The following are required:
1. Patient’s serum
2. Donor’s washed 3% red cell suspension
prepared as follows:
- Transfer 0.2–0.5 ml of red cells from the pilot tube of the donor blood into about 5 ml of physiological saline and mix.
- Centrifuge at high speed (e.g. 1 000 g) for about 2 minutes.
- Discard the supernatant fluid and resuspend the cells in a further 5 ml of saline. Mix, centrifuge, and discard the supernatant fluid.
- Prepare a 3% red cell suspension by adding 1 volume of packed cells to 30 volumes of saline.
3. Antiglobulin polyspecific (Broad-spectrum) reagent, usually colour coded green. AHG reagent will agglutinate red cells sensitized with antibodies and/or coated with detectable levels of complement components.
4. Anti-D serum to make AHG control cells.
5. AHG control IgG sensitized red cells, prepared as follows:
- Wash group O Rh positive red cells (obtained from a group O Rh positive person) three times in saline. Discard the final saline supernatant fluid.
- Add an equal volume of IgG anti-D to the packed red cells and mix.
- Incubate at 37 C for 30 minutes. Wash the cells four times in saline. Remove the final supernatant fluid.
- Suspend the packed cells in saline to make a 5% red cell suspension. When added to AHG reagent, the sensitized cells should show visible agglutination
- Store the sensitized cells at 2–8 C. They can be kept for 2–3 days.
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