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Dialogue: 0,0:00:00.00,0:00:06.00,Default,,0000,0000,0000,,(English captions by Andrea Matsumoto, University of Michigan.) Agglutination assays have been used for decades as a simple method to detect antigenic substances
Dialogue: 0,0:00:06.00,0:00:07.00,Default,,0000,0000,0000,,in biologic samples.
Dialogue: 0,0:00:07.00,0:00:12.00,Default,,0000,0000,0000,,The purpose of this video is to explain how\Nthis method works in practice and to expose
Dialogue: 0,0:00:12.00,0:00:15.00,Default,,0000,0000,0000,,its limitations.
Dialogue: 0,0:00:15.00,0:00:20.00,Default,,0000,0000,0000,,The agglutination assay uses tiny particles,\Nmost often latex beads.
Dialogue: 0,0:00:20.00,0:00:24.00,Default,,0000,0000,0000,,The beads are coated with a specific antibody\Nagainst the antigen that you would like to
Dialogue: 0,0:00:24.00,0:00:27.00,Default,,0000,0000,0000,,detect.
Dialogue: 0,0:00:27.00,0:00:33.00,Default,,0000,0000,0000,,The test is usually performed on a card or,\Nglass or plastic slide, often one with a black
Dialogue: 0,0:00:33.00,0:00:34.00,Default,,0000,0000,0000,,surface.
Dialogue: 0,0:00:34.00,0:00:39.00,Default,,0000,0000,0000,,First you add a suspension of the coated latex\Nbeads to each of the three encircled areas
Dialogue: 0,0:00:39.00,0:00:41.00,Default,,0000,0000,0000,,on the slide.
Dialogue: 0,0:00:41.00,0:00:49.00,Default,,0000,0000,0000,,Note that the suspension is concentrated enough\Nto produce a milky appearance on the background.
Dialogue: 0,0:00:49.00,0:00:53.00,Default,,0000,0000,0000,,Now you add a few drops of the unknown sample\Nthat you are interested testing.
Dialogue: 0,0:00:53.00,0:00:59.00,Default,,0000,0000,0000,,But, you will also need to use one circled\Narea for a negative control solution that
Dialogue: 0,0:00:59.00,0:01:04.00,Default,,0000,0000,0000,,contains no antigen and another for a positive\Ncontrol solution that contains the antigen
Dialogue: 0,0:01:04.00,0:01:12.00,Default,,0000,0000,0000,,of interest.
Dialogue: 0,0:01:12.00,0:01:20.00,Default,,0000,0000,0000,,Next the slide is gently rocked or swirled\Nto mix the beads with the test solutions and
Dialogue: 0,0:01:20.00,0:01:25.00,Default,,0000,0000,0000,,the samples containing the antigen of interest\Nwill begin to agglutinate the beads.
Dialogue: 0,0:01:25.00,0:01:29.00,Default,,0000,0000,0000,,This will produce the appearance of visible\Nclumps and the solution itself will turn from
Dialogue: 0,0:01:29.00,0:01:34.00,Default,,0000,0000,0000,,milky in appearance to clear and transparent.
Dialogue: 0,0:01:34.00,0:01:36.00,Default,,0000,0000,0000,,This transition should occur in the area with\Nthe positive control.
Dialogue: 0,0:01:36.00,0:01:41.00,Default,,0000,0000,0000,,If the antigen is present in the unknown sample\Nthen it will form clumps.
Dialogue: 0,0:01:41.00,0:01:47.00,Default,,0000,0000,0000,,The negative control circle should remain\Nunclumped and opaque.
Dialogue: 0,0:01:47.00,0:01:52.00,Default,,0000,0000,0000,,Recall that the latex beads are coated with\Na specific antibody so that each bead can
Dialogue: 0,0:01:52.00,0:01:54.00,Default,,0000,0000,0000,,bind to numerous antigens.
Dialogue: 0,0:01:54.00,0:02:00.00,Default,,0000,0000,0000,,For agglutination to work the antigen of interest\Nmust also be able to bind to multiple beads.
Dialogue: 0,0:02:00.00,0:02:05.00,Default,,0000,0000,0000,,Therefore in this assay, antigens that can\Nbe detected are limited to large macromolecules
Dialogue: 0,0:02:05.00,0:02:09.00,Default,,0000,0000,0000,,that have repetitive antigenic domains.
Dialogue: 0,0:02:09.00,0:02:15.00,Default,,0000,0000,0000,,Molecules like microbial capsules, flagella,\Nor lipopolysaccharides.
Dialogue: 0,0:02:15.00,0:02:20.00,Default,,0000,0000,0000,,One long repeating antigen molecule can then\Nattach to several beads causing them to clump
Dialogue: 0,0:02:20.00,0:02:23.00,Default,,0000,0000,0000,,together or, agglutinate.
Dialogue: 0,0:02:23.00,0:02:27.00,Default,,0000,0000,0000,,So even very tiny quantities of antigens that\Nhave lots of repeating antigenic domains can
Dialogue: 0,0:02:27.00,0:02:32.00,Default,,0000,0000,0000,,cause visible clumps to form and be detected\Nby this test.
Dialogue: 0,0:02:32.00,0:02:35.00,Default,,0000,0000,0000,,This is the basis of the test.
Dialogue: 0,0:02:35.00,0:02:39.00,Default,,0000,0000,0000,,Finally here are some examples of agglutination\Nassays that are used in clinical practice.