatal period could in some way be learnt as self. A permanent tolerance or unresponsiveness would then be created so that as immunological maturity is reached, there would normally be an inability to respond to self components.

CELLULAR BASIS OF IMMUNE RESPONSE CELLS OF IMMUNE SYSTEM B cells T cells

 NK cells Monocytes/macrophages Neutrophils Eosinophils Basophils T CELLS These cells are recognized by surface glycoprotein. CD3 CD4 CD8

T CELLS can either be CD4+ Cell or a CD8+ Cell. Helper T cells are the major driving force and the main regulators of the immune defense. Their primary task is to activate B cells and killer T cells. However, the helper T cells themselves must be activated. This happens when a macrophage or dendritic cell, which has eaten an invader, travels to the nearest lymph node to present information about the captured pathogen. The phagocyte displays an antigen fragment from the invader on its own surface, a process called antigen presentation. When the receptor of a helper T cell recognizes the antigen, the T cell is activated. Once activated, helper T cells start to divide and to produce proteins that activate B and T cells as well as other immune cells. The killer T cell The killer T cell is specialized in attacking cells of the body infected by viruses and sometimes also by bacteria. It can also attack cancer cells. The killer T cell has receptors that are used to search each cell that it meets. If a cell is infected, it is swiftly killed. Infected cells are recognized because tiny traces of the intruder, antigen, can be found on their surface. Mechanisms of Killing Perforin/Granzyme Killing Cytotoxic T- Lymphocytes (CTLs) have cytoplasmic granules that contain the proteins perforin and granzymes. When the CTL binds to its target, the contents of the granules are discharged by exocytosis. A dozen or more perforin molecules insert themselves into the plasma membrane of target cells forming a pore that enables granzymes to enter the cell. Granzymes are serine proteases. The two most abundant ones are

Granzyme A: - Once inside the cell, it enters the mitochondria and cleaves a subunit of complex I (the NADH dehydrogenase) of the electron transport chain producing reactive oxygen species (ROS) that kill the cell.

Granzyme B: - Once inside the cell, it proceeds to cleave the precursors of caspases thus activating them to cause the cell to selfdestruct by apoptosis. Fas ligand (FasL) Killing The most potential CTL targets express a receptor for Fas ligand designated Fas. When cytotoxic T cells recognize (bind to) their target, they produce more FasL at their surface. This binds with the Fas on the surface of the target cell leading to its death by apoptosis. B Cells The B lymphocyte cell searches for antigen matching its receptors. If it finds such antigen it connects to it and inside the B cell, a triggering signal is set off. The B cell now needs proteins produced by helper T cells to become fully activated. When this happens, the B cell starts to divide to produce clones of itself. During this process, two new cell types are created, plasma cells and B memory cells. PLASMA CELLS The plasma cell is specialized in producing a specific protein, called an antibody, which will respond to the same antigen that matched the B cell receptor. Antibodies are released from the plasma cell so that they can seek out intruders and help destroy them. Plasma cells produce antibodies at an amazing rate and can release tens of thousands of antibodies per second. MEMORY CELLS The Memory Cells are the second cell type produced by the division of B cells. These cells have a prolonged life span and can thereby "remember" specific intruders. T cells can also produce memory cells with an even longer life span than B memory cells. The second time an intruder tries to invade the body, B and T memory cells help the immune system to activate much faster. The invaders are wiped out before the infected human feels any symptoms. The body has achieved immunity against the invader. MACROPHAGES These arise from circulating monocytes which after a brief period of about 8 hours in blood migrate into various tissues & differentiate into macrophages. The primary

function of macrophages is phagocytosis. They trap the antigen & provide it in an optimal concentration to the lymphocytes. DENDRITIC CELLS Cells of the dendritic cell (DC) lineage are bone marrow derived. In the skin they are known as Langerhans Cells (LC). These cells efficiently process antigen but cannot present it to T cells. LC have been shown to pick up antigen in skin and carry it via afferent lymphatic vessels to lymph nodes. Dendritic cells in lymph are known as "veiled" cells. In lymph nodes the cells, now known as tissue dendritic cells or interdigitating cells, may efficiently present antigen if they encounter the right T cell. GRANULOCYTES There are three types of granulocyte distinguished according to their histological staining patterns. Neutrophils, also known as polymophonuclear leukocytes, express receptors for immunoglobulin and complement and are involved in the acute inflammatory response. Eosinophils carry receptors for IgE, are involved in the destruction of IgE coated parasites, such as helminths, and contribute to the response to allergens. Basophils are the circulating counterpart of tissue mast cells. They express high affinity receptors for IgE and are stimulated to secrete the chemicals responsible for immediate hypersensitivity following antigen induced aggregation of these receptors CYTOKINES These are small secreted proteins which mediate & regulate immunity. They must be produced de novo in response to an immune stimulus. These act over short distances & short time spans & at very low concentration. Lymphokines (Cytokines made by lymphocytes ) Monokines (Cytokines made by Monocytes ) Interleukin (Cytokines made by one leukocytes & acting on other leukocytes) Chemokines (Cytokines with Chemotactic activities) Natural Killer (NK) Cells These are cells that can lyse and kill cancer cells and virus-infected cells by secreting cytotoxin similar to that of cytotoxic T lymphocytes. They are called as NK

cell as they are active without prior exposure to the virus & not enhanced by exposure, and are not specific for any virus. They can kill without antibody but antibody IgG enhances their effectiveness. IL-12 & Gamma interferon are potent activators. Natural killer cells are a small, distinct group of large granular lymphocytes that react nonspecifically and eliminate cancerous and virus-infected cells. They kill their target cells by releasing perforins and other cytolytic chemicals. They secrete potent chemicals that enhance the inflammatory response. NK cells lack T cell receptor, CD3 proteins & surface IgM & IgG . Humans who lack NK cells are predisposed to life threatening infections with Varicella Zoster virus & cytomegalovirus.

THEORETICAL ASPECTS OF IMMUNE RESPONSETHE SELECTIVE THEORY-The initial formation of the selective theory can be traced to Paul Ehrlich in 1900. In an effort to give an explanation for the origin of serum antibody, Ehrlich projected that blood cells expressed a multiplicity of receptors called side-chain receptors, that have the capacity to react with infectious agents and later inactivate them., Ehrlich proposed that binding of the receptor with an infectious agent was similar to the fit between a lock and key by correlating his concept with that used by Emil Fischer in 1894 to explain the interaction between an enzyme and its substrate. Ehrlich recommended that interaction between a cell-bound receptor and an infectious agent would encourage the cell to fabricate and discharge more receptors with the same specificity. According to Ehrlichs theory, the specificity of the receptor was resolute before its introduction to antigen, and later the antigen selected the appropriate receptor.

