Haemolytic Anaemias and Haemoglobinopathies

Sickle Cell Disease and Thalassaemias

Sickle Cell Disease

Sickle cell disease (SCD) is an autosomal recessive haemoglobinopathy caused by a missense mutation in the β-globin gene (HBB) on chromosome 11: glutamic acid is replaced by valine at position 6, producing HbS. The nonpolar valine creates a hydrophobic “sticky patch,” so deoxygenated HbS polymerises, distorting red cells into rigid sickle shapes. Polymerisation is promoted by low O₂ tension, acidosis, dehydration, infection, and increased 2,3-BPG. Sickled cells cause both chronic extravascular haemolysis and microvascular occlusion.

Genotype	Key electrophoresis findings	Clinical significance
HbSS	HbS >85–90%, HbF 2–20%, no HbA	Classic severe SCD
HbAS	HbA >50%, HbS <45%	Sickle trait; usually asymptomatic, renal papillary necrosis possible
HbSC	HbS and HbC, no HbA	Often milder haemolysis, more retinopathy
HbS/β0-thalassaemia	HbS, no HbA	Severe, resembles HbSS
HbS/β+-thalassaemia	HbS with reduced HbA	Variable severity

At birth, symptoms are absent because HbF (α₂γ₂) inhibits HbS polymerisation. Disease appears after approximately 6 months, when γ-globin is replaced by β-globin. Laboratory findings reflect haemolysis: normocytic anaemia, reticulocytosis, increased unconjugated bilirubin, increased LDH, decreased haptoglobin, and sickled cells with Howell–Jolly bodies due to functional asplenia.

Major complications are high-yield because they follow directly from vaso-occlusion and splenic dysfunction. Dactylitis is often the first presentation in infants. Recurrent splenic infarction causes autosplenectomy, predisposing to encapsulated organisms: Streptococcus pneumoniae, Haemophilus influenzae type b, and Neisseria meningitidis. Salmonella osteomyelitis is classically associated with SCD, whereas Staphylococcus aureus remains the most common cause overall. Acute chest syndrome presents with fever, chest pain, pulmonary infiltrate, and hypoxaemia; triggers include infection, fat emboli from marrow infarction, and pulmonary vaso-occlusion. Other complications include stroke, avascular necrosis of the femoral head, leg ulcers, priapism, pigment gallstones, and renal papillary necrosis causing painless haematuria and inability to concentrate urine.

Hydroxyurea is the key pharmacologic therapy tested on Step 1. It inhibits ribonucleotide reductase and increases HbF, reducing HbS polymerisation. Typical adult starting dose is 15 mg/kg/day orally, titrated every 8–12 weeks to a maximum around 35 mg/kg/day while monitoring neutrophils and platelets. The landmark Multicenter Study of Hydroxyurea (MSH, 1995) showed approximately a 44% reduction in painful crises and fewer acute chest episodes and transfusions. Supportive principles include folate supplementation, vaccination, penicillin prophylaxis in childhood, hydration, analgesia, and transfusion for selected severe complications; curative therapy is haematopoietic stem-cell transplantation, but this is not the Step 1 emphasis.

Thalassaemias

Thalassaemias are inherited disorders of decreased globin-chain synthesis, causing imbalanced haemoglobin production, ineffective erythropoiesis, and microcytic hypochromic anaemia. Unlike SCD, the haemoglobin structure is usually normal; the problem is quantity, not quality. Iron studies are typically normal or increased, distinguishing thalassaemia from iron deficiency.

α-Thalassaemia

α-globin is encoded by four genes, two on each chromosome 16. α-thalassaemia usually results from gene deletions. Severity depends on the number of deleted α-globin genes.

Deleted α genes	Name	Findings
1	Silent carrier	Usually normal CBC
2	α-thalassaemia trait	Mild microcytosis, mild or no anaemia
3	HbH disease	β4 tetramers; chronic haemolytic anaemia, splenomegaly
4	Hydrops fetalis	Hb Bart’s γ4; severe fetal hypoxia, oedema, usually fatal in utero

HbH and Hb Bart’s have high oxygen affinity, so they hold oxygen tightly and deliver it poorly to tissues. Electrophoresis may be normal in α-thalassaemia trait because HbA, HbA₂, and HbF proportions can remain near normal; genetic testing is more definitive.

