What Does a Positive Coombs Test Mean?
A positive Coombs test indicates that antibodies and/or complement proteins are bound to the surface of red blood cells, signaling immune-mediated red cell destruction or sensitization. 1, 2
Understanding the Two Types of Coombs Tests
Direct Antiglobulin Test (DAT) - Direct Coombs Test
The direct Coombs test detects immunoglobulin (IgG, IgA, IgM) and/or complement (C3) already attached to the patient's red blood cells. 1, 3 This test is performed directly on the patient's red cells without any additional incubation steps. 2
A positive direct Coombs test indicates:
- Autoimmune hemolytic anemia (AIHA) - the most common clinical implication, where the patient's immune system produces antibodies against their own red blood cells 4, 5
- Hemolytic disease of the fetus/newborn - due to maternal-fetal blood group incompatibility (Rh or ABO) 6, 2
- Drug-induced hemolytic anemia - certain medications trigger antibody formation against red cells 4, 5
- Transfusion-related hemolysis - antibodies reacting against transfused blood 1, 3
- Passenger lymphocyte syndrome - after stem cell or solid organ transplantation 1
Indirect Antiglobulin Test (IAT) - Indirect Coombs Test
The indirect Coombs test detects circulating antibodies in the patient's serum that are not yet bound to red cells but have the potential to cause hemolysis. 6 This test requires incubating the patient's serum with test red cells before adding antiglobulin reagent. 3
A positive indirect Coombs test indicates:
- Alloimmunization during pregnancy - maternal antibodies against fetal red cell antigens, particularly anti-Rh antibodies 6
- Pre-transfusion antibody screening - identifies patients at risk for transfusion reactions 6
- Antibody identification - determines which specific red cell antigens the patient has developed antibodies against 3
Clinical Interpretation Framework
When Direct Coombs is Positive
Evaluate for hemolysis by checking: 4, 7
- Haptoglobin (decreased in hemolysis)
- LDH (elevated in hemolysis)
- Indirect bilirubin (elevated in hemolysis)
- Reticulocyte count (elevated if bone marrow responding appropriately)
- Peripheral blood smear morphology
If hemolysis is confirmed, determine the pattern: 7, 5
- IgG positive alone - warm autoimmune hemolytic anemia (most common)
- C3 positive alone - cold agglutinin disease or paroxysmal cold hemoglobinuria
- IgG + C3 positive - mixed-type AIHA or severe warm AIHA
- Schistocytes present on smear - consider thrombotic microangiopathy (TMA), though classic TMA is typically Coombs-negative 4, 7
When Direct Coombs is Negative Despite Clinical Hemolysis
Approximately 2-10% of true autoimmune hemolytic anemia cases have a negative direct Coombs test. 5, 8 This occurs due to:
- IgG levels below detection threshold - sensitization too low for commercial reagents to detect 8
- Low-affinity IgG antibodies - removed during standard washing procedures 8
- IgA or monomeric IgM sensitization alone - not detected by standard anti-IgG/anti-C3 reagents 8
In suspected DAT-negative hemolytic anemia, request: 8
- More sensitive detection methods for low-level IgG
- Testing with anti-IgA and anti-IgM reagents
- Low ionic strength washing techniques to preserve low-affinity antibodies
Critical Pitfalls and False Positives
Technical False Positives
False positive results occur from: 1, 3
- Improper washing technique during test performance
- Incorrect centrifugation speed or duration
- Spontaneous red cell agglutination in the patient sample
- Specimen contamination or improper handling
Clinical False Positives (Positive Test Without Hemolysis)
A positive direct Coombs test without evidence of hemolysis can occur in: 9
- Markedly elevated serum IgG levels - hypergammaglobulinemia from chronic inflammation, infection, or autoimmune disease causes nonspecific antibody coating of red cells 9
- Recent intravenous immunoglobulin (IVIG) administration - exogenous antibodies transiently coat red cells 3
- Chronic lymphocytic leukemia or lymphoma - up to 35% have positive DAT without hemolysis 4, 3
When DAT is positive without hemolysis, investigate: 4, 9
- Serum immunoglobulin levels (IgG, IgA, IgM)
- Recent medication history, particularly immune checkpoint inhibitors
- Underlying lymphoproliferative disorders
- Recent IVIG or blood product administration
Special Clinical Scenarios
In Pregnancy
All pregnant women should have indirect Coombs testing (antibody screening) to detect alloimmunization. 6 If Rh antibody titres reach critical levels, serial monitoring with middle cerebral artery Doppler studies is indicated to assess for fetal anemia. 6
For newborns with jaundice: 6
- Direct Coombs test on cord blood is strongly recommended if mother is Rh-negative or has no prenatal typing
- Positive result indicates ABO or Rh incompatibility requiring close monitoring for hyperbilirubinemia
In Cancer Patients on Immunotherapy
Immune checkpoint inhibitors can cause immune-related hemolytic anemia with positive direct Coombs test. 4, 5 Management depends on severity:
- Grade 1-2 (Hb ≥8 g/dL): Continue therapy with close monitoring; initiate prednisone 0.5-1 mg/kg/day 4
- Grade 3 (Hb <8 g/dL with clinical consequences): Hold immunotherapy; initiate prednisone 1-2 mg/kg/day; hematology consultation 4
- Grade 4 (life-threatening): Permanently discontinue immunotherapy; methylprednisolone 1g IV daily × 3 days 4
In Anemia Workup
Coombs testing should be considered when evaluating anemia in patients with: 4
- Chronic lymphocytic leukemia or non-Hodgkin lymphoma
- History of autoimmune disease
- Unexplained anemia with elevated reticulocyte count
- Evidence of hemolysis (low haptoglobin, elevated LDH, elevated indirect bilirubin)
The reticulocyte index helps distinguish production versus destruction: 4
- Low reticulocyte index with positive Coombs suggests concurrent bone marrow suppression or nutritional deficiency
- High reticulocyte index with positive Coombs confirms appropriate marrow response to immune-mediated hemolysis