TheInstructionalTheory

Later in the 1930s and 1940s, challenges confronted the selective theory imposed by various instructional theories, according to which, an antigen had a key role in determining the specificity of the antibody molecule. According to these theories, an antibody would fold around a particular antigen that would serve as a template. The antibody molecule would thereby take for granted a configuration complementary to that of the antigen template. This concept was first hypothesized about 1930 by Friedrich Breinl and Felix Haurowitz and, later in the 1940s, redefined by Linus Pauling in terms of protein folding. But the instructional theories in the 1960s were formally disproved, as information was emerging about the structures of the DNA, RNA, and proteins that would reveal new insightsto solve the problem of how an individual could make antibodies against almost anything. The body has preformed antibodies whose production is further stimulated by the entry of the antigen. The antigen acts instructively as a template around which a standard unfolded gamma globulin chain is molded to provide the appropriate complimentary shape. CLONAL SELECTION THEORY (BRUNET): The four basic principles of the clonal selection hypothesis 1. Each lymphocyte bears a single type of receptor of a unique specificity. 2. Interaction between a foreign molecule and a lymphocyte receptor capable of binding that molecule with high affinity leads to lymphocyte activation. 3. The differentiated effector cells derived from an activated lymphocyte will bear receptors of an identical specificity to those of the parental cell from which that lymphocyte was derived. 4. Lymphocytes bearing receptors specific for self molecules are deleted at an early stage in lymphoid cell development and are therefore absent from the repertoire of mature lymphocytes.

CLONAL EXPANSION THEORY- The discovery of B cells led to a modern theory of antibody
production called Clonal Expansion Theory (sometimes still referred to as Clonal Selection Theory). In Clonal Expansion Theory, B and T cells are created with random antibodies, then screened for selfreactivity. When antigen enters the system, it eventually binds to any B cell displaying an antibody specific to that antigen. This binding event triggers the following three steps:

1. Each activated B cell reproduces, to create an expanding population of identical B cell clones. This population is called a clone. 2. Some members of the clone become plasma cells. Plasma cells produce and secrete copies of the antibody displayed on the B cell surface. 3. Other members of the clone are stored as memory cells. Thus, there is a large population of cells to create a strong response when the antigen enters the system again. Clonal Expansion Theory explains: 1. Specificity, since only antigen-reactive clones are triggered and only antigen-specific antibody is produced. Diversity is not explained but incorporated into this theory. 2. Memory, since clonal expansion explains why subsequent responses to an antigen are exponentially stronger than the initial response. 3. Tolerance, since B and T cells with potential self-reactivity are destroyed or rendered anergic (unable to respond).

TYPES OF IMMUNITY FIRST LINE OF DEFENSE: - SKIN & MUCOUS MEMBRANE SECOND LINE OF DEFENCE: -INNATE ARM OF IMMUNE SYSTEM THIRD LINE OF DEFENSE: -ACQUIRED (ADAPTIVE ) ARM OF IMMUNE SYSTEM

INNATE ARM OF IMMUNE SYSTEM Innate immunity is resistance that exists prior to exposure to the microbe (antigen). Its involved in the killing of invading microorganisms. Its function is that it helps in activating the acquired immune response. The cells of this arm include neutrophils, complement cells, macrophages/monocytes,mast cells,platelets and natural killer cells. If microorganisms breach first line of defense, then innate arm is available to destroy the invaders. It is non-specific, acts within minutes (as the components are preformed & fully active). Innate immunity does not improve after exposure and has no memory. Innate immunity comprises:

physicochemical barriers molecules normally present in body fluids e.g. lysozyme, complement, antiproteases phagocytic & cytotoxic cells such as neutrophils, macrophages, natural killer cells

The Respiratory Burst aka the oxidative burst membrane-bound NADPH system produces superoxide radicals hyperchlorous acid H2O2 chloramines A reliable means of protecting the host in the first instance against many extracellular organisms It is a property of every living organism Unable to deal with all intracellular organisms (e.g protozoa, viruses & certain bacteria are not killed)

ACQUIRED IMMUNITY

It occurs after exposure to an agent and is highly specific. It requires several days before becoming active and has a long term memory for a specific antigen. It removes the antigen upon repeated exposures. Acquired immunity can be activated only after the innate arm has recognized the microbe Specific & has immunologic memory Dedicated immune cells - the lymphoid cells (lymphocytes) Molecules that immunoglobulins) specifically counteract antigens (antibodies or

 Specific immune systems associated with barrier surfaces e.g. MALT, GALT Lymphocyte secreted cytokines The development of acquired immunity begins with a primary immune response: an afferent phase involving APCs T-cell transformation from a resting to an active state an effector phase - induction of other cells (B-cells & macrophages) by active T-cells The primary immune response is accompanied by the appearance of antigen-specific T-cells & Ig-secreting B-cells The secondary immune response: on second (& subsequent) exposure to the same antigen, antigen-specific memory T- & B-cells are recruited much sooner & more efficiently Ig levels are consequently much higher Extracellular foreign antigen is normally cleared by the innate immune system, with some assistance from B-cell activity Intracellular foreign antigen is handled by the acquired immune system in which self & foreign antigen are jointly recognised All cellular defence mechanisms involve interactions of cell surface molecules (receptors) with complementary molecules (ligands)

Active immunity occurs when when one makes his/her own antibodies. This type of immunity is long term. Getting the disease: if one gets an infectious disease (like chicken pox), often times, that stimulates the production of memory cells which are then stored to prevent the infection in the future. Active immunity can be natural or artificial. Natural active immunity It results from either a clinical or an inapparent infection by a microbe. Artificial active immunity

It is the resistance induced by vaccines. Vaccines are preparations of live or killed microorganisms or their product used for immunization. LIVE VACCINE: - It initiates an infection without causing any injury / disease. The immunity following live vaccine administration therefore parallels that following natural infection though it may be of lower order. The immunity lasts for several years but booster doses may be necessary. Live Vaccines may be administered orally (as with the Sabin Vaccine for Polymyelitis) or parenterally ( as with the measles vaccine ) KILLED VACCINE: - Killed vaccines are generally less immunogenic & protection lasts for a short time period only. They have therefore to be administered repeatedly, generally 2 doses being required for the production of immunity. The first dose is known as Primary Dose& the subsequent doses as booster doses .Killed vaccines may be given orally but generally are not effective. Parentral administration provides humoral antibody response. Passive Immunity occurs when the antibodies come from some other source. This type of immunity is short term. ARTIFICIAL PASSIVE IMMUNITY: - It is the resistance passively transferred to a recipient by the administration of antibodies. The agents used for this purpose are hyper-immune sera, convalescent sera & pooled human gamma globulin. This immunization is indicated for immediate & temporary protection in a non-immune host faced with the threat of an infection, when there is an insufficient time for active immunization to take effect
Cells involved: Lymphocytes which make B lymphocytes (B cells) and T lymphocytes (T cells). Antigen presenting cells (APCs) which include macrophages, B cells and Dendritic cells. Lymphocytes: Circulate through blood and lymphatic system. They produce and display receptors for antigen binding. They are further classes into B-cells, T-cells, or T-lymphocytes. B-Cells, B-lymphocytes come from the bone marrow and mature there. B-cells have receptors that are membrane bound antibody molecules. They are inactive (nave) before exposure to an antigen. Once activated they proliferate into memory cells and antibody secreting effector cells or plasma cells. T-Cells, T-lymphocytes migrate to a lymphoid organ such as the thymus where they mature. The mature T cell express a novel antigen binding receptor called the T cell receptor (TCR). TCRs only recognize antigens that are associated with cell membrane proteins known as MHC (Major Histocompatibility Complex) molecules. Cytotoxic T-cells defend against infections by viruses and bacteria, diseases, tumors cells and transplanted tissues. Antigen Presenting Cells (APC) is a cell that holds a foreign antigen complexed with MHC on it surface. T-cells may recognize the complex with the TCR. Many cells can present antigens to T cells via MHC I molecules but the term is usually limited to cells that prime T cells. Dendritic cell is APC and can be found in the skin, mucosa and lymphoid tissues. They are involved in initiation of immune responses by activating lymphocytes and secreting cytokines. They have long membrane processes.