β-Thalassaemia

β-globin is encoded by two genes on chromosome 11. β-thalassaemia usually results from point mutations affecting promoters, splice sites, or nonsense codons. Mutations are classified as β⁺ when β-chain production is reduced and β⁰ when absent. Reduced β chains lead to excess α chains, which precipitate in erythroid precursors, causing membrane damage, ineffective erythropoiesis, marrow expansion, and extramedullary haematopoiesis.

Disorder	Genotype/physiology	Clinical and laboratory findings
β-thalassaemia minor	Heterozygous β mutation	Mild microcytic anaemia, target cells, increased RBC count; HbA2 >3.5%
β-thalassaemia intermedia	Variable β production	Moderate anaemia; variable transfusion need
β-thalassaemia major	Severe β0/β0 or β0/β+	Presents after 6 months; severe anaemia, growth failure, hepatosplenomegaly

Classic β-thalassaemia major findings include crew-cut skull from marrow expansion, chipmunk facies, extramedullary haematopoiesis, and iron overload from transfusions and increased intestinal absorption. Electrophoresis shows decreased or absent HbA, increased HbF, and increased HbA₂. The Mentzer index, MCV/RBC count, is often <13 in thalassaemia and >13 in iron deficiency anaemia. Management concepts include transfusion support and iron chelation with deferoxamine, deferasirox, or deferiprone; for Step 1, focus on genetics, electrophoresis, and the mechanism of ineffective erythropoiesis.

Hereditary Spherocytosis

Hereditary spherocytosis is an inherited haemolytic anaemia caused by defects in red blood cell membrane cytoskeletal proteins, producing spherical, less deformable erythrocytes that are trapped and destroyed in the spleen. It is the most common inherited haemolytic anaemia in individuals of Northern European ancestry, with an estimated prevalence of approximately 1 in 2,000. Most cases are autosomal dominant, although autosomal recessive and de novo forms occur.

Pathophysiology

Normal erythrocytes are biconcave discs with a high surface-area-to-volume ratio, allowing them to deform through splenic sinusoids. This shape depends on vertical interactions between the lipid bilayer and the underlying spectrin-actin cytoskeleton. In hereditary spherocytosis, mutations disrupt these vertical linkages, causing loss of membrane fragments as microvesicles. The cytoplasmic volume is relatively preserved, but surface area falls, producing a spherocyte: a dense, round red cell lacking central pallor.

Protein affected	Role	Association
Ankyrin	Links spectrin to band 3	Most common defect
Spectrin	Main cytoskeletal scaffold	Can cause severe disease if markedly deficient
Band 3	Anion exchanger; membrane anchor	Autosomal dominant forms
Protein 4.2	Stabilises band 3-ankyrin interaction	Less common; may be recessive

Spherocytes are poorly deformable and are selectively retained in splenic cords. Splenic macrophages remove membrane further and ultimately phagocytose the cell, producing extravascular haemolysis. Therefore, laboratory findings reflect macrophage-mediated red cell destruction: increased unconjugated bilirubin, increased lactate dehydrogenase, low or normal haptoglobin, and reticulocytosis if marrow compensation is intact.

Clinical Features

The classic triad is anaemia, jaundice, and splenomegaly, although severity varies widely. Patients may present in infancy with jaundice or later with fatigue, pallor, episodic scleral icterus, or incidental anaemia. Chronic bilirubin overproduction predisposes to pigment gallstones, especially in adolescence and adulthood. Splenomegaly is common because the spleen is the major site of red cell destruction.

High-yield complications include aplastic crisis, classically triggered by parvovirus B19, which infects erythroid precursors via the P antigen and transiently halts erythropoiesis. In hereditary spherocytosis, red cells have shortened survival, so a brief marrow shutdown can cause abrupt severe anaemia with a low reticulocyte count. This contrasts with baseline hereditary spherocytosis, where reticulocytes are elevated.

