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Detection of Intestinal Parasites



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Sandwich Assay

Slide 28

March 2009

To illustrate the principle behind our immunoassays, I’d like to use the following animation. A similar method is used for all of the immunoassays that we offer in the parasitology lab.

We start with a plastic microtiter well that is lined with antibody to Giardia antigen. The patient’s processed fecal sample is then added to the well. In this case, antigen is present, as denoted by these red circles. This antigen then binds to the antibodies.

Next, a second antibody is added to the well, which is also specific for the Giardia antigen. This is where the phrase “sandwich assay” comes from, because the antigen is sandwiched between two antibodies. The second antibody is conjugated to an enzyme called horseradish peroxidase. Washing of the well occurs between steps to remove any unbound antibody.

Finally, a substrate for this enzyme is added to the mix. If antigen is present, then the antibody conjugated to enzyme will be bound and will not have washed away in the previous steps. The enzyme now acts on its substrate which turns yellow after addition of the final reagent. The yellow color is then read by a spectrophotometer which can detect varying degrees of yellow color. As I mentioned earlier, this is the same method used for detection of Cryptosporidium antigen as well, although the antibodies are different.

Sandwich Assay

 



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