(English captions by Andrea Matsumoto, University of Michigan.) Agglutination assays have been used for decades as a simple method to detect antigenic substances
in biologic samples.
The purpose of this video is to explain how this method works in practice and to expose
its limitations.
The agglutination assay uses tiny particles, most often latex beads.
The beads are coated with a specific antibody against the antigen that you would like to
detect.
The test is usually performed on a card or, glass or plastic slide, often one with a black
surface.
First you add a suspension of the coated latex beads to each of the three encircled areas
on the slide.
Note that the suspension is concentrated enough to produce a milky appearance on the background.
Now you add a few drops of the unknown sample that you are interested testing.
But, you will also need to use one circled area for a negative control solution that
contains no antigen and another for a positive control solution that contains the antigen
of interest.
Next the slide is gently rocked or swirled to mix the beads with the test solutions and
the samples containing the antigen of interest will begin to agglutinate the beads.
This will produce the appearance of visible clumps and the solution itself will turn from
milky in appearance to clear and transparent.
This transition should occur in the area with the positive control.
If the antigen is present in the unknown sample then it will form clumps.
The negative control circle should remain unclumped and opaque.
Recall that the latex beads are coated with a specific antibody so that each bead can
bind to numerous antigens.
For agglutination to work the antigen of interest must also be able to bind to multiple beads.
Therefore in this assay, antigens that can be detected are limited to large macromolecules
that have repetitive antigenic domains.
Molecules like microbial capsules, flagella, or lipopolysaccharides.
One long repeating antigen molecule can then attach to several beads causing them to clump
together or, agglutinate.
So even very tiny quantities of antigens that have lots of repeating antigenic domains can
cause visible clumps to form and be detected by this test.
This is the basis of the test.
Finally here are some examples of agglutination assays that are used in clinical practice.