Coomb's test
[Ref: KB2:p186; PK1:p252-253]
Coomb's reagent
- Coomb's reagent (aka antiglobulin serum)
* Contains polyspecific rabbit or sheep IgG antibodies which are directed against human IgG antibodies (i.e. anti-human-IgG IgG antibodies)
- Anti-human globulin (AHG)
* Produced in animals after injection of human globulin, complement, or specific immunoglobulin
* May contain polyspecific antibodies or monospecific
* ??Coomb's reagent is a subtype of AHG
Direct Coomb's test
- Aim
= To detect antibody or complement on the RBC that have been sensitized in the patient's body
- Patient's RBC are washed first
- Then AHG is added to the washed cells
- If agglutination occurs
--> Positive direct Coomb's test
--> Patient has been sensitised (in vivo) to some antigen on the RBC (i.e. producing antibodies against the RBC cells)
Direct Coomb's test is positive in
- Haemoytic disease (Rh) of the newborn
- Autoimmune haemolytic anaemia
- Drug-induced immune haemolysis
- Haemolytic transfusion reactions
Indirect Coomb's test
- A two stage test
* Incubation, then add reagent
* Direct Coomb's does not require incubation
- Aim
= To detect the presence of any IgG antibodies (in recipient's plasma) directed against donor's RBC membrane antigen
- These (incomplete) antibodies are not able to cause agglutination by themselves during other testing procedures but may cause problem when transfused
Step 1: Incubation
Donor's RBCs are incubated in recipient's serum at 37C
--> Allow any IgG antibodies (from the recipient) to bind to the corresponding antigen on donor RBC
* i.e. First stage
Step 2: Washing
Removes all serum and unbound IgG in the serum
--> IgG antibodies bound to donor RBC remains
Step 3: Testing with Coomb's reagent
Coomb's reagent is added to the washed RBC
If there are IgG antibodies (from recipient's serum) bound to donor RBC
--> Anti-human-IgG IgG antibodies will bind to the IgG on two RBCs
--> Join them together
--> Agglutination
--> Positive Coomb's test
If no IgG are bound to donor RBC
--> No agglutination
--> Negative Coomb's test
Step 4: Coomb's control test
Coomb's control cells
* RBC known to be coated with IgG
Coomb's control cells are added to the negative Coomb's test
--> Will definitely cause agglutination if the reagent was active
If no agglutination when Coomb's control cells are added
--> Reagent was inactive
Indirect Coomb's test is positive when
- Recipient has antibody in serum against donor RBC (during crossmatching)
- Atypical blood group antibodies (during antibody screening)
- Blood group antibodies in pregnant women
- Serum antibodies in autoimmune haemolytic anaemia