An effective immune response involves lymphocytes and antigen presenting cells.

Humoral Responses Two classes of adaptive immune responses: Humoral (antibody) and Cell Mediated immune responses. Humoral immune responses are carried out by B-lymphocytes. Primary focus on exogenous antigens. B-cells are activated to secrete antibodies. Cell Mediated Immune Response (CMI) Cell mediated immune responses are carried out by T-lymphocytes. Primary function endogenous antigens. Activated T cells react directly with a presented antigen. CMI responses are carried out by TH cells and TC cells. A function of TC cell would be to kill a host cell that is infected by a virus and is displaying viral antigens. Th cells produce signal molecules that activate macrophages to destroy the microbes that they have phagocytoses. .

Clonal Selection T-Cells Clonal selection is similar to that of B cells. T-cell population results in the clone of effector T-cells and memory T-cells. Effector T-cells include T helper cells and cytotoxic T lymphocytes.

Processing and Presentation of Antigen An antigen must be degraded into small units (peptides) and complexed with MHC I or II molecules in order for a T-cell to recognize it. Antigen processing and presentation is the conversion of antigens into MHC associated fragments. The route that an antigen enters a cell determines if it will be processes and presented with class I or class II MHC molecules (extracellular or intracellular entry). Exogenous antigens are degraded by APCs (macrophages, B-cells, dendritic cells) and complexed with class II MHC and displayed on the cell surfaced. Endogenous antigens like tumor or viral proteins which alters self cells are degraded in the cytoplasm and displayed with class I MHC molecules on the cell surface.

ANTIBODIES TYPES OF ANTIBODIES Monoclonal antibodies are monospecific antibodies that are identical because they are produced by one type of immune cell that are all clones of a single parent cell. Given (almost) any substance, it is possible to create

monoclonal antibodies that specifically bind to the substance; they can then serve to detect or purify that substance. Polyclonal antibodies (or antisera) are antibodies that are derived from different B-cell lines. They are a mixture of immunoglobulin molecules secreted against a specific antigen, each recognizing a different epitope.

Antibodies exist free in body fluids, e.g. serum, or may be membrane bound to B lymphocytes. Their function when membrane bound is to capture antigen for which they have specificity, after which the B lymphocytes will take the antigen into its cytoplasm for further processing. Free antibodies have the following functions: Agglutination of particulate matter, including bacteria and viruses. IgM is particularly suitable for this, as it is able to change its shape from a star form to a form resembling a crab.

Opsonization i.e. coating of bacteria for which the antibody's Fab region has specificity (especially IgG). This facilitates subsequent phagocytosis by cells possessing an Fc receptor, e.g. neutrophil polymorphonuclear leucocytes ("polymorphs").Thus it can be seen that in opsonization and phagocytosis, both the Fab and the Fc portions of the immunoglobulin molecule are involved. Neutralization of toxins released by bacteria. E.g. tetanus toxin is neutralized when specific IgG antibody binds, thus preventing the toxin binding to motor end plates and causing persistent stimulation, manifest as sustained muscular contraction which is the hallmark of tetanic spasms. This applies particularly to IgG. In the case of viruses, antibodies can hinder their ability to attach to receptors on host cells. Here, only Fab is involved. Immobilization of bacteria: - Antibodies against bacterial ciliae or flagellae will hinder their movement and ability to escape the attention of phagocytic cells. Again, only Fab is involved. Complement activation (classical pathway) especially by the Fc region of IgM and IgG, leads eventually to death of bacteria by the terminal complement components which punch holes in the cell wall, leading to an osmotic death.. Mucosal protection. This is provided mainly by IgA, and to a lesser degree, IgG. IgA acts chiefly by inhibiting pathogens from gaining attachment to mucosal surfaces. This is a Fab function.

 Antibody dependent cell mediated cytotoxicity (ADCC). Antibodies bind to organisms via their Fab region. Large granular lymphocytes (Natural Killer cells - abbreviated NK cells), attach via Fc receptors, and kill these organisms not by phagocytosis but by release of toxic substances called perforins. Conferring immunity to the foetus by the transplantal passage of IgG. IgG is the only class (isotope) of immunoglobulin that can cross the placenta and enter the foetal circulation, where it confers immune protection. This is of great importance to the foetus in the first 3 months. STRUCTURE OF ANTIBODY Digestion with proteases is useful for characterizing antibody structure. Papain: derived from papaya latex: - Papain yields F(ab) and Fc fragments, F(ab) = fragment, antigen binding and Fc = fragment, crystallizable Pepsin: derived from pig intestine: -Pepsin yields F(ab)2 + other small fragments -mercaptoethanol reduces disulfides and yields two polypeptide chains (H and L) Although different immunoglobulins can differ structurally, they all are built from the same basic units. A. Heavy and Light Chains: - All immunoglobulins have a four chain structure as their basic unit. They are composed of two identical light chains (23kD) and two identical heavy chains (50-70kD) B. Disulfide bonds Inter-chain disulfide bonds - The heavy and light chains and the two heavy chains are held together by inter-chain disulfide bonds and by non-covalent interactions. The number of inter-chain disulfide bonds varies among different immunoglobulin molecules. Intra-chain disulfide binds - Within each of the polypeptide chains there are also intra-chain disulfide bonds. C. Variable (V) and Constant (C) Regions When the amino acid sequences of many different heavy chains and light chains were compared, it became clear that both the heavy and light chain could be divided into two regions based on variability in the amino acid sequences. These are the:

1. Light Chain - VL (110 amino acids) and CL (110 amino acids) 2. Heavy Chain - VH (110 amino acids) and CH (330-440 amino acids) D. Hinge Region This is the region at which the arms of the antibody molecule form a Y. It is called the hinge region because there is some flexibility in the molecule at this point. E. Domains Three dimensional images of the immunoglobulin molecule show that it is not straight. Rather, it is folded into globular regions each of which contains an intrachain disulfide bond (figure 2B-D). These regions are called domains. 1. Light Chain Domains - VL and CL 2. Heavy Chain Domains - VH, CH1 - CH3 (or CH4) F. Oligosaccharides Carbohydrates are attached to the CH2 domain in most immunoglobulins. However, in some cases carbohydrates may also be attached at other locations. STRUCTURE AND SOME PROPERTIES OF Ig CLASSES AND SUBCLASSES A. IgG 1. Structure All IgG's are monomers (7S immunoglobulin). The subclasses differ in the number of disulfide bonds and length of the hinge region. 2. Properties IgG is the most versatile immunoglobulin because it is capable of carrying 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 is mediated by a receptor on placental cells for the Fc region of IgG. Not all subclasses cross equally well; IgG2 does not cross well.

d) Fixes complement - Not all subclasses fix equally well; IgG4 does not fix complement e) Binding to cells - Macrophages, monocytes, PMNs and some lymphocytes have Fc receptors for the Fc region of IgG. Not all subclasses bind equally well; IgG2 and IgG4 do not bind to Fc receptors. A consequence of binding to the Fc receptors on PMNs, monocytes and macrophages is that the cell can now internalize the antigen 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 Structure IgM normally exists as a pentamer (19S 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 mu 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. Properties a) IgM is the third most common serum Ig. b) IgM is the first Ig to be made by the fetus and the first Ig to be made by a virgin B cells when 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 agglutinating Ig . Thus, IgM antibodies are very good in clumping microorganisms for eventual elimination from the body. e) IgM binds to some cells via Fc receptors. f) B cell surface Ig

