Discrepancies in hemoglobin A2 measurement between capillary electrophoresis and high-performance liquid chromatography in β-thalassemia trait

Background
Accurate quantification of hemoglobin A2 (HbA2) is crucial for detecting β-thalassemia trait. High-performance liquid chromatography (HPLC) and capillary electrophoresis (CE) are widely used methods; however, discrepancies in their results remain a concern. The aim of this study is to evaluate the concordance of HbA2 measurements by HPLC and CE in β-thalassemia trait and study the potential factors causing discrepancies between both methods.


Patients and methods
This cross-sectional study included 55 newly diagnosed β-thalassemia trait children and adolescents in addition to 40 healthy controls presented to the Pediatrics Department of the Faculty of Medicine at Menoufia University in Egypt. Both HPLC and CE were used to quantify HbA2 levels, and intraclass correlation coefficient analysis was conducted to evaluate the degree of agreement. Multivariate analysis was conducted to find independent determinants of measurement variations.


Results
HbA2 levels were slightly higher by CE compared with HPLC (P<0.001). Discrepancies between both methods (ΔHbA2) were influenced by mean corpuscular hemoglobin (MCH) (positive correlation, P=0.014), HbF (negative correlation, P=0.018), and red blood cell distribution width (negative correlation, P=0.030). Multivariate analysis identified MCH and HbF as independent predictors of difference in HbA2 measurements by both methods.


Conclusion
HPLC and CE show good agreement in HbA2 measurement among β-thalassemia trait patients with a slight systematic bias. MCH and HbF are independent predictors of discrepancies between the two methods necessitating the need to account for these factors in diagnostic evaluations.