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Recognizing hemolysis and its mechanism

15 min

  • Recognize hemolysis from haptoglobin, indirect bilirubin, LDH, and reticulocyte results
  • Tell immune, fragmentation, and intrinsic hemolysis apart using the DAT and film
  • Recognize when a normal G6PD result cannot exclude deficiency
  • Recognize a spherocyte in the monolayer of a blood film

Read the full reference

Try first

Try first

A hospitalized patient's hemoglobin has fallen over 2 days, and the lactate dehydrogenase (LDH) comes back markedly increased. Nothing else has been checked yet. Does the LDH result by itself establish hemolysis?

The next section explains it.

Right. The next section explains why.

The next section explains it.

The next section explains it.

Get the idea

The markers move together

No single result proves that red cells are dying faster than the marrow can replace them.1 Hemolysis raises the reticulocyte count once the marrow responds, raises indirect bilirubin and LDH as cells break down, and lowers haptoglobin as it binds the released hemoglobin.1 LDH also rises with liver disease, tissue injury, ineffective erythropoiesis, and a hemolyzed collection, so a raised LDH alone does not establish hemolysis. A low haptoglobin, a raised indirect bilirubin, and a rising reticulocyte count on a properly collected specimen, read together, support red-cell destruction.

Where the cells break down

PatternKey findings
ExtravascularReticulocytosis, indirect bilirubin and LDH increase, haptoglobin decrease, spherocytes in some disorders
IntravascularMarked haptoglobin depletion, plasma free hemoglobin, hemoglobinuria, schistocytes in fragmentation disorders

Immune, fragmentation, or inherited

The direct antiglobulin test (DAT) and the film sort the mechanism apart. Spherocytes form in warm autoimmune hemolysis and in hereditary spherocytosis, so a reactive DAT for IgG or C3d, read with objective hemolysis and the transfusion and drug history, supports an immune cause.2 A nonreactive DAT with a family pattern points toward membrane studies such as eosin-5'-maleimide (EMA) binding.4 Schistocytes point toward fragmentation.

A normal G6PD result during an episode

Reticulocytes carry more glucose-6-phosphate dehydrogenase (G6PD) activity than older red cells, and the most deficient cells are lost first during hemolysis, so an assay run during or soon after an episode can read normal in a deficient patient. Recent transfusion masks it the same way.3 State the limitation in the report, and repeat the quantitative assay at steady state, commonly 2 to 3 months later.3

References
  1. Keohane EM, Preston MM, Mirza KM, Walenga JM, eds. Rodak's Hematology: Clinical Principles and Applications. 7th ed. Elsevier; 2025. Accessed September 26, 2026. https://www.us.elsevierhealth.com/rodaks-hematology-9780323936507.html
  2. Michel M, Crickx E, Fattizzo B, Barcellini W. Autoimmune haemolytic anaemias. Nat Rev Dis Primers. 2024;10(1):82. doi:10.1038/s41572-024-00566-2
  3. Luzzatto L, Ally M, Notaro R. Glucose-6-phosphate dehydrogenase deficiency. Blood. 2020;136(11):1225-1240. doi:10.1182/blood.2019000944
  4. Wu Y, Liao L, Lin F. The diagnostic protocol for hereditary spherocytosis: 2021 update. J Clin Lab Anal. 2021;35(12):e24034. doi:10.1002/jcla.24034

Watch one

A hemoglobin of 8.9 g/dL is falling further over 2 days. The reticulocyte count is increased, indirect bilirubin and LDH are increased, and haptoglobin is undetectable. The film shows spherocytes in the monolayer and no schistocytes. The DAT is positive for IgG. The patient has no personal or family history of anemia and no recent transfusion. What mechanism does the pattern support?

  1. Check the markers together: reticulocytes, bilirubin, and LDH are up, and haptoglobin is undetectable. Hemolysis is present.

    Establishing that hemolysis is present comes before asking what kind.

  2. Read the film: spherocytes are present and no schistocytes are seen, which argues against a fragmentation process.

    Shape narrows the mechanism before any serologic test is read.

  3. Check the DAT: positive for IgG, which supports an immune cause when read with objective hemolysis.

    Spherocytes alone cannot separate an immune cause from a hereditary one.

  4. Check the history: no personal or family history of anemia and no recent transfusion, so nothing points away from an acquired immune cause.

    A family pattern or long-standing mild anemia would point toward hereditary spherocytosis.

The pattern supports immune-mediated hemolysis: spherocytes with a positive DAT for IgG, no schistocytes, and no family or transfusion history to suggest another cause.

Your turn

Problem 1 of 3

Results show decreased haptoglobin, hemoglobinemia, and hemoglobinuria. Which clearance route does this pattern reflect?

Correct. Red cells rupturing in the circulation release hemoglobin into plasma. Once haptoglobin and hemopexin are saturated, free hemoglobin appears in plasma and urine, and hemosiderinuria can follow with ongoing loss.

Incorrect. Haptoglobin can fall in either route, but macrophage clearance releases no free hemoglobin into plasma and urine. Extravascular hemolysis raises indirect bilirubin and lactate dehydrogenase (LDH) with reticulocytosis.

Incorrect. Pitting removes inclusions such as Howell-Jolly bodies and returns the red cell to the circulation, so it releases no hemoglobin.

