RBC surface in-vitro, the antihuman globulin will agglutinate the RBCs and the direct Coombs test will be positive. (A visual representation of a positive direct Coombs test is shown in the upper half of the schematic). Indirect Coombs test The indirect Coombs test, also referred to as the indirect antiglobulin test (IAT), 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. In antenatal care, the IAT 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. Uses Blood transfusion preparation 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. Donor blood for transfusion is also screened for infections in separate processes. Antibody screening 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 (i.e. blood types). Cross matching The indirect Coombs test is used to test a sample of the recipient's serum for antibodies against a sample of the blood donor's RBCs. This is sometimes called cross-matching blood. Antenatal antibody screening 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 haemolytic disease of the newborn. Laboratory method The IAT is a two-stage test. (A cross match is shown visually in the lower half of the schematic as an example of an indirect Coombs test). First stage Nonpatient, washed red blood cells (RBCs) with known antigens are incubated with patient serum containing unknown antibody content. If the serum contains antibodies to antigens on the RBC surface, the antibodies will bind to the surface of the RBCs. 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 (also known Coombs reagent) in this stage, and the indirect Coombs