Investigations
1st investigations to order
haemoglobin (Hb)
mean corpuscular volume (MCV)
Test
The MCV correlates with phenotypic and genotypic worsening of the disease. Patients with alpha-thalassaemia silent carrier have a normal to slightly low MCV, whereas those with deletional alpha-thalassaemia trait usually have an MCV of <78 femtolitres.[48] Those with deletional Hb H disease will have an MCV in the mid-60s.[43] Patients with Hb H/Constant Spring will have an MCV that is normal to mildly low, and those homozygous for Hb Constant Spring will often have a normal MCV.
Result
low
mean corpuscular haemoglobin (MCH)
Test
MCH will be normal to low (24-29 picograms/cell) in those with 1 alpha-thalassaemia silent carrier, and it will be low (<27 picograms/cell) in those with >1 alpha-globin deletion or in those with non-deletional mutations. Mean MCH is approximately 20 picograms/cell in Hb H disease.[43]
Result
low
red blood cell count
Test
Should be performed in all patients.
Result
increased; variable, corresponding to severity
peripheral smear
reticulocyte percentage
Test
Elevated (5% to 10%), and may be further increased during acute infections or haemolytic crises.[48]
Result
increased
serum iron
Test
Result
normal or elevated
Investigations to consider
brilliant cresyl blue staining of red blood cells
Test
Numerous inclusion bodies are typically seen in Hb H disease.[16] Felt by some experts to be more reliable than Hb electrophoresis for diagnosis of Hb H disease.
May be reduced or absent in the presence of heterozygous beta-thalassaemia such as Hb S, Hb C, Hb E, or beta-thalassaemia.[43][54]
Result
demonstrates Hb H inclusions in peripheral blood red cells
Hb electrophoresis
Test
Hb H and Hb Bart can be detected as fast-moving haemoglobins. Will not pick up alpha-thalassaemia silent carrier or alpha-thalassaemia trait outside of the newborn period (Hb Barts), and will not differentiate deletional from non-deletional forms of Hb H (except for Hb Constant Spring). Patients with Hb H disease and concomitant heterozygous beta-haemoglobinopathies such as Hb S, Hb C, Hb E, or beta-thalassaemia have low or absent Hb H, which may confound the diagnosis of Hb H disease.
Result
presence of Hb H, Hb Bart, and concomitant haemoglobinopathies (Hb E, Hb S, Hb C, Hb D)
Hb fractionation by high-performance liquid chromatography (HPLC)
Test
Hb fractionation by HPLC provides fast separation and precise estimation of haemoglobin variants.
Will not pick up alpha-thalassaemia silent carrier or alpha-thalassaemia trait outside of the newborn period (Hb Barts), and will not differentiate deletional from non-deletional forms of Hb H (except for Hb Constant Spring). Patients with Hb H disease and concomitant heterozygous beta-haemoglobinopathies such as Hb S, Hb C, Hb E, or beta-thalassaemia have low or absent Hb H, which may confound the diagnosis of Hb H disease.
Result
presence of Hb H, Hb Bart, and concomitant haemoglobinopathies (Hb E, Hb S, Hb C, Hb D)
gap-polymerase chain reaction (gap-PCR)
Test
Seven of the most common alpha-thalassaemia deletions (-alpha(3.7), -alpha(4.2), --(FIL), --(THAI), --(MED), -(alpha)(20.5) , --(SEA)) can be diagnosed by gap-PCR.[48]
Result
detects specific deletions
multiplex ligation-dependent probe amplification
Test
Used to detect deletions and duplications in the alpha-globin gene cluster causing alpha-thalassaemia.[48]
Result
detects deletion alleles
direct sequencing/reverse dot blot
MRI (hepatic or cardiac)
Test
Hepatic iron overload can be assessed by R2 or R2* MRI.[51]
MRI is the gold standard for measurement of hepatic iron concentration.[16]
Cardiac iron loading is assessed by T2* cardiac MRI.[48] Cardiac iron loading is uncommon in non-transfused patients.
Result
hepatic or cardiac iron in patients with iron overload is demonstrated by difference in organ-to-muscle signal intensity
superconducting quantum interference devices (SQUID)
liver biopsy
Test
If iron status is significantly elevated as evident by a serum ferritin >1797.6 picomol/L (>800 nanograms/mL), hepatic iron overload can be assessed by liver biopsy.[50][51]
MRI is preferred to liver biopsy for assessment of iron overload where possible as it has better accuracy and fewer procedural risks. Liver biopsy may be used where assessment of histology is needed.[21]
Result
liver iron ≥5 mg/gram dry weight on liver biopsy indicates need for iron chelation therapy
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