Severity Classification

Severity	Typical haemoglobin	Reticulocytes	Bilirubin	Clinical pattern
Mild	11–15 g/dL	3–6%	1–2 mg/dL	Often compensated; may be asymptomatic
Moderate	8–12 g/dL	>6%	>2 mg/dL	Jaundice, splenomegaly, gallstones
Severe	6–8 g/dL	Often >10%	Elevated	Transfusion may be needed; growth delay possible

Laboratory Diagnosis

The peripheral smear shows spherocytes: small, hyperchromic-appearing red cells without central pallor. Because spherocytes contain relatively more haemoglobin per unit volume, the mean corpuscular haemoglobin concentration is often increased; a value >36 g/dL is suggestive, whereas the normal range is approximately 32–36 g/dL. Mean corpuscular volume may be normal or slightly low, and red cell distribution width is often increased.

• Reticulocyte count: elevated at baseline due to marrow compensation; normal adult reticulocytes are approximately 0.5–2.5%.
• Direct antiglobulin test: negative, helping distinguish hereditary spherocytosis from autoimmune haemolytic anaemia, which can also produce spherocytes.
• Osmotic fragility test: increased fragility in hypotonic saline because spherocytes have reduced surface-area-to-volume ratio and lyse earlier than normal biconcave cells.
• Eosin-5-maleimide binding test: flow cytometry assay assessing band 3-associated membrane proteins; commonly used screening test with reported sensitivity and specificity often around 90–95% in appropriate clinical settings.

The osmotic fragility principle is a useful Step 1 concept: when extracellular fluid becomes hypotonic, water enters red cells. Normal biconcave cells can swell before rupturing, but spherocytes have little excess membrane surface area, so they lyse at less hypotonic conditions.

Management Principles and Step 1 Associations

Treatment depends on severity and is aimed at preventing complications rather than correcting the membrane defect. Folic acid is often given in moderate to severe chronic haemolysis because increased erythropoiesis raises folate demand; common dosing is 1 mg orally daily. Red cell transfusion is reserved for severe symptomatic anaemia or aplastic crisis.

Splenectomy decreases extravascular haemolysis by removing the major site of red cell destruction. It improves anaemia and reticulocytosis but does not correct the membrane defect, so spherocytes persist on smear. Because splenectomy increases lifetime risk of sepsis from encapsulated organisms, it is generally avoided before age 5–6 years unless disease is severe. Patients should receive vaccines against Streptococcus pneumoniae, Haemophilus influenzae type b, and Neisseria meningitidis, ideally at least 2 weeks before elective splenectomy. Postsplenectomy smears may show Howell-Jolly bodies, reflecting impaired splenic clearance of nuclear remnants.

The key differential diagnosis is warm autoimmune haemolytic anaemia: both can show spherocytes and extravascular haemolysis, but autoimmune disease has a positive direct Coombs test, whereas hereditary spherocytosis is Coombs-negative and often has a family history.

G6PD Deficiency

Core biochemistry and pathogenesis

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is an X-linked recessive enzymopathy causing episodic haemolytic anaemia under oxidative stress. G6PD catalyses the first, rate-limiting step of the hexose monophosphate shunt: glucose-6-phosphate → 6-phosphogluconolactone, producing NADPH. In erythrocytes, NADPH is essential to maintain glutathione in its reduced form (GSH) via glutathione reductase. Reduced glutathione detoxifies hydrogen peroxide and other reactive oxygen species via glutathione peroxidase.

Red blood cells are uniquely vulnerable because they lack mitochondria and nuclei, cannot synthesize new proteins, and rely heavily on G6PD-derived NADPH for antioxidant defence. When oxidant stress exceeds capacity, haemoglobin sulfhydryl groups oxidize and denature, forming Heinz bodies. Splenic macrophages remove Heinz body–containing membrane portions, producing bite cells and blister cells. Haemolysis is predominantly extravascular in the spleen, but severe oxidative injury can also cause intravascular haemolysis with haemoglobinaemia and haemoglobinuria.

