The detection and diagnosis of hemoglobin A₂′ by high-performance liquid chromatography

Hemoglobin (Hb) A₂′ is a hematologically silent variant of HbA₂ that is detected easily by high-performance liquid chromatography (HPLC), where it elutes in the S window. Our purposes were to define diagnostic criteria for the HbA₂′ trait using the Variant II (Bio-Rad, Hercules, CA) and to determine the prevalence of HbA ₂′ in a metropolitan patient population. All Hb screens (N = 5,862) performed during a 26-month period were reviewed for new hemoglobinopathies. We identified 57 cases of HbA₂′ trait, making it the fourth most prevalent Hb variant detected in this population after HbS, HbC, and β-thalassemia minor. For HbA₂′ trait cases, the mean HbA₂ level was 1.7% (SD, 0.17%), and the mean HbA ₂′ level was 1.3% (SD, 0.18%). Six possible HbA ₂′/β-thalassemia double heterozygotes were identified, for whom the sum of the HbA₂ and HbA₂′ exceeded 4% of total Hb. Hb variants that might interfere with detection of HbA ₂′ include HbS, glycosylated HbC, and HbG₂. Diagnostic criteria proposed for the HbA₂′ trait by HPLC are HbA₂ of 2% or less, S window peak of 1% to 2%, no previous diagnosis of HbS, and absence of HbG and HbC.