Supplemental Lecture (98/05/24 update) by Stephen T. Abedon (abedon.1@osu.edu)

  1. Chapter title: Humoral Immunity
    1. A list of vocabulary words is found toward the end of this document
    2. We will limit our discussion of specific host defences in this lecture to humoral immunity (but discuss cell mediated immunity when discussing viral cytopathic effects). Humoral immunity is that mediated by antibodies. Note that just because I am limiting this discussion to humoral immunity in no way implies that humoral immunity is any more or less important than cell mediated immunity. However, one is often more important than the other, depending on the pathogen.
  2. Acquired immunity
    1. Acquired immunity is the protection an animal develops against microorganisms or foreign substances.
    2. Acquired immunity is generally split into two types:
      1. that associated with the humoral arm of the immune system
      2. that associated with cell-mediated immunity
  3. Humoral immunity [antibody-mediated immunity, lymphocytes, B cells]
    1. Antibodies:
      1. Humoral immunity is that aspect of specific immunity which:
        1. is mediated by B lymphocytes
        2. involves the binding of antibodies to antigens
      2. Contrast humoral immunity with cell mediated (cellular) immunity. Together they constitute specific (as well as acquired) immunity.
    2. Characteristics:
      1. Acquired humoral immunity is the recognition of antigens associated with microorganisms or foreign substances.
      2. This recognition is coupled with the ability to initiate appropriate actions (e.g., antibody production) against these microorganisms or foreign substances.
    3. Acquired humoral immunity may be subdivided into:
      1. that acquired actively
      2. that acquired passively
  4. Antigen [immunogen]
    1. An antigen (a.k.a, immunogen) is a molecule which is capable of stimulating specific immunity.
    2. Cellular immunity:
      1. For immunity mediated by cytoxic T cells, an antigen is always a protein or a portion of a protein.
      2. This is because only peptides (protein fragments) interact with MHC (major histocompatibility complex) and cell mediated immunity requires interaction with these MHC-associated peptides.
      3. For humoral immunity, an antigen is anything an antibody can bind to, though usually this is a protein or a carbohydrate.
    3. Antibody dependent cellular cytotoxicity:
      1. For antibody dependent cellular cytotoxicity (ADCC), an antigen, too, is anything an antibody can bind to.
      2. Note, though, that ADCC is limited to interacting with antibodies associated with components found on the surface of eucaryotic cells.
  5. Antibody [immunoglobulin, Ig, IgG]
    1. Antigen binding proteins:
      1. An antibody is any one of a number of related host-produced proteins.
      2. Antibodies bind to antigens at very specific sites called antigenic determinants or epitopes.
      3. Generally, antibodies have binding domains (which are found on the antibody) and these binding domains interact with specific sites on antigens.
      4. This interaction is analogous to the binding of a virion adsorption protein to the receptor molecule found on a host cell.
      5. Antibodies are produced in high numbers and in great variety. Generally they are able to bind to antigens in numerous ways and also to many different parts of antigens.
    2. High specificity of antibody binding:
      1. Individually, antibodies tend to be limited to very binding to very specific regions of specific antigens.
      2. There exist mechanisms which lead to the evolution, over short periods (days), of antibody variants which display high specificity as well as strong binding to specific antigens.
      3. Simultaneously, the production of antibodies which can bind to host proteins is actively suppressed.
      4. Antibody binding consequently tags an antigen as foreign.
  6. Epitope [antigenic determinant]
    1. Antibody binding site:
      1. An epitope is the specific site on an antigen to which an antibody binds.
      2. Epitopes generally are significantly smaller than the antigens which contain them (and much smaller than the size of an antibody).
      3. A single antigen often contains numerous epitopes.
    2. Many antibodies to single antigen:
      1. A humoral immune response very often consists of numerous antibodies which collectively recognize many but not all the epitopes found on a given antigen.
      2. Very often individual epitopes may be recognized by more than one kind of antibody.
  7. Active immunity
    1. Learned immunity:
      1. A normal functioning immune system is capable of recognizing antigens and amplifying antigen-recognizing cells.
      2. The ability of an animal to recognize and respond to an antigen consequently increases with exposure (at least to certain limits).