Genetics, epidemiology, and evolutionary association

The G6PD gene is located on Xq28. Hemizygous males are classically affected; heterozygous females may be symptomatic because of lyonization with skewed X-inactivation. The disorder is common in populations from Africa, the Mediterranean, the Middle East, and parts of Asia. This geographic distribution reflects partial protection against severe Plasmodium falciparum malaria, an important Step 1 parallel to sickle cell trait and thalassaemias.

Variant/class	Residual enzyme activity	Typical clinical pattern	Classic association
WHO class I	Usually <10%	Chronic nonspherocytic haemolytic anaemia	Rare severe variants
WHO class II	<10%	Intermittent severe haemolysis with oxidant exposure	Mediterranean variant; favism common
WHO class III	10–60%	Usually episodic, self-limited haemolysis	African A− variant
WHO class IV	60–150%	Usually asymptomatic	No significant haemolysis
WHO class V	>150%	No haemolysis	Increased activity; clinically rare

Triggers and pharmacology-relevant associations

Haemolytic episodes typically occur 24–72 hours after oxidative exposure. Common triggers include infections, fava beans, and oxidant drugs. Infection is often the most common precipitant because activated neutrophils and macrophages generate reactive oxygen species.

• Antimalarials: primaquine and tafenoquine can cause severe haemolysis. Primaquine is commonly used for hypnozoite eradication in P. vivax/P. ovale at 30 mg base orally daily for 14 days in adults when appropriate testing is normal; tafenoquine is given as a 300 mg single dose for radical cure but requires quantitative G6PD testing because it has a long half-life of approximately 14–16 days.
• Antibiotics: sulfonamides, dapsone, nitrofurantoin, and occasionally fluoroquinolones are high-yield oxidant exposures.
• Other drugs: rasburicase and pegloticase generate hydrogen peroxide during uric acid metabolism and are contraindicated in G6PD deficiency. Methylene blue is ineffective and potentially harmful in G6PD deficiency because its reduction to leukomethylene blue requires NADPH.
• Foods/chemicals: fava beans cause favism, especially in Mediterranean variants.

Clinical and laboratory findings

Patients may develop fatigue, pallor, jaundice, dark urine, back or abdominal pain, and splenomegaly after a trigger. Newborns may present with unconjugated hyperbilirubinaemia, increasing risk for kernicterus if severe. Laboratory findings show a haemolytic pattern: falling haemoglobin, increased reticulocyte count after marrow response, increased indirect bilirubin, increased lactate dehydrogenase, and low haptoglobin. Urinalysis may be positive for blood with few or no red cells, reflecting haemoglobinuria.

Peripheral smear is classic: Heinz bodies are seen with supravital stains such as crystal violet or brilliant cresyl blue, while routine Wright-Giemsa smear may show bite cells and blister cells. A key exam concept is that haemolysis may stop spontaneously because the oldest erythrocytes, which have the lowest G6PD activity, are preferentially destroyed; younger reticulocytes have relatively higher enzyme activity.

Diagnosis and testing pitfalls

Diagnosis is confirmed by measuring G6PD enzyme activity, typically reported as units per gram of haemoglobin; normal adult values are laboratory-dependent, often approximately 7–10 U/g Hb. Qualitative fluorescent spot tests detect NADPH generation but may miss heterozygous females and milder deficiency. Quantitative spectrophotometric assays are preferred when drug decisions depend on the result.

A major Step 1 pitfall is false-normal testing during or immediately after an acute haemolytic episode. The most enzyme-deficient older cells have already lysed, leaving reticulocyte-rich blood with higher apparent G6PD activity. If suspicion remains high, repeat testing after haemolysis resolves, commonly after about 2–3 months, approximating red cell population turnover. Current malaria guidance requires G6PD testing before primaquine or tafenoquine; tafenoquine is generally avoided unless activity is clearly normal, often using a threshold of ≥70% of normal activity.