C. IgA 1. Structure Serum IgA is a monomer but IgA found in secretions is a dimer. When IgA exits as a dimer, a J chain is associated with it. When IgA is found in secretions is also has another protein associated with it called the secretory piece or T piece; sIgA is sometimes referred to as 11S immunoglobulin. Unlike the remainder of the IgA which is made in the plasma cell, the secretory piece is made in epithelial cells and is added to the IgA as it passes into the secretions (Figure 12). The secretory piece helps IgA to be transported across mucosa and also protects it from degradation in the secretions. 2. Properties a) IgA is the 2nd most common serum Ig. b) IgA is the major class of Ig in secretions - tears, saliva, colostrum, mucus. Since it is found in secretions secretory IgA is important in local (mucosal) immunity. c) Normally IgA does not fix complement, unless aggregated. d) IgA can binding to some cells - PMN's and some lymphocytes.

D. IgD 1. Structure IgD exists only as a monomer. 2. Properties a) IgD is found in low levels in serum; its role in serum is uncertain. 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-alpha and Igbeta 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 in allergic symptoms. c) IgE also plays a role in parasitic helminth diseases. Since serum IgE levels rise in parasitic diseases, measuring IgE levels is helpful in diagnosing parasitic infections. Eosinophils have Fc receptors for IgE and binding of eosinophils to IgE-coated helminths results in killing of the parasite. d) IgE does not fix complement.

ANTIGENS Immunogen: - A substance that induces a specific immune response. Antigen (Ag): - A substance which when introduced parentally into the body stimulates the production of an antibodywith which it reacts specifically to produce an immune response. Tolerogen: - An antigen that induces immunological tolerance (or unresponsiveness to an antigen that is induced by prior exposure to the antigen. Allergen: - Antigen that induces anaphylaxis. Hapten: - A substance that is non-immunogenic but which can react with the products of a specific immune response. Haptens are small molecules which could never induce an immune response when administered by themselves but which can when coupled to a carrier molecule. Free haptens, however, can react with products

of the immune response after such products have been elicited. Haptens have the property of antigenicity but not immunogenicity. Epitope or Antigenic Determinant: - That portion of an antigen that combines with the products of a specific immune response. Classification of Antigens Based on immunogenicity Complete antigen: -substances which can induce both immunogenicity and immunoreactivity. By convention, we call a complete antigen as an antigen. Incomplete antigen: -substances with immunoreactivity only. A measure of the immune reaction caused by an antigen. (Haptens) they need a carrier to express immunogenicity.
Immunogenicity is the ability of a particular substance, such as an antigen or epitope, to provoke an immune response in the body of a human or animal.

Based on chemical nature A. Proteins: - The vast majority of immunogens are proteins. These may be pure proteins or they may be glycoproteins or lipoproteins. In general, proteins are usually very good immunogens. B. Polysaccharides: - Pure polysaccharides and lipopolysaccharides are good immunogens. C. Nucleic Acids: - Nucleic acids are usually poorly immunogenic. However, they may become immunogenic when single stranded or when complexed with proteins. D. Lipids: - In general lipids are non-immunogenic, although they may be haptens. Based on source of the antigen Xenoantigen: - An antigen that is found in more than one species. Alloantigen (ABO system) Autoantigen: - Release of Sequestered antigen eg. The lens protein when released into bloodproduces an immune response which produces inflammation of the lens. Heterophile antigen (Forsmann Antigen): - Common antigens shared by different species. Eg. The M protein of Streptococcus bears common antigenic determinant with basement membrane of the kidney. This can cause Post-streptococcal glomerulonephritis.

SUPERANTIGENS When the immune system encounters a conventional T-dependent antigen, only a small fraction (1 in 104 -105) of the T cell population is able to recognize the antigen and become activated (monoclonal/oligoclonal response). However, there are some antigens which polyclonally activate a large fraction of the T cells (up to 25%). These antigens are called superantigens. Examples of superantigens include: Staphylococcal enterotoxins (food poisoning), Staphylococcal toxic shock toxin (toxic shock syndrome), Staphylococcal exfoliating toxins (scalded skin syndrome) and Streptococcal pyrogenic exotoxins (shock). Although the bacterial superantigens are the best studied there are superantigens associated with viruses and other microorganisms as well. The diseases associated with exposure to superantigens are, in part, due to hyper activation of the immune system and subsequent release of biologically active cytokines by activated T cells. COMPLEMENT SYSTEM The complement system is a series of serum proteins (9 factors) which, through sequential proteolysis, non-specifically increase immunity to infectious organisms and proteins. The initial, inactive complement components are named C1 - C9. Each complement component is synthesized in an inactive form. Once initial activation occurs, the inactive complement component (named C1-C9) is split into fragments, designated by letters (e.g. C1q, C3a, C3b, etc.). synthesised by macrophages or hepatocytes usually circulate as inactive proenzymes heat labile (c.f. Ig is heat stable) Functions opsonisation-C3b,C5b, (MAC)C5b-C9 chemotaxisC3a,C5a, immune adherence

acceleration of acute inflammationC3a,C5a immune cytolysis virus neutralisation(HIV-alternative pathway)

The complement system may be activated in 3 distinct ways, named:

 The Classical Pathway - activated by antigen/antibody complexes The Alternative Pathway- activated by many Gram-negative (mot significantly, N. meningitidis and N. Gonorrhoea), some Gram-positive bacteria and certain viruses and parasites, and results in the lysis of these organisms. The Lectin Pathway - activated by bacterial lectins. The end result of this activation is always the same. Only the initial steps differ. The ultimate end-result of complement activation is the formation of the Membrane Attack Complex, formed by components C5 through C9, which literally punches holes in membranes. In addition, intermediate products such as C3a, C5a, and C3b may play a role in upregulating the immune response through chemotaxis or opsonization. CLASSICAL PATHWAY

C1 activation: -C1, a multi-subunit protein containing three different proteins

(C1q, C1r and C1s), binds to the Fc region of IgG and IgM antibody molecules that have interacted with antigen. C1 binding does not occur to antibodies that have not complexed with antigen and binding requires calcium and magnesium ions. The binding of C1 to antibody is via C1q and C1q must cross link at least two antibody molecules before it is firmly fixed. The binding of C1q results in the activation of C1r which in turn activates C1s. The result is the formation of an activated C1qrs, which is an enzyme that cleaves C4 into two fragments C4a and C4b.

C4 and C2 activation (generation of C3 convertase): - The C4b fragment

binds to the membrane and the C4a fragment is released into the microenvironment. Activated C1qrs also cleaves C2 into C2a and C2b. C2a binds to the membrane in association with C4b, and C2b is released into the microenvironment. The resulting C4bC2a complex is a C3 convertase, which cleaves C3 into C3a and C3b.

