What is the appropriate workup and management for a patient suspected of having autoimmune hemolytic anemia (AIHA)?

Workup for Autoimmune Hemolytic Anemia

Begin with a comprehensive laboratory panel including CBC with differential, reticulocyte count, peripheral blood smear, direct antiglobulin test (DAT/Coombs) with monospecific antisera, LDH, haptoglobin, and bilirubin (direct and indirect) to establish the diagnosis of AIHA. 1, 2

Essential Initial Laboratory Tests

Core Hemolysis Markers

• Complete blood count (CBC) showing anemia with macrocytosis is the foundation of diagnosis 1, 3
• Reticulocyte count should be elevated (typically >2-3%), indicating active bone marrow compensation for hemolysis 1, 4
• Peripheral blood smear to identify spherocytes (characteristic of warm AIHA), agglutination (cold AIHA), schistocytes, or other red cell morphology abnormalities 1, 3
• Lactate dehydrogenase (LDH) will be markedly elevated, particularly in intravascular hemolysis 1, 4
• Haptoglobin will be decreased or absent due to binding of free hemoglobin 1, 4
• Bilirubin with elevated indirect/unconjugated fraction confirms hemolysis 1, 4

Critical Diagnostic Test

• Direct antiglobulin test (DAT/Coombs) with monospecific antisera is mandatory and must be performed before initiating any treatment 1, 2
  • Positive for IgG only suggests warm AIHA 3, 2
  • Positive for C3d only suggests cold agglutinin disease (CAD) or paroxysmal cold hemoglobinuria 3, 2
  • Positive for both IgG and C3d indicates mixed AIHA 3, 2

Important caveat: DAT-negative AIHA occurs when antibody levels are below detection thresholds or involve IgM/IgA antibodies not detected by standard testing. If clinical suspicion remains high despite negative DAT, proceed with treatment as for warm AIHA and assess response to corticosteroids. 1, 3

Excluding Secondary Causes and Mimics

Medication Review

• Obtain detailed drug history evaluating for common causative agents: 1, 5
  • Fludarabine (contraindicated in AIHA, can cause life-threatening hemolysis) 5
  • Fluoroquinolones (ciprofloxacin, levofloxacin) 1, 5
  • Penicillins and cephalosporins 1, 5
  • Rifampin, dapsone, NSAIDs, quinine/quinidine 1
  • Interferon, lorazepam, diclofenac 1

Nutritional and Metabolic Causes

• B12, folate, copper, and iron studies to exclude deficiencies that can mimic or coexist with hemolysis 1
• Thyroid function tests as thyroid disorders can be associated with AIHA 1
• Glucose-6-phosphate dehydrogenase (G6PD) activity to exclude G6PD deficiency (note: may be falsely normal during acute hemolysis due to young RBCs) 1

Infectious Workup

• Viral studies including HIV, hepatitis B and C, CMV serology, and parvovirus B19 to identify triggers 1
• Consider Mycoplasma pneumoniae and EBV in appropriate clinical contexts 3

Exclude Thrombotic Microangiopathy

• Maintain high index of suspicion for TTP/HUS if schistocytes are present on smear 1
• Check ADAMTS13 activity if TTP is suspected 1
• Immediate hematology consultation and plasma exchange may be life-saving 1

Identifying Underlying Lymphoproliferative or Autoimmune Disorders

Secondary AIHA Evaluation

• Lymphoproliferative disorders: Obtain flow cytometry, lymph node imaging (CT chest/abdomen/pelvis), and consider bone marrow biopsy if lymphoma or CLL is suspected 3, 2
• Systemic autoimmune diseases: Check ANA, anti-dsDNA, complement levels (C3, C4), and rheumatoid factor for SLE or other connective tissue diseases 3, 2
• Immunodeficiency syndromes: Consider immunoglobulin levels and lymphocyte subsets, particularly in younger patients 3

Determining AIHA Subtype

Warm AIHA (Most Common)

• DAT positive for IgG with or without C3d 3, 2
• Spherocytes on peripheral smear 3
• Hemolysis occurs at body temperature 2

Cold Agglutinin Disease (CAD)

• DAT positive for C3d only 3, 2
• Cold agglutinin titer typically >1:512 with monoclonal IgM 2, 6
• Hemolysis worsens with cold exposure 2
• Distinguish primary CAD (clonal lymphoproliferative disorder) from secondary cold agglutinin syndrome 2, 6

Mixed AIHA

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

Paroxysmal Cold Hemoglobinuria (PCH)

• DAT positive for C3d only 3
• Donath-Landsteiner test positive (biphasic cold-reactive IgG) 3
• Often self-limiting, associated with viral infections 3

Special Monitoring Considerations

Reticulocytopenia warning: 20-40% of AIHA cases present with inadequate reticulocyte response due to marrow involvement, nutritional deficiencies, infections, or autoimmune reaction against erythroid precursors—this is a poor prognostic factor requiring more aggressive management. 4

Thrombocytopenia alert: If present alongside hemolysis, consider Evans' syndrome (combined AIHA and immune thrombocytopenia) or thrombotic microangiopathy, both requiring modified treatment approaches. 4