Management principles and high-yield distinctions

Step 1 focuses on mechanism and trigger avoidance rather than advanced management. Treatment is supportive: stop the offending exposure, treat infection, provide hydration, and transfuse only if clinically significant anaemia occurs. Folate may be used in chronic haemolysis because erythropoiesis increases folate demand. Unlike autoimmune haemolytic anaemia, the direct antiglobulin test is negative. Unlike hereditary spherocytosis, osmotic fragility is not the defining defect; the primary lesion is impaired antioxidant defence causing oxidant denaturation of haemoglobin.

Pyruvate Kinase Deficiency

Core Biochemical Defect

Pyruvate kinase (PK) deficiency is an inherited enzymopathy causing chronic extravascular hemolytic anemia. It is classically autosomal recessive and most often due to mutations in PKLR, the gene encoding the erythrocyte and liver isoforms of pyruvate kinase. The key Step 1 concept is that mature red blood cells lack mitochondria and therefore depend entirely on anaerobic glycolysis for ATP generation.

Pyruvate kinase catalyzes the final ATP-generating step of glycolysis:

Phosphoenolpyruvate + ADP → pyruvate + ATP

When PK activity is reduced, erythrocytes cannot generate adequate ATP. ATP is required to maintain red cell membrane ion gradients via pumps such as Na⁺/K⁺-ATPase. ATP depletion causes membrane rigidity, dehydration, and premature destruction by splenic macrophages. Thus, unlike G6PD deficiency, which primarily causes oxidant-induced hemolysis, PK deficiency causes hemolysis from an energy failure of the red cell.

Pathophysiology and High-Yield Consequences

Because pyruvate kinase deficiency blocks a late step in glycolysis, upstream glycolytic intermediates accumulate, especially 2,3-bisphosphoglycerate (2,3-BPG). Increased 2,3-BPG binds deoxygenated hemoglobin and stabilizes the T state, shifting the oxygen-hemoglobin dissociation curve to the right. This promotes oxygen unloading to tissues and can partially compensate for anemia. Therefore, some patients tolerate surprisingly low hemoglobin concentrations compared with other causes of anemia.

Feature	Pyruvate Kinase Deficiency	Step 1 Significance
Inheritance	Autosomal recessive	Often presents in infancy or childhood; variable severity
Metabolic problem	↓ ATP production in RBCs	RBCs depend exclusively on glycolysis
Hemolysis type	Predominantly extravascular	Splenomegaly, jaundice, pigment gallstones
2,3-BPG	Increased	Right-shifted O2-Hb curve; improved tissue oxygen unloading
Blood smear	Echinocytes, also called burr cells	Irregular, short membrane projections

Clinical Presentation

Severity ranges from neonatal jaundice to mild compensated hemolysis discovered incidentally. Typical manifestations include fatigue, pallor, scleral icterus, jaundice, splenomegaly, and pigment gallstones from chronic unconjugated hyperbilirubinemia. Severe neonatal disease may present with marked hyperbilirubinemia requiring phototherapy or exchange transfusion. In chronic hemolysis, the marrow increases erythropoiesis, so reticulocytosis is expected unless marrow reserve is impaired.

Laboratory findings resemble other hemolytic anemias: decreased hemoglobin, increased reticulocyte count, increased lactate dehydrogenase, increased indirect bilirubin, and decreased haptoglobin. Mean corpuscular volume may be normal or mildly elevated because reticulocytes are larger than mature erythrocytes. Direct antiglobulin testing is negative, helping distinguish PK deficiency from autoimmune hemolytic anemia.

Diagnosis

The diagnosis is suspected in a patient with chronic Coombs-negative hemolytic anemia, especially with echinocytes on peripheral smear. Confirmation is by quantitative pyruvate kinase enzyme activity assay or PKLR genetic testing. Enzyme assays can be misleading after recent transfusion because donor erythrocytes have normal PK activity. Testing is ideally performed before transfusion or interpreted with caution.