C3 activation (generation of C5 convertase): - C3b binds to the membrane in

association with C4b and C2a, and C3a is released into microenvironment. The resulting C4bC2aC3b is a C5 convertase. generation of C5 convertase is the end of the classical pathway. the The

Several of the products of the classical pathway have potent biological activities that contribute to host defenses. Some of these products may also have detrimental effects if produced in an unregulated manner. If the classical pathway were not regulated there would be continued production of C2b, C3a, and C4a. Thus, there occurs regulation of the activity of the classical pathway. LECTIN PATHWAY The lectin pathway is very similar to the classical pathway. It is initiated by the binding of mannose-binding lectin (MBL) to bacterial surfaces with mannosecontaining polysaccharides (mannans). Binding of MBL to a pathogen results in the association of two serine proteases, MASP-1 and MASP-2 (MBL-associated serine proteases). MASP-1 and MASP-2 are similar to C1r and C1s, respectively and MBL is similar to C1q. Formation of the MBL/MASP-1/MASP-2 tri-molecular complex results in the activation of the MASPs and subsequent cleavage of C4 into C4a and C4b. The C4b fragment binds to the membrane and the C4a fragment is released into the microenvironment. Activated MASPs also cleave C2 into C2a and C2b. C2a binds to the membrane in association with C4b and C2b is released into the microenvironment. The resulting C4bC2a complex is a C3 convertase, which cleaves C3 into C3a and C3b. C3b binds to the membrane in association with C4b and C2a and C3a is released into the microenvironment. The resulting C4bC2aC3b is a C5 convertase. The generation of C5 convertase is the end of the lectin pathway. The biological activities and the regulatory proteins of the lectin pathway are the same as those of the classical pathway. ALTERNATIVE PATHWAY The alternative pathway begins with the activation of C3 and requires Factors B and D and Mg++ cation, all present in normal serum. 1. Amplification loop of C3b formation: - In serum there is low level spontaneous hydrolysis of C3 to produce C3i. Factor B binds to C3i and becomes susceptible to Factor D, which cleaves Factor B into Bb. The C3iBb complex acts as a C3 convertase and cleaves C3 into C3a and C3b. Once C3b is formed, Factor B will bind to it and becomes susceptible to cleavage by Factor D. The resulting C3bBb complex is a C3 convertase that will continue to generate more C3b, thus amplifying C3b production. If this process continues unchecked, the result would be the consumption of all C3 in the serum. Thus, the spontaneous production of C3b is tightly controlled. 2. Stabilization of C convertase by activator (protector) surfaces: - When bound to an appropriate activator of the alternative pathway, C3b will bind Factor B, which is

enzymatically cleaved by Factor D to produce C3 convertase (C3bBb). The complex is further stabilized by properdin binding to C3bBb. Activators of the alternate pathway are components on the surface of pathogens and include: LPS of Gramnegative bacteria and the cell walls of some bacteria and yeasts. Thus, when C3b binds to an activator surface, the C3 convertase formed will be stable and continue to generate additional C3a and C3b by cleavage of C3. 3. Generation of C5 convertase: - Some of the C3b generated by the stabilized C3 convertase on the activator surface associates with the C3bBb complex to form a C3bBbC3b complex. This is the C5 convertase of the alternative pathway. The generation of C5 convertase is the end of the alternative pathway. The alternative pathway can be activated by many Gram-negative (most significantly, Neisseria meningitidis and N. gonorrhoea), some Gram-positive bacteria and certain viruses and parasites, and results in the lysis of these organisms. Thus, the alternative pathway of C activation provides another means of protection against certain pathogens before an antibody response is mounted. The alternate pathway may be the more primitive pathway and the classical and lectin pathways probably developed from it. Alternative pathway provides a means of non-specific resistance against infection without the participation of antibodies and hence provides a first line of defense against a number of infectious agents. Many gram negative and some gram positive bacteria, certain viruses, parasites, heterologous red cells, aggregated immunoglobulins (particularly, IgA) and some other proteins (e.g. proteases, clotting pathway products) can activate the alternative pathway. One protein, cobra venom factor (CVF), has been extensively studied for its ability to activate this pathway. MEMBRANE ATTACK (LYTIC) PATHWAY C5 convertase from the classical (C4b2a3b), lectin (C4b2a3b) or alternative (C3bBb3b) pathway cleaves C5 into C5a and C5b. C5a remains in the fluid phase and the C5b rapidly associates with C6 and C7 and inserts into the membrane. Subsequently C8 binds, followed by several molecules of C9. The C9 molecules form a pore in the membrane through which the cellular contents leak and lysis occurs. Lysis is not an enzymatic process; it is thought to be due to physical damage to the membrane. The complex consisting of C5bC6C7C8C9 is referred to as the membrane attack complex (MAC). C5a generated in the lytic pathway has several potent biological activities. It is the most potent anaphylotoxin. In addition, it is a chemotactic factor for neutrophils and stimulates the respiratory burst in them and it stimulates inflammatory cytokine production by macrophages. Its activities are controlled by inactivation by carboxypeptidase B (C3-INA). MAJOR HISTOCOMPATIBILITY COMPLEX

MHC molecules were initially discovered during studies aimed at understanding the molecules responsible for rejection of transplanted tissues. Hence the name Major Histocompatibility Complex(MHC). The term Major Histocompatibility Complex actually refers to a region of the genome that encodes a number of genes (hence Complex) that play an important (hence Major) role in tissue transplantation (hence Histocompatibility). The term MHC molecule or MHC antigen refers to a molecule encoded by a gene within this region. The function of this complex is to transfer information about proteins within a cell to the cell surface. MHC I are expressed on the great majority of cells and recognized by CD8+ T cells. MHC II are expressed on B cells, macrophages, dendritic cells and recognized by CD4+ T cells. They are the cells responsible for graft rejection and are found on chromosome 6 in human and 17 in mouse. T cells only recognize antigen associated with MHC molecules on cell surfaces ANTIGEN-ANTIBODY REACTIONS AGGLUTINATION: - When the antigen is particulate, the reaction of an antibody with the antigen can be detected by agglutination (clumping) of the antigen. The general term agglutinin is used to describe antibodies that agglutinate particulate antigens. When the antigen is an erythrocyte the term hemagglutination is used. All antibodies can theoretically agglutinate particulate 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. Commonly used agglutination test is test to determine ABO blood group. Passive hemagglutination: - The agglutination test only works with particulate antigens. However, it is possible to coat erythrocytes with a soluble antigen (e.g. viral antigen, a polysaccharide or a hapten) and use the coated red blood cells in an agglutination test for antibody to the soluble antigen. This is called passive hemagglutination. The test is performed just like the agglutination test. Applications include detection of antibodies to soluble antigens and detection of antibodies to viral antigens. Coomb's Test (Antiglobulin Test) Direct Coomb's Test: -When antibodies bind to erythrocytes, they do not always result in agglutination. This can result from the antigen/antibody ratio being in antigen excess or antibody excess or in some cases electrical charges on the red blood cells preventing the effective cross linking of the cells. These antibodies that bind to but do not cause agglutination of red blood cells are sometimes referred to as incomplete antibodies. In no way is