      3. For humoral immunity, the cells which are amplified (increased in number/stimulated to divide) are those which produce antibody, B cells.
    2. Active immunity simply is this normal "recognize-respond" action of the immune system.
  8. Passive immunity
    1. Transfer of active immunity:
      1. In contrast to active immunity, passive immunity is the transfer of active immunity from one individual to another.
      2. Because antibodies are soluble proteins found in the blood, this transferance is relatively simple and requires, for example, only a simple transfusion of the antibody component of blood (often called gamma globulin).
    2. Mother to child:
      1. The most important natural occurrence of passive immunity is that from mother to infant.
      2. This occurs either via transplacental transfer or via breast milk, especially colostrum.
    3. Powerful but temporary:
      1. Passively acquired immunity can be very powerful in fighting disease.
      2. The effects of passive immunity, however, are both temporary and cannot be restored without additional passive infusion.
      3. Passive immunity lasts only as long as infused antibody lasts, which is only on the order of weeks.
    4. Medically, antitoxin and antiserum play important roles in disease fighting via administration of passive immunity.
  9. Antiserum [gamma globulin]
    1. Liquid portion of blood:
      1. Serum is the yellow liquid left over following the clotting of blood.
      2. Serum is packed with antibodies.
    2. Serum specificity:
      1. Serum from individuals who have been exposed to a pathogen (or vaccine) usually contains antibodies that recognize the pathogen (thus accounting for some fraction of the active immunity displayed by that individual against the pathogen).
      2. Such serum, when employed in passive immunity, is referred to as antiserum.
    3. Antiserum used for passive immunity is often derived from humans or horses though each source has its problems:
      1. the former is a human blood product and therefore human-derived antiserum potentially carries human pathogens (though these may be removed by employing various procedures)
      2. the latter (horse serum) is recognized as foreign by the (human) recipient consequently making horse-derived passive immunity a treatment which may be safely employed only once or only a few times before active immunity against horse antigens develop
  10. Artificially acquired immunity
    1. Various medical procedures can induce immunity artifically.
    2. The administration of antiserum is an example of artificially acquired passive immunity.
    3. Vaccinaition, on the other hand, is an example of artificially acquired active immunity.
  11. Antibody diversity
    1. The ability of an animal to recognize an antigen by antibody-binding is a two-fold process:
      1. antibodies that bind to the antigen must exist
      2. antibodies that bind to the antigen must exist in sufficient number for recognition to be acted upon meaningfully
    2. generating variation:
      1. The first criteria is achieved in the course of the replication of antibody-producing cells (B cells) through a combination of recombination between the genes associated with many antibody types, and later through the random mutation of antibody genes (the latter called somatic point mutation).
    3. Amplification of recognizing antibody:
      1. The second criteria is achieved through the amplification of those antibody-producing cells that produce antibodies which bind novel (not self) antigens.
      2. Humoral immunity, thus, is not so much "acquired" as amplified and refined in response to exposure to non-self antigens.
    4. Memory:
      1. Once this amplification has occurred it is institutionalized in so-called memory cells.
      2. The existence of memory cells is the physiological manifestation of the priming of humoral immunity, i.e., for future recognition of specific antigens.
  12. Consequence of antibody binding
    1. Once an antigen is bound by an antibody, any of four actions may occur:
      1. the antigen may no longer function properly (see virus inactivation by antibody)
      2. the antigen and/or attached microorganism may clump (a process called agglutination) or even precipitate out of solution
      3. complement may be activated
      4. antibody-dependent cell-mediated cytotoxicity may occur if the antigen is found on the surface of one of the body's own cells
    2. Agglutination:
      1. Agglutination can occur because antibodies display multivalence: An ability to bind more than one epitope simultaneously (i.e., most antibodies have more than one antigen binding site).
      2. A consequence of agglutination is increased susceptibility to phagocytosis.
  13. Monoclonal antibodies [Mabs]
    1. Single type of antibody:
      1. Monoclonal antibodies (MAbs) are a way of isolating and making large quantities of only a single type of antibody.
      2. Remember that though a full complement of antibodies may show great variation in specificity, individual antibodies tend to be fairly specific in what they will bind and how strongly they will bind.
