Diagnostic Workup for Alpha Thalassemia Trait
Begin with a complete blood count (CBC) showing microcytosis (MCV <80 fL, often below the 5th percentile for age) with normal or elevated red blood cell count and normal RDW (≤14.0%), followed by serum ferritin to exclude iron deficiency, then proceed to definitive DNA testing for alpha globin gene deletions or mutations. 1, 2
Initial Laboratory Screening
Complete Blood Count Parameters
The CBC reveals characteristic findings that distinguish alpha thalassemia trait from iron deficiency anemia:
- Mean Corpuscular Volume (MCV): Values typically <80 fL and frequently below the 5th percentile for age 1, 2
- Red Blood Cell (RBC) Count: Normal or elevated, which is a critical differentiating feature from iron deficiency where RBC count is low 1, 3
- Red Blood Cell Distribution Width (RDW): Usually normal (≤14.0%), whereas iron deficiency typically shows RDW >14.0% 1, 3
- Hemoglobin: May show mild anemia or be normal 4, 5
Iron Studies
- Serum Ferritin: Must be normal to confirm alpha thalassemia trait and exclude concurrent iron deficiency 1, 3
Critical Pitfall: If iron deficiency is present (low ferritin), you must provide iron replacement therapy before proceeding with hemoglobin analysis, as iron deficiency can mask thalassemia characteristics and falsely lower HbA2 levels 3
Definitive Diagnostic Testing
DNA-Based Molecular Testing
DNA testing for alpha globin gene deletions or point mutations is the definitive diagnostic test and should be performed once iron deficiency is excluded 2, 6
The most common mutations to identify include:
- 3.7 kb single gene deletion (α+-thalassemia): Most common mutation, found in approximately 39% of cases 6
- 20.5 kb double gene deletion (α0-thalassemia): Found in approximately 21% of cases 6
- MED double gene deletion: Found in approximately 18% of cases 6
- 4.2 kb single gene deletion: Less common α+-thalassemia variant 6, 7
Modern molecular techniques include:
- PCR-based strategies: Have largely replaced Southern blotting methodology for straightforward identification 7
- Commercial strip assays: Available for detecting common deletions and point mutations 6
- Screening systems: Including DGGE and SSCP followed by direct sequencing for point mutations 7
Newborn Screening Correlation
In infants, a "fast hemoglobin variant" (Hemoglobin Bart's, 2-8%) detected on newborn screening is highly specific for alpha thalassemia trait, with a prevalence of approximately 2.5% in screened populations 8
Ethnicity Considerations
Failure to consider ethnicity is a common diagnostic pitfall 2:
- Alpha thalassemia is most prevalent in Southeast Asian, Mediterranean, Middle Eastern, and African populations 2, 3
- In Southeast Asian populations, alpha thalassemia accounts for 28-55% of non-immune hydrops fetalis cases 1, 2
Genetic Counseling and Family Implications
Once alpha thalassemia trait is confirmed:
- Cascade testing approach: Test the affected individual first to identify the specific mutation, then offer targeted testing to at-risk family members 2
- Reproductive counseling: Critical for couples where both partners are carriers, as they face a 25% risk with each pregnancy of having a child with severe disease (Hemoglobin H disease or Hemoglobin Bart's hydrops fetalis) 1, 3
- Partner screening: Mandatory when one partner is identified as a carrier, including CBC with MCV measurement and DNA testing if MCV <80 fL 3
Differential Diagnosis Algorithm
When encountering microcytic anemia:
- Check MCV: If <80 fL, proceed to step 2 2, 3
- Assess RBC count and RDW:
- Measure ferritin: Normal ferritin supports thalassemia; low ferritin indicates iron deficiency 1, 3
- Perform DNA testing: Definitive diagnosis of alpha thalassemia trait 2, 6
Common Pitfall: Alpha thalassemia should always be considered in the differential diagnosis of hypochromic microcytic anemia, especially in cases without iron deficiency and without beta-thalassemia carrier state 6