this meant to indicate that the antibodies are different in their structure, although this was once thought to be the case. Rather, it is a functional definition only. In order to detect the presence of non-agglutinating antibodies on red blood cells, one simply adds a second antibody directed against the immunoglobulin (antibody) coating the red cells. This antiimmunoglobulin can now cross link the red blood cells and result in agglutination. It is known as the Direct Coomb's test. Indirect Coomb's Test: - If it is necessary to know whether a serum sample has antibodies directed against a particular red blood cell and you want to be sure that you also detect potential non- agglutinating antibodies in the sample, an Indirect Coomb's test is performed. This test is done by incubating the red blood cells with the serum sample, washing out any unbound antibodies and then adding a second anti-immunoglobulin reagent to cross link the cells. PRECIPITATION: - In this test antigen is in solution. The antibody cross links antigen molecule in variable proportions & aggregates (precipitates) form. In the zone of equivalence, optimal proportions of antigen & antibody combine and the maximal amount of precipitate forms. In the zone of antibody excess, there is too much antibody for efficient lattice formation. In the zone of antigen excess, all antibody has combined , but precipitation is reduced because many antigen-antibody complexes are too small to precipitate. Radial Immunodiffusion (Mancini): - In radial immunodiffusion, antibody is incorporated into the agar gel as it is poured and different dilutions of the antigen are placed in holes punched into the agar. As the antigen diffuses into the gel, it reacts with the antibody and when the equivalence point is reached a ring of precipitation is formed. Immunoelectrophoresis: -In immunoelectrophoresis, a complex mixture of antigens is placed in a well punched out of an agar gel and the antigens are electrophoresed so that the antigen are separated according to their charge. After electrophoresis, a trough is cut in the gel and antibodies are added. As the antibodies diffuse into the agar, precipitin lines are produced in the equivalence zone when an antigen/antibody reaction occurs. Countercurrent electrophoresis: - In this test the antigen and antibody are placed in wells punched out of an agar gel and the antigen and antibody are electrophoresed into each other where they form a precipitation line as illustrated in Figure 15. This test only works if conditions can be found where the antigen and antibody have opposite charges. This test is primarily qualitative, although from the thickness of the band you can get some measure of quantity. Its major advantage is its speed.

Radioimmunoassay (RIA)/Enzyme Linked Immunosorbent Assay (ELISA): - Radioimmunoassays (RIA) are assays that are based on the measurement of radioactivity associated with immune complexes. In any particular test, the label may be on either the antigen or the antibody. Enzyme Linked Immunosorbent Assays (ELISA) are those that are based on the measurement of an enzymatic reaction associated with immune complexes. In any particular assay, the enzyme may be linked to either the antigen or the antibody. 1. Competitive RIA/ELISA for Ag Detection: - By using known amounts of a standard unlabeled antigen, one can generate a standard curve relating radioactivity (cpm) (Enzyme) bound versus amount of antigen. From this standard curve, one can determine the amount of an antigen in an unknown sample. Noncompetitive RIA/ELISA for Ag or Ab: -Noncompetitive RIA and ELISAs are also used for the measurement of antigens and antibodies. The bead is coated with the antigen and is used for the detection of antibody in the unknown sample. The amount of labeled second antibody bound is related to the amount of antibody in the unknown sample. This assay is commonly employed for the measurement of antibodies of the IgE class directed against particular allergens by using a known allergen as antigen and anti-IgE antibodies as the labeled reagent. It is called the RAST test (radioallergosorbent test). The amount of labeled second antibody that binds is proportional to the amount of antigen that bound to the first antibody. Immunofluorescence Immunofluorescence is a technique whereby an antibody labeled with a fluorescent molecule (fluorescein or rhodamine or one of many other fluorescent dyes) is used to detect the presence of an antigen in or on a cell or tissue by the fluorescence emitted by the bound antibody. Direct Immunofluorescence Indirect immunofluorescence Flow Cytometry COMPLEMENT FIXATION: - It consists of two steps. Antigen & antibody are mixed & measured amount of complement is added. If the antigen & antibody match, they will combine & take up the complement. An indicator system consisting of sensitized red blood cells (i.e. RBC+ anti RBC antibody ) is now added. If the antibody matched the antigen in the first step,

2.

complement is fixed & less or none is available to attach to the sensitized RBC. The RBC remain non-hemolyzed i.e. the test is +ve, because the patients serum had antibodies to that antigen. However, if the antibody did not attach to the antigen in the first step, the complement is free to attach to the sensitized red blood cells & they are lysed i.e. test is negative. Complement must be sensitized & the patients serum must be heated to 56 C for 30 min to inactivate any human complement activity. NEUTRALISATION TEST: - These use the ability of antibody to block the effect of toxins or the infectivity of viruses. These can be used in cell culture or in the host animals.

HYPERSENSTIVITY Hypersensitivity is an immune response that damages the body's own tissues. They are divided into four classes (Type I IV) based on the mechanisms involved and the time course of the hypersensitive reaction. TYPE 1 HYPERSENSTIVITY: - Common allergens are Pollen, Animal Danders, Foods (nuts, shellfish ) and various drugs. The mechanism of reaction involves preferential production of IgE, in response to certain antigens (often called allergens). The precise mechanism as to why some individuals are more prone to type-I hypersensitivity is not clear. However, it has been shown that such individuals preferentially produce more of TH2 cells that secrete IL-4, IL-5 and IL-13 which in turn favor IgE class switch. IgE has very high affinity for its receptor (Fc; CD23) on mast cells and basophils. A subsequent exposure to the same allergen cross links the cell-bound IgE and triggers the release of various pharmacologically active substances (figure 1). Cross-linking of IgE Fc-receptor is important in mast cell triggering. Mast cell degranulation is preceded by increased Ca++ influx, which is a crucial process; ionophores which increase cytoplasmic Ca++also promote degranulation, whereas, agents which deplete cytoplasmic Ca++ suppress degranulation. The agents released from mast cells and their effects are listed in Table 1. Mast cells may be triggered by other stimuli such as exercise, emotional stress, chemicals (e.g., photographic developing medium, calcium ionophores, codeine, etc.),anaphylotoxins (e.g., C4a, C3a, C5a, etc.). These reactions, mediated by agents without IgE-allergen interaction, are not hypersensitivity reactions, although they produce the same symptoms. CLINICAL MANIFESTATIONS include Urticaria, Eczema, Rhinitis, Conjunctivitis and Asthma.

TREATMENT & PREVENTION: - Administration of antihistamines, which either inhibit the production of histamine or block histamines at receptor sites. After the administration of anti-histamines, E antibody receptor sites on the mast cells are blocked, thereby preventing the release of the histamines that cause the allergic reactions. The allergens are still there, but the body's allergic reactions are suspended for the period of time that the antihistamines are active. Antihistaminics, also constrict the smaller blood vessels and capillaries, thereby removing excess fluids. Decongestants can bring relief as well, but they can be used for only a short time, since their continued use can irritate and intensify the allergic reaction. In cases of extreme allergic reaction leading to anaphylactic shock, the patient may require an injection of epinephrine (also sometimes called adrenaline), a hormone that the body produces for responding to situations of fear and danger. In the case of anaphylactic shock, which involves such severe constriction of the breathing passages that the patient runs a risk of suffocation, epinephrine causes the passages to open, making it possible to breathe again. It also constricts the blood vessels, increasing the pressure and making the blood move more rapidly throughout the body. The body's own supply of epinephrine is not enough to counteract anaphylactic shock, however, and therefore a person experiencing that condition must receive an emergency injection containing many times the amount of the hormone naturally supplied by the body. It may be administered at a hospital, though doctors usually advise people with severe allergies to keep an emergency supply on hand. Maintenance of airway, support of respiratory & cardiac function should be provided. Identification and reactions. MEDIATORS Histamine increases bronchoconstriction, mucus secretion, vasodilatation, vascular permeability Serotonin increases capillary dilation, vascular permeability and smooth muscle contraction SRS A ECF-A Prostaglandins increase vascular permeability, smooth muscle contraction Attracts Eosinophils and releases histaminase & arylsulfatase Cause dilation & increase permeability of capillaries avoidance of allergen in future prevents an such untoward

Newly formed mediators leukotriene B4 leukotriene C4 , D 4 prostagland ins D2 PAF basophil attractant same as histamine but 1000x more potent edema and pain platelet aggregation and heparin release: microthrombi

TYPE 2 HYPERSENSTIVITY: - It occurs when antibody directed at antigens of cell membrane activates the complement. This generates a membrane attack complex which damages the cell membrane. The antibodies attach to antigen via its Fab region & acts as a bridge to complement via its Fc region. This phenomenon is seen in Autoimmune hemolytic Anemia Erythroblastosis fetalis Goodpastures syndrome Treatment involves anti-inflammatory and immunosuppressive agents.