      3. Normally pure, large quantities of any individual antibody are difficult to produce within an animals.
      4. Given the existence of large numbers of pure antibody, however, there exist numerous techniques one can employ these antibodies in. Particular, these techniques have in common a requirement for very specific antibody binding.
      5. The primary advantage of monoclonal antibodies (i.e., they come from a single B cell clone) is that their properties are uniform (within type) and they bind to only a single epitope. Thus, they have high specificity and predictability.
      6. A downside of MAbs are that their biological properties are very limited compared with antiserum (polyclonal antibodies) and therefore their ability to recognize different varieties of individual antigens is limited.
    2. Example: diagnostics:
      1. MAb-based diagnostic tests measure the ability of specific MAbs to bind to specific antigens, the presence of which might indicate the existence of a pathology such a pathogen infection.
      2. Other uses of monoclonal antibodies range from numerous applications in basic research to, potentially, treatment of disease via passive immunization.
    3. Creating MAbs:
      1. MAbs are created by inducing a humoral immune response in an animal (usually a mouse) then collecting the mouse's B cells (usally they are isolated from the animals spleen).
      2. These B cells are fused with mouse myeloma (cancer) cells to make them immortal (otherwise they would eventually die out during propagation in tissue culture). The resulting cell is called a hybridoma.
      3. The antibody produced by individual hybridomas is characterized.
      4. Desirable hybridomas (i.e., those making antibodies with desirable properties) may be grown and antibody produced via standard tissue culture techniques.
    4. Incompatibility across species:
      1. A problem with using mouse monoclonal antibodies in situ (i.e., within patients for therapeutic reasons) is the same as that experienced when employing horse antiserum, i.e., recognition by the host as foreign and formation of a host directed humoral immune response against infused antibody.
      2. These problems may be eliminated, however, by making mouse monoclonal antibodies more human-like through bioengineering.
      3. Additionally, it is now possible to make human monoclonal antibodies from human B cells.
  14. Distinguishing self from nonself
    1. How does the body pull off the seemingly miraculous feat of producing a huge variety (and number) of antibodies seemingly capable of recognizing nearly any antigen the body is exposed to, but simultaneously not produce antibodies which recognized (bind to) host cells? For T cell-mediated -immunity, the answer is that T cells mature in the thymus (hence the name T cell) where they are exposed to all the body's potential antigens and those which are capable of binding to self are induced to die. B cells, however, are not matured in the thymus. How is it that B cells which would otherwise bind to self antigens are rare?
    2. "For nearly 50 years, immunologists have thought that during embryonic development or early life the immune system undergoes a critical education in which it learns how to tolerate the body's own tissues while retaining the ability to mount an attack on bacteria, viruses, and any other foreign invaders." (p. 1665, Pennisi, 1996). However, this dogma, that the steps involved in distinguishing self from nonself only early in life, turns out to be not necessarily correct.
    3. Instead, the answer, based on recent research (see Pennisi, 1996), may be that antigens to which the immune system is exposed simply in very large amounts do not stimulate an immune response. However, antigens to which the body is exposed in moderate amounts, especially in a context which the body recognizes as having the hallmarks of infection ("danger!"), do stimulate active immunity. Thus, host antigens, which are highly prevalent and usually healthy, must suppress the propagation of an immune response against themselves.
    4. This model may in part explain why it is that infants are less capable of mounting immune responses especially, for example, against such things as vaccines. These very small people may perceive the antigen found in a normal vaccination to be so large (relative to themselves or, actually, the number of their "T cells and antigen-presenting cells" p. 1666, Pennisi, 1996) that it is perceived as "self." Later in life, when these small people are no longer quite so small, the ratio of antigen to body size (i.e., "T cells and antigen-presenting cells") is sufficiently larger that the antigen is not recognized as "self" and consequently can induce a reasonable immune response.
