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Coombs test

Also known as an antiglobulin test

Refers to either of two clinical blood tests used in immunohematology and immunology.

The two Coombs tests are the direct Coombs test (DCT, also known as 2242in test or IAT)

The direct Coombs test is used to test for autoimmune hemolytic anemia, whereby the immune system lysis or breaks down RBC membranes causing RBC destruction.

In auto immune diseases a patients blood may contain IgG antibodies that can bind to antigens on the RBC surface membrane, and their circulating RBCs can become coated with IgG alloantibodies and/or IgG autoantibodies.

Complement may subsequently bind to the bound antibodies and cause RBC destruction.

The direct Coombs test is used to detect these antibodies or complement proteins that are bound to the surface of red blood cells.

A blood sample’s RBCs are washed and then incubated with anti-human globulin (Coombs reagent),

If agglutination of RBCs occurs, the direct Coombs test is positive

This agglutination is a visual indication that antibodies are bound to the surface of red blood cells.

The indirect Coombs test is used in prenatal testing of pregnant women and in testing blood prior to a blood transfusion.

The indirect Coombs test detects antibodies against RBCs that are present unbound in the patient’s serum.

In the indirect Coombs test, serum is extracted from the blood sample taken from the patient, and is incubated with RBCs of known antigenicity, then anti-human globulin is added.

If agglutination occurs, the indirect Coombs test is positive.

The Coombs tests are based on the fact that anti-human antibodies, which produced by immunizing non-human species with human serum, will bind to human antibodies, commonly IgG or IgM.

Uses of the Coombs test include: cross matching for blood for transfusion, atypical antibodies in the blood plasma of pregnant women as part of antenatal care, and detection of antibodies for the diagnosis of immune-mediated hemolytic anemias.

The direct Coombs test is used to detect if antibodies or complement system factors have bound to RBCs surface antigens in vivo.

The direct Coombs test is used clinically when immune-mediated hemolytic anemia is suspected.

A positive Coombs test indicates that an immune mechanism is attacking the patient’s own RBCs, through autoimmunity, alloimmunity or a drug-induced immune-mediated mechanism.

Alloimmune hemolysis occurs with hemolytic disease of the newborn, Rh D hemolytic disease of the newborn, ABO hemolytic disease of the newborn. Anti-Kell hemolytic disease of the newborn. Rh c hemolytic disease of the newborn Rh E hemolytic disease of the newborn and other blood group incompatibility, and alloimmune hemolytic transfusion reactions.

Autoimmune hemolysis/immunohemolytic hemolysis occur with:

Warm antibody autoimmune hemolytic anemia

Idiopathic autoimmune hemolysis.

Systemic lupus erythematosus.

Evans’ syndrome (antiplatelet antibodies and hemolytic antibodies).

Cold antibody immunohemolytic anemia.

Idiopathic cold hemagglutinin syndrome.

Waldenström’s macroglobulinemia

Infectious mononucleosis

Paroxysmal cold hemoglobinuria

Drug-induced immune-mediated hemolysis

Methyldopa (IgG mediated type II hypersensitivity)

Penicillin at high dose

Quinidine

During the process the patient’s RBCs are washed to remove the patient’s own serum, and then centrifuged with antihuman globulin (Coombs reagent).

If immunoglobulin or complement factors have been fixed on to the RBC surface in-vivo, the antihuman globulin will agglutinate the RBCs and the direct Coombs test will be positive.

The indirect Coombs test is used to detect in-vitro antibody-antigen reactions.

It is used to detect very low concentrations of antibodies present in a patient’s plasma/serum prior to a blood transfusion.

It is used to screen pregnant women for antibodies that may cause hemolytic disease of the newborn.

The IAT can also be used for compatibility testing, antibody identification, RBC phenotyping, and titration studies.

The indirect Coombs test is used to screen for antibodies in the preparation of blood for blood transfusion.

The donor’s and recipient’s blood must be ABO and Rh D compatible.

A blood sample from the recipient and a blood sample from every unit of donor blood are screened for antibodies with the indirect Coombs test.

Each sample is incubated against a wide range of RBCs that together exhibit a full range of surface antigens.

The indirect Coombs test, called cross-matching is used to test a sample of the recipient’s serum for antibodies against a sample of the blood donor’s RBCs.

The indirect Coombs test is used to screen pregnant women for IgG antibodies that are likely to pass through the placenta into the fetal blood and cause hemolytic disease of the newborn.

Washed test red blood cells are incubated with a known human serum.

If the serum contains antibodies to antigens on the RBC surface, the antibodies will bind onto the surface of the RBCs.

In a second stage the RBCs are washed three or four times with isotonic saline solution and then incubated with antihuman globulin.

If antibodies have bound to RBC surface antigens in the first stage, RBCs will agglutinate when incubated with the antihuman globulin in this stage, and the indirect Coombs test will be positive.

Coombs reagent, known as Coombs antiglobulin or antihuman globulin, is used in both the direct Coombs test and the indirect Coombs test.

Coombs reagent is antihuman globulin. made by injecting human globulin into animals, which produce polyclonal antibodies specific for human immunoglobulins and human complement system factors.

Historically, the test was done in test tubes, but today it is commonly done using microarray and gel technology.

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