Direct Coombs Test (DAT)


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Subject: Direct Coombs Test (DAT)

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  • The DAT is used to detect IgG antibodies or complement components on the RBC surface membrane by incubating Coombs (rabbit anti-human globulin [AHG]) reagent with the patient's washed RBCs. The Coombs reagent can selectively bind IgG or C3 or be polyspecific and contain both anti-IgG and anti-C3.


  • DAT is useful in diagnosing autoimmune hemolysis [see p. 377]), hemolytic diseases of the newborn [see p. 380]), drug-induced hemolysis, and hemolytic transfusion reactions [see p. 757]).

  • The DAT is often used whenever hemolysis of red cells is suspected as being caused by autoantibodies. The assay determines if red cells have been coated in vivo with immunoglobulins, complement, or both.


Positive DAT

  • The DAT is positive whenever the patient's red cells are coated with autoantibodies that developed against the patient's own red cells.

  • Hemolytic disease of the newborn (erythroblastosis fetalis).

  • Warm autoimmune hemolytic anemias.

    • Idiopathic.

    • Evan syndrome (ITP and hemolytic anemia).

  • Cold autoimmune hemolytic anemia/cold agglutinin disease.

  • It is also positive when alloantibodies in a recipient's circulation react with antigens on recently transfused red cells, as well as alloantibodies in maternal circulation, which cross the placenta and coat fetal red cells.

    • Alloimmune (acute/delayed) hemolytic transfusion reactions.

    • Hemolytic disease of the newborn.

    • If the DAT is positive following recent transfusions, the antibodies can be eluted from RBCs and identified.

  • Drug-induced reactions, such as

    • Alpha-methyldopa

    • l-Dopa

    • High doses of penicillin

    • Quinidine

  • In patients who have not been transfused within the preceding 3 months, a positive DAT almost always reveals autoimmune antibodies.

Negative DAT

  • Hemolytic anemias caused by an intrinsic RBC defect (e.g., G6PD, hemoglobinopathies [see p. 363])


  • Finding of a positive DAT indicates the presence of red cell autoantibodies, alloantibodies following transfusions, or of coating of red cells with excess immunoglobulins. It requires additional workup to elucidate the etiology of the immunoglobulins by performing tests for antibody specificity: cold agglutinins (see p. 377 under hemolytic anemias), Donath-Landsteiner antibody (see p. 379), and also serum protein electrophoresis or immunofixation when a plasmacytic disease (see p. 432) is suspected. The administration of certain drugs (α-methyldopa, IV penicillin, or procainamide) and recent transfusions must also be excluded.

  • False-positive results may occur in plasma cell myeloma and lymphoplasmacytic lymphoma.

  • 1:10,000 normal blood donors have a positive DAT.

  • A negative DAT can rarely be seen in patients with autoimmune hemolytic anemia if only a small amount of IgG is bound to the RBC membrane.

  • A negative DAT does not rule out hemolysis. For instance, DAT is negative in some cases of drug-induced hemolytic anemias, hemoglobinopathies, hereditary spherocytosis, and other hereditary hemolytic anemias.