Agglutination Tests in Diagnosis
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Direct answer
Agglutination tests detect antigen or antibody by visible clumping of particles such as bacteria, latex beads or red cells. Direct and slide agglutination identify organisms, while passive and latex methods coat carrier particles to detect soluble antigen or antibody. The Coombs tests detect antibody bound to red cells or free in serum, and the Widal test is the classic tube agglutination for enteric fever.
What you must remember
- Direct agglutination: a particulate antigen clumps visibly with specific antibody, used in slide tests for serotyping bacteria and grouping blood.
- Widal test: tube agglutination using Salmonella Typhi O and H antigens, where a single titre is weak evidence and paired sera are more useful.
- Passive agglutination: a soluble antigen is adsorbed on latex or red cells to detect antibody, as in latex tests for rheumatoid factor and ASO.
- Reverse passive agglutination: antibody on latex detects antigen, as in cryptococcal antigen testing in CSF.
- Coombs direct test: detects antibody or complement on the patient's own red cells, positive in autoimmune haemolysis and haemolytic disease of the newborn.
- Coombs indirect test: detects free antibody in serum, used for antibody screening and crossmatching before transfusion.
- Haemagglutination inhibition: antibody blocks virus-induced agglutination of red cells, used in influenza serology.
- Paul-Bunnell test: heterophile antibodies agglutinate sheep red cells in infectious mononucleosis, with lower sensitivity in young children.
Matching each test to its principle and use
Agglutination tests answer different questions, so the choice depends on whether the laboratory is trying to identify an organism, detect antibody in serum or examine antibody on cells. The table summarises the six formats most often tested.
| Test | Principle | Typical use |
|---|---|---|
| Slide agglutination | Particulate antigen clumps with antibody on a glass slide | Serotyping and ABO grouping |
| Widal (tube) | Serial dilutions of serum against Salmonella O and H antigens | Enteric fever, read with caution |
| Latex agglutination | Antigen or antibody coated on latex particles | RF, ASO, cryptococcal antigen |
| Coombs direct (DAT) | Anti-human globulin detects antibody on the patient's red cells | Autoimmune haemolysis, HDN |
| Coombs indirect (IAT) | Anti-human globulin detects free antibody in serum | Antibody screen and crossmatch |
| Haemagglutination inhibition | Antibody prevents virus from agglutinating red cells | Influenza serology on paired sera |
The Widal test is the most often misread of the group, because vaccination and endemic exposure can raise titres in people who do not have active disease. The Coombs tests look similar but test different compartments, one on the cells and the other in the serum.
Where the exam tries to catch you
Agglutination questions usually test which compartment or which antigen the test measures, so the wording of the question decides the answer.
- Trap: calling a single Widal titre diagnostic of typhoid Correct: a single raised titre is weak evidence because vaccination and past infection cause false positives, so a fourfold rise between paired sera is more informative.
- Trap: using the direct Coombs test for crossmatching Correct: crossmatching uses the indirect test with donor cells and recipient serum, whereas the direct test examines the patient's own red cells.
- Trap: taking a positive latex RF test as proof of rheumatoid arthritis Correct: RF is positive in many other conditions and in some healthy people, so anti-CCP is the more specific marker.
Frequently asked questions
Why is the direct Coombs test positive in haemolytic disease of the newborn?
The direct Coombs test is positive in haemolytic disease of the newborn because maternal IgG, such as anti-D, has crossed the placenta and coats the infant's red cells. The anti-human globulin reagent detects this coating, which confirms antibody-mediated haemolysis.
What does the Widal test detect in enteric fever?
The Widal test detects agglutinating antibodies against Salmonella Typhi O and H antigens in the patient's serum. Because vaccination and past infection can raise the titres, a single result is weak evidence, and a rising titre in paired sera is more useful.
Which agglutination test detects cryptococcal antigen in cerebrospinal fluid?
Latex agglutination for cryptococcal antigen detects the capsular polysaccharide of Cryptococcus neoformans in cerebrospinal fluid and serum. Antibody-coated latex particles clump when the antigen is present, which makes it a rapid test in suspected cryptococcal meningitis.
What is the Paul-Bunnell test used to diagnose?
The Paul-Bunnell test is used to diagnose infectious mononucleosis, which is caused by Epstein-Barr virus. Heterophile antibodies in the patient's serum agglutinate sheep red cells, and the test is less sensitive in young children than in older patients.