TYPE III HYPERSENSITIVITY: - Type III hypersensitivity occurs as a result of immune complex deposition. They usually form when antigen is produced in excess of antibody. They may be localized to the site of antigen production or they may be found in the circulation. Immune complexes are usually cleared by the classical complement pathway or by the transfer of immune complexes by red blood cells to the liver or spleen for phagocytosis. The clearing mechanisms may be inadequate when there is excessive production of immune complexes. IgG in immune complexes activates complement as well as macrophages and neutrophils through Fc receptors to cause a hypersensitivity reaction. Complement activation aids in clearing the immune complexes but it also increases the permeability of blood vessels and is chemotactic. Activation of neutrophils, macrophages and platelets cause the release of proteolytic enzymes which damage blood vessels and initiate inflammation.

TYPE 4 HYPERSENSTIVITY: - This type of hypersensitive (allegic) reaction occurs when an antigen interacts with antigen-specific lymphocytes that release inflammatory and toxic substances, which attract other white blood cells and results in tissue injury. In type IV hypersensitivity, CD8 cytotoxic T-cells and CD4 helper T-cells recognize either intracellular or extracellular synthesized antigen when it is complexed with class I or class II MHC molecules, respectively. Macrophages function as antigen-presenting cells and release interleukin-1 , which promotes the proliferation of helper T-cells. Helper T-cells release interferon -gamma and interleukin-2, which together regulate delayed hypersensitivity reactions centered on macrophage activation and T-cell mediated immunity. When an individual is first exposed to protein antigen of tubercle bacilli, nave CD4+ T cell recognize peptides derived from these antigen. This is followed by differentiation of nave CD4+ T cell to Th-1 cell. Some of Th-1 cell enter circulation & enter the memory pool of T cells for long period. On second exposure, these Th-1 cell secrete cytokines (IFN gamma) which causes DTH. Interferon gamma is Powerful activator of Macrophages Activated macrophages are altered in several ways. Their ability to Phagocytose is greatly augmented. They express more Class 2 molecule on the surface and secrete TNF, IL-1 & Chemokines which promote inflammation. They produce more IL-12 which amplify Th-I response. Thus activated macrophages serve to eliminate offending antigen. CONTACT DERMATITIS On repeat exposure to antigen, sensitized CD4+ cells of Th-1 accumulates in the dermis. They then migrate toward the antigen within epidermis. Here they release cytokines that damage keratinocytes, causing sepration of these cell & formation of an intraepidermal vesicle. In certain other forms of DTH, especially those follow viral infections, cytokine producing CD8+ cells may be dominate effector cells. Type V This is an additional type that is sometimes (often in the UK) used as a distinction from Type 2.[5]

Instead of binding to cell surface components, the antibodies recognise and bind to the cell surface receptors, which either prevents the intended ligand binding with the receptor or mimics the effects of the ligand, thus impairing cell signaling. Some clinical examples:

Graves' disease Myasthenia gravis

The use of Type 5 is rare. These conditions are more frequently classified as Type 2, though sometimes they are specifically segregated into their own subcategory of Type 2.

Antibody Dependant Cell Mediated Cytotoxicity (ADCC) Type VI Hypersensitivity Type VI reaction according to the Gell and Coombs Classification . A phenomenon in which target cells, coated with antibody, are destroyed by specialized killer cells ( NK cells , Killer T-cells and macrophages ), which bear receptors for the Fc portion of the coating antibody (Fc receptors). These receptors allow the killer cells to bind to the anti-body-coated target.Eosinophils kill helminths (parasitic larvae infections) by ADCC. Comparison of Different Types of hypersensitivity type-I (anaphyla ctic) IgE exogenous 15-30 minutes weal & flare basophils type-II (cytotoxic ) IgG, IgM cell surface minuteshours lysis and necrosis antibody type-III (immun e comple x) IgG, IgM soluble 3-8 hours erythem a and edema, necrosis complem type-IV (delayed type) None tissues & organs 48-72 hours erythema and induratio n monocyte

characteri stics

antibody antigen response time

appearance

histology

and eosinophil transferred with Antibody

and complemen t antibody erythroblas tosis

ent and neutroph ils antibody SLE, farmer's lung disease

s and lymphocy tes T-cells tuberculi n test, poison ivy, granulom a

examples

allergic asthma, hay fever

fetalis, Goodpastur e's nephritis

IMMUNODEFICIENCY DISORDERS Immunodeficiencies occur when a part of the immune system is not present or is not working properly. Sometimes a person is born with an immunodeficiency these are called primary immunodeficiencies. Although primary immunodeficiencies are conditions that a person is born with, symptoms of the disorder sometimes may not show up until later in life.

Characteristic infections of the primary immunodeficiencies compon ent T-cells primary pathogen intracellular, bacteria viruses, protozoa, fungi, pneumococcus, streptococcus, haemophilus enteric bacteria and viruses phagocy tes Staphylococcal, Klebsiella Pseudomonas, primary site clinical example

non-specific

SCID, DiGeorge IgG, IgM deficiency

lung, skin, CNS IgG, IgM deficiency GI, nasal, eye lung, skin, regional lymph node IgA deficiency chronic granulomatous disease (CGD)

B-cells

comple ment

neisseria, Haemophilus, pneumococcus, streptococcus

CNS lung skin

C3, Factors I and H, late C components

NON-SPECIFIC IMMUNE SYSTEM - DEFECTS IN THE MYELOID LINEAGE Primary immunodeficiencies of the non-specific immune system include defects in phagocytic and NK cells and the complement system. Congenital Agranulomatosis Patients have a decrease in the neutrophil count. This is due to a defect in the myeloid progenitor cell differentiation into neutrophils. These patients are treated with granulocyte-macrophage colony stimulating factor (GM-CSF) or G-CSF. Defects of the phagocytic system Defects of phagocytic cells (numbers and/or functions) can lead to increased susceptibility to a variety of infections. Cyclic neutropenia This is marked by low numbers of circulating neutrophil approximately every three weeks. The neutropenia lasts about a week during which the patients are susceptible to infection. The defect appears to be due to poor regulation of neutrophil production. Chronic granulomatous disease (CGD) CGD is characterized by marked lymphadenopathy, hepato- splenomegaly and chronic draining lymph nodes. Leukocytes have poor intracellular killing (figure 5) and low respiratory burst. In majority of these patients, the deficiency is due to a defect in NADPH oxidase (cytochrome b558 : gp91phox, or rarely gp22phox) or other cofactor proteins (gp47phox, gp67phox) that participate in phagocytic respiratory burst. These patients can be diagnosed on the basis of poor Nitroblue tetrazolium (NBT) reduction which is a measure of respiratory burst. Interferon-gamma therapy has been successful. Leukocyte Adhesion Deficiency In this disease, T cells and macrophages lack the complement receptor CR3 due to a defect in CD11 or CD18 peptides and consequently they cannot respond to C3b opsonin. Alternatively there may a defect in integrin molecules, LFA-1 or mac-1 arising from defective CD11a or CD11b peptides, respectively. These molecules are involved in diapedesis and hence defective neutrophils cannot respond effectively to chemotactic signals. Treatment is with bone marrow (devoid of T cells and MHCmatched) transplantation or gene therapy. Chediak-Higashi syndrome Chediak-Higashi syndrome is marked by reduced (slower rate) intracellular killing and chemotactic movement accompanied by inability of phagosome and lysosome fusion and proteinase deficiency. Giant lysosomes (intracellular granules) are often

seen (figure 6). The respiratory burst is normal. Accompanying NK cell defects and platelet and neurological disorders are noted.

