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Antigen is mixed with antibodies in tubes. Precipitate forms and falls to the bottom of the tube.

Increasing amounts of antigen added

Increasing amounts of antigen added

Soluble antigen

Zone of antigen _ excess

Zone of optimal proportions

Zone of antibody excess

Soluble antigen

Antigen is carefully layered over serum to prevent mixing. In the excess of soluble antigen molecules, cross-linking cannot occur and no precipitate forms.

In the zone of optimal proportions, antigen and antibody molecules mix and are extensively cross-linked and precipitation occurs.

Serum containing specific antibodies; antibody molecules in excess of antigen, so cross-linking cannot occur and no precipitate forms.

Figure 17.3 Antigen-Antibody Precipitation Reactions (a) Precipitin curve in the tube precipitation test. (b) The ring precipitation test.Why does the amount of precipitate first increase and then decrease as more antigen is added?

Antigen is carefully layered over serum to prevent mixing. In the excess of soluble antigen molecules, cross-linking cannot occur and no precipitate forms.

In the zone of optimal proportions, antigen and antibody molecules mix and are extensively cross-linked and precipitation occurs.

Serum containing specific antibodies; antibody molecules in excess of antigen, so cross-linking cannot occur and no precipitate forms.

Figure 17.3 Antigen-Antibody Precipitation Reactions (a) Precipitin curve in the tube precipitation test. (b) The ring precipitation test.Why does the amount of precipitate first increase and then decrease as more antigen is added?

Precipitation occurs in a zone of optimal proportion— that is, the proportion at which both antigen and antibody are fully incorporated in the precipitate. When soluble antigen and antibody are mixed together in tubes, the precipitate will form and fall to the bottom of the tube (figure 17.3a). If, however, antiserum is placed in a small tube and the antigen is carefully layered over it to avoid mixing, the precipitate will form in a line where antigen and antibody meet in optimal proportions

430 Chapter 17 Applications of Immune Responses

(figure 17.3b). This is called the ring test, and it illustrates an important principle of antigen-antibody precipitation reactions: if there is a great excess of either antigen or antibody, a cross-linking lattice cannot form; consequently, no precipitate is formed. When these same interactions occur in the body, rather than in a laboratory test, large complexes are removed from the circulation by phagocytes, but small primary immune complexes with a slight antigen excess tend to remain in the circulation and can cause disease. â–  immune complex diseases, p. 448

Immunodiffusion Tests

Immunodiffusion tests are precipitation reactions carried out in agarose or other gels. A simple application of the immunodiffu-

sion technique is the radial immunodiffusion test, which is a quantitative test. It can be used, for example, to measure the amount of IgG in the serum if an immune abnormality is suspected. Antibodies specific for human IgG molecules are added to melted agar. After the agar hardens, wells are cut in the agar and the person's serum is added to a well. In this case the IgG in the serum is the antigen being tested for. A precipitin ring forms where the antigen and antibody meet in optimal proportion. The diameter of the ring that forms is compared with a standard curve prepared from samples with known concentrations of the antigen IgG. The concentration of IgG in the patient's serum can be read directly from the standard curve (figure 17.4).

Increasing concentrations of known antigen are placed in wells cut in a gel that contains antibody mixed throughout.

Increasing concentrations of known antigen are placed in wells cut in a gel that contains antibody mixed throughout.

Unknown samples A and B are placed in wells.

Antigen diffuses from the wells into the antibody-containing gel.

Unknown samples A and B are placed in wells.

Antigen diffuses from the wells into the antibody-containing gel.

Precipitation occurs where the antigen and antibody meet in optimal proportions; the diameter of each ring is measured.

Precipitation occurs where the antigen and antibody meet in optimal proportions; the diameter of each ring is measured.

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