DAT is mainly done on blood samples of newborns, especially when a newborn is suffering from jaundice.
1. Place the sample of RBCs to be tested in a 5 % suspension of isotonic saline.
2. Using a clean pipette, add 1 drop of the RBCs suspension prepared into a small test tube.
3. Wash 3 times using normal saline in order to remove all serum traces. After last washing step, decant completely.
4. Then add 2 drops of antihuman serum and mix thoroughly. …show more content…
These antibodies, if present in the serum, will agglutinate bacterial suspension which carries homologous antigens.
If agglutination occurs with O antigen then it is considered positive for Salmonella typhi. If agglutination occurs in A or B antigen then it is confirmed as positive for Salmonella paratyphi. Agglutination will occur in H antigen for all the cases of antigens like O, A, and B.
This Widal test can be performed either by slide agglutination test or by tube agglutination test. The test done was that of the slide agglutination qualitative test, which procedure is detailed below.
1. Place all reagents needed at room temperature and mix thoroughly before use.
2. On the test card, put 1 drop of test sample in each reaction circle containing O, H, AH, and BH.
3. Add 1 drop of antigen solutions of O and H for Salmonella typhi, AH and BH for Salmonella paratyphi, respectively in their labeled reaction circles on the test card.
4. Using a stick, mix well and rotate gently.
5. Observe any formation of agglutination.
6. Record the grade of agglutination, if any visible.