Immunodeficiencies can also be acquired through infection or produced by drugs. These are sometimes called secondary immunodeficiencies. Immunodeficiencies can affect B lymphocytes, T lymphocytes, or phagocytes. IgA deficiency is the most common immunodeficiency disorder. IgA is an immunoglobulin that is found primarily in the saliva and other body fluids that help guard the entrances to the body. IgA deficiency is a disorder in which the body doesn't produce enough of the antibody IgA. People with IgA deficiency tend to have allergies or get more colds and other respiratory infections, but the condition is usually not severe. Severe combined immunodeficiency (SCID) is also known as the "bubble boy disease" after a Texas boy with SCID who lived in a germ-free plastic bubble. SCID is a serious immune system disorder that occurs because of a lack of both B and T lymphocytes, which makes it almost impossible to fight infections. DiGeorge syndrome (thymic dysplasia), a birth defect in which children are born without a thymus gland, is an example of a primary T-lymphocyte disease. The thymus gland is where T lymphocytes normally mature. Chediak-Higashi syndrome and chronic granulomatous disease both involve the inability of the neutrophils to function normally as phagocytes. Secondary immunodeficiencies include: HIV (human immunodeficiency virus) infection/AIDS (acquired immunodeficiency syndrome) is a disease that slowly and steadily destroys the immune system. It is caused by HIV, a virus which wipes out certain types of lymphocytes called T-helper cells. Without T-helper cells, the immune system is unable to defend the body against normally harmless organisms, which can cause life-threatening infections in people who have AIDS. Research into HIV pathogenesis continued to reveal new aspects of human immunology. Clark et al4 showed that the number and frequency of TH1/TH17 CD4 T cells were reduced in HIV-infected patients and were only partially reconstituted with antiretroviral treatment, indicating a possible contributing factor to the increased prevalence of allergic disease in this patient population. Also in these patients Pallikkuth et al5 reported that increases in memory B-cell frequencies and serum levels of B-cell activating factor and a proliferation inducing ligand were associated with good antibody titers against influenza vaccine, suggesting that

there was a threshold of B-cell function necessary to produce specific antibody responses. Immunodeficiencies caused by medications. Some medicines suppress the immune system. One of the drawbacks of chemotherapy treatment for cancer, for example, is that it not only attacks cancer cells, but other fast-growing, healthy cells, including those found in the bone marrow and other parts of the immune system. In addition, people with autoimmune disorders or who have had organ transplants may need to take immunosuppressant medications. These medicines can also reduce the immune system's ability to fight infections and can cause secondary immunodeficiency.

Immunodeficiencies associated with aging

These include a progressive decrease in thymic cortex, hypo-cellularity of and reduction in the size of thymus, a decrease in suppressor cell function and hence an increase in auto-reactivity, a decrease in CD4 cells functions. By contrast B cells functions may be somewhat elevated. Immunodeficiencies associated with malignancies and other diseases B cell deficiencies have been noted in multiple myeloma, Waldenstrom's macroglobulinemia, chronic lymphocytic leukemiaand well differentiated lymphomas. Hodgkin's disease and advanced solid tumors are associated with impaired T-cell functions. Most chemotherapeutic agents used for treatment of malignancies are also immunosuppressive. Other conditions in which secondary immunodeficiencies occur are sickle cell anemia, diabetes mellitus, protein calorie malnutrition, burns, alcoholic cirrhosis, rheumatoid arthritis, renal malfunction, etc. Immunology of primary immunodeficiencies
An increase in TH17 cell counts in patients with immunodysregulation, polyendocrinopathy, enteropathy, X-linked syndrome (IPEX) was demonstrated by Paserini et al,11 who also pointed out that dysfunctional, epigenetically imprinted regulatory T cells were present and might contribute to the inflammatory manifestations of this condition. Papinazath et al12 studied thymocyte development in a murine model of purine nucleoside phosphorylase deficiency and reported on the essential role of this enzyme in the survival of double-positive thymocytes, which underwent apoptosis when exposed to deoxyguanosine. T-cell proliferation and T-cell receptor (TCR) diversity were not affected, and the restoration of purine nucleoside phosphorylase prevented the observed apoptosis. Considering the potential role of matrix metalloproteinases (MMPs) in the development of pneumatoceles and aneurysms and the presence of signal transducer and activator of transcription 3binding sites in promoter regions of genes encoding these enzymes, plasma levels of these enzymes were investigated in patients with hyper-IgE syndrome (HIES).13 MMP8 and MMP9 levels were found to be increased along with decreased expression of MMP3 in patients with HIES compared with levels seen in control subjects, suggesting a possible pathogenetic mechanism.

AUTOIMMUNE DISORDERS Its a group of disorders in which a person's body actually rejects itself. This condition comes about when the ability of the immune system to read MHCs becomes scrambled, such that it fails to recognize cells from within the body and instead rejects them as though they came from outside. As a result, the body sets in

motion the same destructive operation against its own cells that it normally would carry out against bacteria, viruses, and other such harmful invaders. The reasons why the immune system becomes dysfunctional are not well understood, but most researchers agree that a combination of genetic, environmental, and hormonal factors plays into autoimmunity. They also speculate that certain mechanisms may trigger it. First, a substance normally restricted to one part of the body, and therefore not usually exposed to the immune system, is released into other areas, where it is attacked. Second, the immune system may mistake a component of the body for a similar foreign component. Third, cells of the body may be altered in some way, by drugs, infection, or some other environmental factor, so that they are no longer recognizable as "self" to the immune system. Fourth, the immune system itself may be dysfunctional, for instance, because of a genetic mutation. In autoimmune disorders, the immune system mistakenly attacks the body's healthy organs and tissues as though they were foreign invaders. Autoimmune diseases include: Lupus is a chronic disease marked by muscle and joint pain and inflammation. The abnormal immune response may also involve attacks on the kidneys and other organs. Juvenile rheumatoid arthritis is a disease in which the body's immune system acts as though certain body parts such as the joints of the knee, hand, and foot are foreign tissue and attacks them.

Scleroderma is a chronic autoimmune disease that can lead to inflammation

and damage of the skin, joints, and internal organs. Ankylosing spondylitis is a disease that involves inflammation of the spine and joints, causing stiffness and pain. Juvenile dermatomyositis is a disorder marked by inflammation and damage of the skin and muscles.

REFERENCES

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