Antibodies reaction with antigen

The classical methods of demonstrating specificity of antibody reactions with antigens, such as immunodiffusion plates, are obsolete. Immunofluorescence is a very sensitive method that requires controls for specificity of equal sensitivity. The most important controls are not just those that demonstrate the presence of an antibody that reacts with the antigen of interest, but more importantly, the absence of antibodies that react with other things. 

The attachment of protein to the transducer surface for immunoassay is a difficult problem, since it must be achieved without interfering with the active site. If an unlabeled assay is to be performed it must also be such that subsequent non specific interaction with the surface can be inhibited. This requirement is often contrary to those for effective antibody-antigen complex formation. A surface close packed with antibody will be sterically hindered and its reaction with antigen inhibited. On the other hand, a suitably spaced packing allows non-specific interactions to occur and large false positive signals to be recorded. As mentioned earlier, this was demonstrated by Cullen and Lowe who used the surface plasmon resonance technique to probe specific and non-specific protein interactions at metal surfaces. (23). 

Industrial operations and chemical processing provide a suitable atmosphere for the induction of immunologic reactions as seen with skin sensitization and the respiratory tract. The antigen antibody reaction with a complex cascade-type system of serum proteins results in a variety of sensitizing reactions of the worker and disturbs his or her normal health. 

The assumption of bi- or multivalence of antibodies has been amply verified. IgG, IgE, and presumably IgD are bivalent electron micrographs show that IgA may exist as a bivalent molecule or as a tetravalent dimer, and IgM in various species may be a tetramer, pentamer, or hexamer with valences of 8, 10, and 12, respectively, although all of these may not be available for reaction with antigen simultaneously. Bivalence of IgG has been established by equilibrium dialysis, - fluores-... 

The formation of antigen-antibody complexes can lead to clinical syndromes such as the Arthus reaction. In this model, a high level of preformed specific IgG antibody combines with antigen to produce a localized edematous, erythematous reaction within 5 to 8 hours. The reaction involves local formation of insoluble antigen-antibody complexes, complement activation with release of C3a and C5a collectively referred to as anaphylatoxins, mast cell degranulation, and influx of polymorphonuclear cells. 

Two secoiridoids, hydramacrosides A (109) and B (110), isolated from the leaves of Hydrangea macrophylla var. thunbergii (Saxifragaceae) showed inhibitory effect on the histamine release from rat mast cell induced by antigen-antibody reaction with inhibition of 9.1% and 21.3%, respectively, at a concentration of 10 M [135]. 

A base, formed by the bacterial degradation of histidine, and present in ergot and in many animal tissues, where it is liberated in response to injury and to antigen-antibody reactions. If injected it causes a condition of shock with dilatation of many blood vessels, loss of plasma from the capillaries to the tissues and a rapid fall in blood pressure. It is normally prepared from protein degradation products. 

Enzyme Immunosensors. Enzyme immunosensors are enzyme immunoassays coupled with electrochemical sensors. These sensors (qv) require multiple steps for analyte determination, and either sandwich assays or competitive binding assays maybe used. Both of these assays use antibodies for the analyte of interest attached to a membrane on the surface of an electrochemical sensor. In the sandwich assay type, the membrane-bound antibody binds the sample antigen, which in turn binds another antibody that is enzyme-labeled. This immunosensor is then placed in a solution containing the substrate for the labeling enzyme and the rate of product formation is measured electrochemically. The rate of the reaction is proportional to the amount of bound enzyme and thus to the amount of the analyte antigen. The sandwich assay can be used only with antigens capable of binding two different antibodies simultaneously (53). 

Not all antigen-antibody reactions are of benefit to the body, as sometimes the complexes (or their subsequent interaction with body tissues) may result in tissue damage. This must be regarded as a malfunction of the immune system and is known as a hypersensitive reaction. These reactions can be categorized into five main types. The first three involve the interaction between antigen and humoral antibody, and as the onset of the reaction is rapid, the condition is termed immediate hypersensitivity. The fourth type (delayed hypersensitivity) involves T cells and the symptoms of the reaction appear after 24 hours. The fifth type is where antibody stimulates cell function. 

The limitations of ELISA methods include the specificity of antibodies, the concentrations of primary antibody and antigen, and the type of reaction solution. Nonspecific binding of either of the antibodies to related antigens, unrelated proteins of other bacteria, or even the microtiter plate may lead to false positive reactions.49,52 57 Use of a monoclonal antibody may decrease crossreactivity with other antigens. For detection of low numbers of bacteria, as in drinking water, the sample may be filtered to concentrate the cells or cultured in a selective broth until it reaches the minimum detection limit for ELISA.49,58 Commercial test kits using dipsticks, immunoblots, and sandwich ELISA methods have been developed for the identification of pathogenic bacteria.58,59... 

Fig. 10.5. Immunoblotting analysis of the antibody response of jirds against various stages of A viteae. (A) Reaction with sera of jirds vaccinated with irradiated L3 (B) reaction of sera of A. w feae-infected jirds (C) reaction with sera of naive jirds. 1, Male antigens 2, female antigens 3, mf antigens 4, L3 antigens. Note that the reaction of vaccinated jird sera is predominantly directed against chitinase bands of 205 kDa and 67 kDa (arrows) as well as against a 17 kDa protein.

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