• Peripheral smear: echinocytes/burr cells; nonspecific but high-yield association.
• Direct Coombs test: negative.
• Reticulocyte count: increased in compensated hemolysis; normal adult reticulocyte percentage is approximately 0.5%–2.5%.
• Confirmatory testing: reduced erythrocyte PK activity or biallelic pathogenic PKLR variants.

Differential Diagnosis

Disorder	Key Mechanism	Classic Smear Finding	Distinguishing Test
Pyruvate kinase deficiency	↓ ATP from glycolysis	Echinocytes	↓ PK activity; PKLR mutations
G6PD deficiency	↓ NADPH; oxidative injury	Bite cells, Heinz bodies	↓ G6PD activity, often after recovery from episode
Hereditary spherocytosis	Membrane cytoskeleton defect	Spherocytes	↑ EMA binding abnormality or osmotic fragility
Warm autoimmune hemolytic anemia	IgG-mediated RBC destruction	Spherocytes	Positive direct Coombs test

Management Principles and Pharmacology

For Step 1, the major emphasis is mechanism and diagnosis, not advanced management. Supportive care includes folate supplementation because chronic hemolysis increases erythropoietic demand. Typical folic acid dosing used in chronic hemolysis is 1 mg orally daily. Transfusions may be needed in severe anemia, aplastic crisis, pregnancy, or perioperative settings. Splenectomy can reduce transfusion requirements because much of the hemolysis is extravascular, but it increases lifelong risk of infection by encapsulated organisms such as Streptococcus pneumoniae, Neisseria meningitidis, and Haemophilus influenzae type b.

Mitapivat is an oral allosteric activator of pyruvate kinase approved for adults with pyruvate kinase deficiency. It increases PK activity, improving ATP generation. Dosing commonly begins at 5 mg orally twice daily and may be titrated to 20 mg twice daily and then 50 mg twice daily based on hemoglobin response and tolerability. In the phase 3 ACTIVATE trial, approximately 40% of treated patients achieved a hemoglobin response compared with 0% receiving placebo. This is useful pharmacology to know conceptually: it treats the enzymatic defect by enhancing residual enzyme function, so response depends on having activatable mutant enzyme.

Classic USMLE Associations

• Autosomal recessive chronic hemolytic anemia due to defective glycolysis.
• ATP depletion causes rigid RBCs removed by the spleen.
• Increased 2,3-BPG causes a right shift of the oxygen dissociation curve.
• Echinocytes on smear are the classic morphologic clue.
• Coombs-negative hemolysis helps separate it from autoimmune hemolytic anemia.

Autoimmune Haemolytic Anaemia

Autoimmune haemolytic anaemia (AIHA) is acquired haemolysis caused by autoantibodies directed against red blood cell (RBC) surface antigens. Normal RBC lifespan is approximately 120 days; in haemolysis, premature destruction exceeds marrow compensation, producing anaemia with reticulocytosis unless marrow function is impaired. The key Step 1 concept is that AIHA is classified by the thermal reactivity and immunoglobulin type of the antibody, which determines the site and mechanism of haemolysis.

Core Pathophysiology

RBC-bound antibodies are detected by the direct antiglobulin test (DAT; direct Coombs test). In this test, anti-human globulin reagent is added to patient RBCs; agglutination indicates RBCs are coated in vivo with IgG, complement C3, or both. The indirect Coombs test detects free anti-RBC antibodies in patient serum and is used in pre-transfusion testing and pregnancy antibody screening.

Haemolysis may be extravascular or intravascular. Extravascular haemolysis occurs when splenic or hepatic macrophages remove antibody- or C3b-coated RBCs. Partial phagocytosis of RBC membrane decreases the surface area-to-volume ratio, producing spherocytes. Intravascular haemolysis occurs when complement activation proceeds to the membrane attack complex, causing RBC lysis within blood vessels.

