Astrovirus ELISA

The Astrovirus ELISA uses specific antibodies in a sandwich-type method. Monoclonal antibodies to all known astrovirus serotypes are coated to the well surface of the microwell plate. A pipette is used to place a suspension of the stool sample to be examined as well as control samples into the well of the microwell plate together with biotinylated monoclonal anti-astrovirus antibodies (Conjugate 1) for incubation at room temperature (20 25 °C). After a wash step, streptavidin poly-peroxidase conjugate (Conjugate 2) is added and that is incubated again at room temperature (20–25 °C). With the presence of astroviruses in a stool sample, a sandwich complex will form which consists of immobilized antibodies, the astrovirus antigens, and the antibodies conjugated with the biotin-streptavidin-peroxidase complex. Another wash step removes the unattached streptavidin poly-peroxidase conjugate. After adding the substrate, the attached enzyme changes the color of the previously colorless solution in the wells of the microwell plate to blue if the test is positive. Addition of a stop reagent changes the color from blue to yellow. The extinction is proportional to the concentration of astroviruses found in the sample.