Direct Agglutination

Direct agglutination occurs when antigens are found naturally

on a particle. One of the best examples of direct agglutination
testing involves known bacterial antigens used to test for the
presence of unknown antibodies in the patient. Typically, patient
serum is diluted into a series of tubes or wells on a slide
and reacted with bacterial antigens specific for the suspected
disease. Detection of antibodies is primarily used in diagnosis
of diseases for which the bacterial agents are extremely difficult
to cultivate. One such example is the Widal test, a rapid screening
test used to help determine the possibility of typhoid fever.
A significant finding is a fourfold increase in antibody titer over
time when paired dilutions of serum samples are tested with
any of these antigens. Although more specific tests are now
available, the Widal test is still considered useful in diagnosing
typhoid fever in developing countries and remains in use in
many areas throughout the world.15
If an agglutination reaction involves RBCs, then it is called
hemagglutination. The best example of this occurs in ABO
blood group typing of human RBCs, one of the world’s most
frequently used immunoassays.5 Patient RBCs mixed with antisera
of the IgM type can be used to determine the presence
or absence of the A and B antigens; this reaction is usually performed
at room temperature without the need for any enhancement
techniques. Group A RBCs will agglutinate with anti-A
antibody and Group B RBCs will agglutinate with anti-B antibody.
This type of agglutination reaction is simple to perform,
is relatively sensitive, and is easy to read (Fig. 10–10).
A titer that yields semiquantitative results can be performed
in test tubes or microtiter plates by making serial dilutions of
the antibody. The reciprocal of the last dilution still exhibiting
a visible reaction is the titer, indicating the antibody’s strength.
Interpretation of the test is done on the basis of the cell sedimentation
pattern. If there is a dark red, smooth button at the
bottom of the microtiter well, the result is negative. A positive
result will have cells that are spread across the well’s bottom,
usually in a jagged pattern with an irregular edge. Test tubes
also can be centrifuged and then shaken to see if the cell button
can be evenly resuspended. If it is resuspended with no visible
clumping, then the result is negative. Positive reactions can be
graded to indicate the strength of the reaction (Fig. 10–11).
Passive Agglutination
Passive, or indirect, agglutination employs particles that are
coated with antigens not normally found on their surfaces. A variety
of particles, including erythrocytes, latex, and gelatin, are
used for passive agglutination.4 The use of synthetic beads or
particles provides the advantages of consistency and uniformity.