Type	Antibody	Temperature	DAT Pattern	Mechanism and Associations
Warm AIHA	Usually IgG	37°C	IgG ± C3 positive	Fc receptor-mediated splenic macrophage clearance; spherocytes. Associated with SLE, CLL, lymphomas, and drugs.
Cold agglutinin disease	Usually IgM	Cold exposure, often 0–30°C	C3 positive, IgG negative	IgM binds RBCs in cold peripheral tissues, fixes complement, then dissociates on warming; hepatic clearance of C3b-coated RBCs. Associated with Mycoplasma pneumoniae, EBV, CLL, lymphoplasmacytic disorders.
Paroxysmal cold haemoglobinuria	IgG anti-P	Biphasic	C3 positive	Donath-Landsteiner antibody binds in cold and activates complement on warming; classically post-viral in children, historically syphilis.

Warm Autoimmune Haemolytic Anaemia

Warm AIHA is the most common form. IgG autoantibodies bind RBC antigens at body temperature. Splenic macrophages recognize IgG Fc portions via Fc receptors, phagocytose part or all of the RBC, and generate extravascular haemolysis. Peripheral smear shows spherocytes and polychromasia from reticulocytosis. This can resemble hereditary spherocytosis, but AIHA has a positive DAT, whereas hereditary spherocytosis classically has a negative DAT.

Common causes include idiopathic disease, systemic lupus erythematosus, chronic lymphocytic leukaemia, non-Hodgkin lymphoma, and drugs. Drug-induced immune haemolysis may occur by several mechanisms: α-methyldopa can induce true autoantibodies against RBC antigens; high-dose penicillin can act as a hapten coating RBCs; cephalosporins can cause immune-complex or drug-dependent antibody haemolysis.

Cold Agglutinin Disease

Cold agglutinin disease is usually mediated by IgM, which efficiently fixes complement because pentameric IgM binds C1q strongly. In cooler body regions such as fingers, toes, ears, and nose, IgM binds RBCs and causes agglutination. When blood returns to central warmer temperatures, IgM dissociates, but C3b remains attached. The result is mainly hepatic macrophage clearance of complement-coated RBCs, although severe complement activation can cause intravascular haemolysis. Clinical clues include haemolysis worsened by cold exposure, acrocyanosis, and RBC agglutination on smear or automated CBC artefacts.

Laboratory Findings

• Anaemia: decreased haemoglobin/haematocrit; normal adult haemoglobin is roughly 13.5–17.5 g/dL in males and 12.0–15.5 g/dL in females.
• Reticulocytosis: normal reticulocytes are approximately 0.5–2.5%; haemolysis increases reticulocyte count if marrow response is intact.
• High LDH and high unconjugated bilirubin from haem breakdown.
• Low haptoglobin, especially in intravascular haemolysis, because haptoglobin binds free plasma haemoglobin.
• Positive DAT: the defining test for immune-mediated haemolysis.
• Peripheral smear: spherocytes in warm AIHA; RBC agglutination in cold agglutinin disease.

Clinical Correlation and Treatment Principles

Patients may present with fatigue, pallor, jaundice, dark urine, splenomegaly, or symptoms triggered by cold. Severe haemolysis can produce tachycardia, dyspnoea, or angina due to reduced oxygen delivery. Transfusion may be difficult because autoantibodies complicate cross-matching, but life-threatening anaemia should still be treated with the best-matched blood available.

Condition	First-Line Approach	Additional High-Yield Therapy
Warm AIHA	Glucocorticoids, commonly prednisone about 1 mg/kg/day	Rituximab 375 mg/m² weekly for 4 weeks or splenectomy in refractory disease; treat underlying SLE, CLL, or lymphoma.
Cold agglutinin disease	Avoid cold exposure; warmed transfusions if needed	Rituximab-based therapy for clinically significant disease; corticosteroids are generally less effective than in warm AIHA.
Paroxysmal cold haemoglobinuria	Supportive care and cold avoidance	Often self-limited in children after viral illness.

For Step 1, the essential discriminator is: warm AIHA = IgG, splenic extravascular haemolysis, spherocytes, DAT IgG positive; cold AIHA = IgM-mediated complement fixation, DAT C3 positive, cold-induced agglutination.