What are the clinical features and lab diagnosis of Autoimmune Hemolytic Anemia (AIHA)?

Clinical Features and Laboratory Diagnosis of Autoimmune Hemolytic Anemia (AIHA)

Autoimmune hemolytic anemia is characterized by immune-mediated destruction of red blood cells through autoantibody production, with diagnosis requiring evidence of hemolysis and a positive direct antiglobulin test in most cases. 1

Clinical Presentation

• Patients commonly present with symptoms of anemia including weakness, fatigue, pallor, jaundice, dark-colored urine, and reduced exercise tolerance 1
• Physical examination may reveal tachycardia, heart murmurs, splenomegaly, and jaundice 1
• Onset can be acute or insidious, with varying severity from mild anemia to life-threatening hemolysis 2
• Fever may be present in some cases, particularly with severe hemolysis 1
• Clinical presentation may vary based on AIHA subtype, with cold agglutinin disease often presenting with acrocyanosis and Raynaud's phenomenon during cold exposure 3

Laboratory Evidence of Hemolysis

• Complete blood count (CBC) showing anemia with macrocytosis 1
• Elevated reticulocyte count (typically >2%) reflecting bone marrow compensation 1, 4
• Peripheral blood smear showing spherocytes, polychromasia, and sometimes red cell agglutination 1, 5
• Elevated lactate dehydrogenase (LDH) indicating intravascular hemolysis 1, 4
• Decreased or absent haptoglobin due to binding with free hemoglobin 1, 4
• Elevated indirect (unconjugated) bilirubin 1, 4
• Free hemoglobin in plasma and/or urine in cases of severe intravascular hemolysis 1

Immunological Testing

• Direct antiglobulin test (DAT/Coombs test) is the cornerstone of diagnosis 4
  • Positive for IgG in warm AIHA
  • Positive for C3d in cold agglutinin disease
  • Positive for both IgG and C3d in mixed AIHA 3
• Indirect antiglobulin test to detect free autoantibodies in serum 1
• Thermal amplitude testing to differentiate warm from cold antibodies 3
• DAT-negative AIHA (5-10% of cases) may occur due to low-affinity antibodies or IgA/IgM antibodies below detection threshold 3, 4

AIHA Classification

Based on Antibody Thermal Reactivity

• Warm AIHA (60-70% of cases):

  • Antibodies (usually IgG) react optimally at 37°C
  • DAT positive for IgG and/or C3d 3, 2

• Cold Agglutinin Disease (20-25% of cases):

  • IgM antibodies react optimally at 4°C
  • DAT positive for C3d only 3, 2

• Mixed AIHA (7-8% of cases):

  • Features of both warm and cold antibodies
  • DAT positive for both IgG and C3d 3, 5

• Paroxysmal Cold Hemoglobinuria (rare):

  • Biphasic IgG (Donath-Landsteiner antibody)
  • DAT positive for C3d only 3

Based on Etiology

• Primary/Idiopathic AIHA: No identifiable underlying cause 4
• Secondary AIHA: Associated with underlying conditions 5
  • Lymphoproliferative disorders (CLL, NHL)
  • Autoimmune disorders (SLE, rheumatoid arthritis)
  • Infections (Mycoplasma, EBV, HIV)
  • Drugs (penicillins, cephalosporins, NSAIDs, etc.)
  • Immunodeficiencies
  • Solid tumors
  • Post-transplantation 2, 4

Additional Diagnostic Workup

• Autoimmune serology to identify underlying autoimmune disorders 1
• Viral and bacterial testing, especially for mycoplasma, HIV, and hepatitis 1
• Protein electrophoresis to rule out monoclonal gammopathy 1, 2
• Flow cytometric analysis of lymphocyte subsets in suspected secondary cases 5
• Bone marrow examination in selected cases to rule out underlying hematologic malignancies 1
• Evaluation for drug-induced hemolysis by medication history 1
• Screening for paroxysmal nocturnal hemoglobinuria 1
• Glucose-6-phosphate dehydrogenase assessment to rule out enzymatic causes 1

Diagnostic Pitfalls and Challenges

• Normal IgG levels do not exclude AIHA, especially in acute presentations 6
• Autoantibody titers may vary during disease course; initially seronegative patients may become positive later 6
• Recent blood transfusions can complicate diagnosis and interpretation of test results 1
• Incomplete removal of platelets and leukocytes can lead to false-normal enzyme activity levels 1
• Comprehensive autoimmune serology workup may not be available in all laboratories; reference laboratories should be used in cases of diagnostic uncertainty 1
• Repeated testing may be necessary if initial results are negative but clinical suspicion remains high 6
• In acute severe presentations, typical laboratory findings may be absent or atypical 6

Diagnostic Algorithm

1. Establish presence of anemia and evidence of hemolysis (CBC, reticulocyte count, LDH, haptoglobin, bilirubin) 4
2. Perform DAT to confirm immune-mediated hemolysis 4
3. Characterize antibody type (warm vs. cold) through thermal amplitude testing 3
4. Screen for underlying conditions based on AIHA subtype 5, 2
5. In DAT-negative cases with strong clinical suspicion, consider more sensitive methods or referral to specialized centers 3
6. Evaluate response to initial therapy as a diagnostic criterion in unclear cases 4