Precipitation Reactions

Antibody and soluble antigen interacting in aqueous solution form a lattice that eventually develops into a visible precipitate. Antibodies that aggregate soluble antigens are called precipitins. Although formation of the soluble Ag-Ab complex occurs within minutes, formation of the visible precipitate occurs more slowly and often takes a day or two to reach completion.

Formation of an Ag-Ab lattice depends on the valency of both the antibody and antigen:

■ The antibody must be bivalent; a precipitate will not form with monovalent Fab fragments.

■ The antigen must be either bivalent or polyvalent; that is, it must have at least two copies of the same epitope, or have different epitopes that react with different antibodies present in polyclonal antisera.

Experiments with myoglobin illustrate the requirement that protein antigens be bivalent or polyvalent for a precip-itin reaction to occur. Myoglobin precipitates well with specific polyclonal antisera but fails to precipitate with a specific monoclonal antibody because it contains multiple, distinct epitopes but only a single copy of each epitope (Figure 6-4a). Myoglobin thus can form a crosslinked lattice structure with polyclonal antisera but not with monoclonal antisera. The principles that underlie precipitation reactions are presented because they are essential for an understanding of commonly used immunological assays. Although various modifications of the precipitation reaction were at one time the major types of assay used in immunology, they have been largely replaced by methods that are faster and, because they are far more sensitive, require only very small quantities of antigen or antibody. Also, these modern assay methods are not limited to antigen-antibody reactions that produce a precipitate. Table 6-3 presents a comparison of the sensitivity, or minimum amount of antibody detectable, by a number of immunoas-says.

POLYCLONAL ANTISERUM

Myoglobin

POLYCLONAL ANTISERUM

Myoglobin

Precipitation Reaction Immunology
MONOCLONAL ANTIBODY
Immunology Precipitation Curve

FIGURE 6-4

Antigen-excess zone

Supematants excess Ab excess Ag

Antibody precipitated

FIGURE 6-4

Precipitation reactions. (a) Polyclonal antibodies can form lattices, or large aggregates, that precipitate out of solution. However, if each antigen molecule contains only a single epitope recognized by a given monoclonal antibody, the antibody can link only two molecules of antigen and no precipitate is formed. (b) A precipitation curve for a system of one antigen and its antibodies. This plot of the amount of antibody precipitated versus increasing antigen concentrations (at constant total antibody) reveals three zones: a

Antibody-excess Equivalence zone zone

Antigen-excess zone

Precipitation Reaction

Antigen added zone of antibody excess, in which precipitation is inhibited and antibody not bound to antigen can be detected in the supernatant; an equivalence zone of maximal precipitation in which antibody and antigen form large insoluble complexes and neither antibody nor antigen can be detected in the supernatant; and a zone of antigen excess in which precipitation is inhibited and antigen not bound to antibody can be detected in the supernatant.

Antigen added zone of antibody excess, in which precipitation is inhibited and antibody not bound to antigen can be detected in the supernatant; an equivalence zone of maximal precipitation in which antibody and antigen form large insoluble complexes and neither antibody nor antigen can be detected in the supernatant; and a zone of antigen excess in which precipitation is inhibited and antigen not bound to antibody can be detected in the supernatant.

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Responses

  • kathrin
    Can monoclonal antibodies be used to form lattices?
    8 years ago
  • Marco
    Why myoglobin precipitate with polyclonal antisera?
    8 years ago
  • GILLY
    Do precitation reaction occurs in monovalent antibody and antigen having one epitope?
    8 years ago
  • BALDOVINO LI FONTI
    Why would antibody precipitate out of solution?
    8 years ago
  • Italo Romano
    Why precipitate in polyclonal but not monoclonal?
    8 years ago
  • Paul
    What must both the antigen and antibody be to perform a precipitation reaction?
    8 years ago
  • Fearne
    What is the antigen excess in precipitation?
    7 years ago
  • erik
    Can monoclonal antibodies form large lattices?
    6 years ago
  • Hagosa
    Why does precipitation form in chemical reactions in immun?
    2 years ago
  • CLAIRE
    Can precipitation occur with monovalent Ag and Ab?
    4 months ago
  • emilia rosenbaum
    Does precipitate form if antigen added before antibody?
    3 months ago

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