Hint
  1. Haptoglobin binds free hemoglobin released directly into the plasma.
  2. Macrophage clearance in the spleen and liver does not put hemoglobin into the plasma or urine.

Review Erythrocyte clearance and iron recycling

Problem 2 of 3

A film shows numerous schistocytes with thrombocytopenia, and lactate dehydrogenase (LDH) is markedly elevated; PT and activated partial thromboplastin time (APTT) are near normal. What is the laboratory's action?

Correct. Schistocytes, thrombocytopenia, and a high LDH with relatively preserved clotting times suggest thrombotic thrombocytopenic purpura (TTP) or another thrombotic microangiopathy. Urgent notification comes first, and an ADAMTS13 specimen is collected before treatment when feasible. Activity below 10% supports TTP.

Incorrect. In TTP the PT and APTT are often less disturbed than in overt disseminated intravascular coagulation (DIC), so near-normal clotting times fit the urgent pattern.

Incorrect. Acute DIC shows prolonged clotting times, low fibrinogen, and a positive D-dimer. Schistocytes and thrombocytopenia with near-normal clotting times fit a thrombotic microangiopathy.

Hint
  1. Look at the platelet count and the clotting times together with the film.
  2. A thrombotic microangiopathy needs urgent communication under the laboratory's procedure.

Review Urgent laboratory patterns

Problem 3 of 3

Near the feather edge of a film, several red cells appear dense with no central pallor. In the monolayer, red cells show normal central pallor. What do you report?

Outside the monolayer, overlapping or flattened red cells can lose their central pallor and look dense. Spherocytes are judged where cells lie singly.

Called cells outside the monolayer spherocytes

Outside the monolayer, overlapping or flattened red cells can lose their central pallor and look dense. Calling them spherocytes suggests hereditary spherocytosis or immune hemolysis the specimen does not show. Spherocytes are judged where cells lie singly and normal cells keep their pallor.

In the monolayer, where cells lie singly, the red cells keep their normal central pallor. The dense appearance at the edge is a preparation effect and does not represent true spherocytes.

Central pallor is judged where cells lie singly and are not overlapping or flattened. The monolayer is the correct place to make this call.

Review Red-cell morphology on the stained film

Use it

  • A woman, MRN 0271946, is admitted with fatigue and yellowing of the eyes.
  • Her hemoglobin has fallen since admission 2 days ago.
  • She has no history of transfusion, recent travel, or new medications.
  • The film shows spherocytes in the monolayer and no schistocytes.
  • The blood bank reports the DAT positive for IgG.
  • A quantitative G6PD assay drawn this admission is within the reference interval.
TestResultPreviousReference intervalFlag
Hemoglobin8.9 g/dL11.4 g/dLAdmission12.0–15.5 g/dLLow
Reticulocytes (absolute)185 × 10³/µL62 × 10³/µLAdmission25–100 × 10³/µLHigh
Indirect bilirubin3.8 mg/dL0.6 mg/dLAdmission0.1–1.0 mg/dLHigh
Lactate dehydrogenase410 U/L160 U/LAdmission100–220 U/LHigh
HaptoglobinUndetectable90 mg/dLAdmission30–200 mg/dLLow

Specimen: H 3, L 8, I 2. Serum and EDTA whole blood, drawn on admission day 2

Decision 1 of 3

Does this panel show hemolysis, or a production problem alone?

Haptoglobin is undetectable, indirect bilirubin and LDH are increased, and the reticulocyte count has risen, all read together. This is the pattern of accelerated red-cell destruction with a marrow response.

A low reticulocyte count would suggest reduced production. A rising reticulocyte count here is the marrow's response to destruction.

The reticulocyte count, LDH, indirect bilirubin, and haptoglobin already agree on hemolysis with a marrow response. A biopsy is not needed to make this call.

Review Evidence of increased hemoglobin catabolism

Decision 2 of 3

Which mechanism does the pattern support?

Fragmentation hemolysis shows schistocytes. None are seen here, and the film shows spherocytes.

Spherocytes with a positive DAT for IgG, read with objective hemolysis and no family or transfusion history, support an antibody-mediated cause cleared by the spleen.

The film and DAT already point to an immune cause. A single G6PD result drawn during this episode cannot rule enzyme deficiency in or out either way.

Review Extrinsic and immune hemolysis

Decision 3 of 3

What do you do with the normal G6PD result drawn during this admission?

Acute hemolysis removes the most deficient cells and leaves younger, higher-activity cells behind, so a deficient patient can test normal during an episode. This result cannot exclude deficiency.

Excluded G6PD deficiency from one result during recovery

Reticulocytes carry more G6PD than older cells, and the most deficient cells are lost during hemolysis, so activity measured during or soon after an episode can look normal. Recent transfusion masks it the same way. State the limitation in the report, and repeat the quantitative assay at steady state, commonly 2 to 3 months later.

The result was drawn during active hemolysis, when activity can look falsely normal. The report states that limitation, and a repeat 2 to 3 months later at steady state would be needed if deficiency remains a concern.

A same-day repeat still falls during the same hemolytic episode and would carry the same limitation. The steady-state interval is what removes it.

Review G6PD deficiency testing

The clue that settles this case is the combination on the film and the DAT: spherocytes with no schistocytes, a DAT positive for IgG, and no family or transfusion history. That combination points to immune-mediated hemolysis, and it means the admission's G6PD result, drawn during the episode, cannot be read as excluding deficiency.

Keep

Sources checked