Elevated Hemoglobin <scp> A ₂ </scp> : A Molecular Revisited, and Implications to β‐Thalassemia Screening

ABSTRACT Background In Thailand, the Hb A 2 cut‐off value for β‐thalassemia carrier has been changed from 4.0% to 3.6% since 2015. We examined the molecular basis of β‐thalassemia in a large cohort of Thai subjects with this change. Methods A total of 36,313 retrospective specimens encountered during January 2016 to June 2025 were reviewed, and 5909 subjects (16.7%) with Hb A 2 ≥ 3.6% were recruited. Diagnosis of thalassemia was based on Hb and DNA analyses. Results Among the 5909 subjects, 5395 (91.3%) had Hb A 2 A, 115 (1.9%) carried Hb variants, and the remaining 399 (6.8%) resulted from misinterpretations. Of 5395 subjects, 5117 (94.8%) carried β‐thalassemia, 189 (3.5%) had normal β‐globin genes, and 89 (1.6%) had incomplete molecular analysis. Fifty different β‐thalassemia genes were identified, including nine hitherto undescribed in Thailand and two novel mutations, β CD81(CTC &gt; CTA) and β IVSII‐713(G &gt; A) . Falsely elevated Hb A 2 was found in 364 of 399 (70.8%) subjects with misinterpretation, 20 (3.9%) represented human errors, and 15 (2.9%) resulted from Hb E blood transfusion. A low proportion of β 0 ‐ and a high proportion of β + ‐thalassemias, a normal β‐globin gene, and diagnostic errors were observed at borderline Hb A 2 levels. Conclusions A change in the Hb A 2 cut‐off can alter the molecular spectrum of β‐thalassemia in Thailand. The molecular basis of β‐thalassemia was updated to include 50 mutations, both new and known. Misinterpretation at borderline Hb A 2 should be aware and cost‐effectiveness should be considered at the screening of β‐thalassemia in the region.