Thalassemias

Indications for Testing

Laboratory testing for thalassemia is used to :

• Perform carrier screening in individuals from populations with a high frequency of thalassemia or with a family history of thalassemia, or in reproductive partners of an individual diagnosed with thalassemia
• Diagnose thalassemia in individuals with anemia or isolated microcytosis

Laboratory Testing

Screening

Carrier screening (with genetic counseling) is appropriate before pregnancy for individuals with a family history of thalassemia, in reproductive partners of known thalassemia carriers, and in individuals from populations with a high incidence of thalassemia.  Timely screening can help inform reproductive choices, ensure appropriate maternal care, and facilitate diagnosis in newborns.  Due to the wide range in disease severity, the CDC recommends screening newborns for both α and β thalassemias.  Primary screening is generally performed using HPLC or IEF, and results are confirmed using a second technique. 

Diagnosis

At least two complementary techniques (eg, a combination of HPLC and electrophoresis) should be used in the initial identification of a thalassemia.  Genetic testing may be needed for definitive diagnosis. 

Initial Evaluation

The first step in the evaluation of a suspected thalassemia is a CBC with peripheral smear.  Serum iron studies are also helpful for distinguishing between thalassemias and other microcytic anemias, particularly iron deficiency anemia.

Hemoglobin Assays

Hb testing is appropriate in the following circumstances :

• Hydrops fetalis
• Anemia and low or absent HbA in a neonate or infant
• Unexplained anemia and splenomegaly
• Unexplained microcytosis
• Suspected thalassemia
• Unexplained target cells on peripheral smear

A variety of techniques can be used for Hb analysis. The two most commonly used techniques are HPLC and IEF.  Capillary zone electrophoresis, acid or alkaline gel electrophoresis, and other methods may also be used.  Each technique has advantages and disadvantages, and not all Hb variants can be detected by every technique. 

Hb electrophoresis or equivalent techniques should not be repeated in patients who have a previous test result and do not require therapeutic intervention. Repeat testing should only be used to make a more specific diagnosis, ^,  or if results may have been complicated by the presence of donor Hb from a recent transfusion.  Severe iron deficiency anemia may reduce the HbA2 level by as much as 0.5%. 

Genetic Testing

Molecular genetic testing is recommended to confirm the results of Hb analysis. ^, , 

Alpha Thalassemia

Targeted deletion analysis for common HBA1 and HBA2 variants is recommended as a first genetic test for α thalassemia, followed by sequencing.  If neither targeted deletion analysis nor sequencing yields diagnostic results, full gene-targeted deletion analysis may be required.  Alternatively, a full gene-targeted deletion analysis may be used initially to detect both common and uncommon deletions, followed by a sequence analysis if no deletions are detected. 

For prenatal counseling, it is important to know whether the nonfunctioning α-globin genes lie on the same chromosome (cis) or on opposite chromosomes (trans). When both parents carry a cis deletion of HBA1 and HBA2 (--/αα), the risk for lethal hydrops fetalis associated with Hb Barts in their offspring may be 1:4. 

Beta Thalassemia

More than 200 known HBB variants are categorized into two classes: β zero (β0, no β-globin chain synthesis from the affected allele) and β plus (β+, decreased β-globin chain synthesis from the affected allele).  HBB sequencing, including splice sites and other intronic regions, the proximal promoter region, and the 5’ and 3’ untranslated regions (UTRs), detects or confirms nearly all β thalassemia cases.  In rare cases, β-globin deletion/duplication analysis may be required. 

Monitoring

Patients with β thalassemia major are transfusion dependent, whereas patients with HbH disease or β thalassemia intermedia may require occasional transfusion. Regular transfusion is associated with a number of complications, including infections, development of antibodies to RBCs, hemolytic reactions, and iron overload that leads to organ failure.