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Direct Antiglobulin Test (DAT)

The DAT is a serologic procedure that is used to detect in vivo binding of IgG antibody and/or complement on the red cells (in vivo sensitization of RBCs). It is used primarily for the detection and differential diagnosis of various forms of immune hemolysis including autoimmune hemolytic anemia, hemolytic transfusion reactions, drug-induced hemolysis, and hemolytic disease of the newborn (HDN).
The procedure involves washing the patient's red cells to remove residual plasma proteins and then testing the washed cells with antiglobulin reagent. Depending upon the antiglobulin reagent used, a positive test means that either IgG and/or complement components were bound to the red cells in vivo, or inside of the body.
Initial testing is often done using polyspecific antihuman globulin (AHG) that contains both anti-IgG and at least anti-C3d (In some preparations, anti-C3b and other anti-complement activity may also be present).
Once reactivity is noted with polyspecific AHG, monospecific reagents may be used to differentiate between IgG and complement. This extended testing is useful in suspected cases of paroxysmal cold hemoglobinuria, cold hemagglutinin disease, warm autoimmune hemolytic anemia, and drug-induced hemolytic anemia. However, the use of anti-C3 would not be required for diagnosing HDN as a positive DAT would always be the result of the production of IgG antibody and would not involve complement.