    5. In addition, "If reasearchers can control the type of immune response they induce by adjusting how much antigen they give relative to the amount of T cells and antigen-presenting cells in the recipients, they might, for example, be able to make people undergoing organ transplants tolerant to the donor tissue or arrest autoimmune diseases." (p. 1666, Pennisi, 1996)
  15. Vocabulary
    1. Acquired immunity
    2. Active immunity
    3. Antibody
    4. Antibody diversity
    5. Antibody-mediated immunity
    6. Antigen
    7. Antigenic determinant
    8. Antiserum
    9. Artificially acquired immunity
    10. Consequence of antibody binding
    11. Creating MAbs
    12. Distinguishing self from nonself
    13. Epitope
    14. Gamma globulin
    15. High specificity of antibody binding
    16. Humoral immunity
    17. Ig
    18. Immunogen
    19. Immunoglobulin
    20. MAbs
    21. Monoclonal antibodies
    22. Passive immunity
  16. Practice questions
    1. Acquired immunity specifically obtained through the transfer of serum from one individual to a second in order to fight disease is an example of __________? [PEEK]
    2. Three natural consequences of antibody binding include agglutination, inactivation, and mediation of antibody-dependent cell-mediated cytotoxicity. Name a fourth. [PEEK]
    3. Name a problem associated with using horses as a source of antisera. [PEEK]
    4. Which is smaller? (circle best answer) [PEEK]
      1. antigen.
      2. antibody.
      3. epitope.
      4. B cell.
      5. protein.
      6. viral envelope protein.
      7. toxoid.
    5. __________ is an example of artificial, passive immunity. [PEEK]
    6. Name a difference between humoral immunity and cell-mediated immunity (other than the generic answer: that the latter is mediated by "cells"). [PEEK]
    7. Describe agglutination in terms of the multivalent nature of the antibody. [PEEK]
    8. Name a disease for which artificially acquired passive immunity would be a effective treatment. [PEEK]
    9. Give an example of passively acquired natural immunity. [PEEK]
    10. Another name for an "antigenic determinant" is a(n) __________. [PEEK]
    11. A consequence of the agglutination of a pathogen is an increased tendency by certain immune system cells of your body to __________ the agglutinated pathogen. [PEEK]
    12. The primary advantage of __________ is that within type their properties are uniform and they bind to only a single epitope? (choose best answer to fill blank) [PEEK]
    13. antigen
      1. polyclonal antibody
      2. immunoglobulin
      3. monoclonal antibody
      4. active immunity
      5. acquired immunity
      6. antibody
  17. Practice question answers
    1. passive immunity
    2. activation of complement
    3. Repeated injection of horse sera results in the stimulation of an anti-horse-serum immune response.
    4. iii, epitope.
    5. administration of antitoxin or antiserum.
    6. Humoral immunity: B cells, antibody-mediated, etc.; Cell-mediated immunity: Natural killer cells, Cytotoxic T cells, antigen presented in context of MHC molecule.
    7. Agglutination in a clumping of antigens. It occurs because antibodies are able to bind more than one epitope simultaneously, thus bridging together antigens. Note that the agglutination of more than two individual antigen molecules by IgG requires that more than one type of IgG be present (i.e., a monoclonal antibody, MAb, is not nearly as efficient an agglutinator as polyclonal antibody, a.k.a., serum).
    8. tetanus, diphtheria, etc. Actually, almost any disease for which antibodies are an effective defence might be treated with artificially acquired passive immunity, though passive immunization is often saved for life threatening diseases for which other treatments are not as effective.
    9. Transfer of IgG from mother to infant or unborn child, e.g., across placenta or in colostrum.
    10. epitope.
    11. Phagocytize
    12. iv, monoclonal antibody
  18. References
    1. Black, J.G. (1996). Microbiology. Principles and Applications. Third Edition. Prentice Hall. Upper Saddle River, New Jersey. pp. 480-497.
    2. Pennisi, E. (1996). Teetering on the brink of danger. Science 271:1665-1667.
    3. Tortora, G.J., Funke, B.R., Case, C.L. (1995). Microbiology. An Introduction. Fifth Edition. The Benjamin/Cummings Publishing, Co., Inc., Redwood City, CA, pp. 424-447.