Annals of laboratory medicine · 2026
Assessment of a High-Resolution HPLC System to Quantify HbE for α0-Thalassemia Diagnosis in HbE Heterozygotes.
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Srisuwan Wibhasiri, Satthakarn Surada, Panyasai Sitthichai
Аннотация
BACKGROUND: HbE and α0-thalassemia are highly prevalent in Southeast Asia and co-inherited in asymptomatic carriers. α0-Thalassemia screening in HbE heterozygotes relies on HbE quantification. We evaluated the performance of a high-resolution HPLC system in quantifying HbE and HbA2 to distinguish HbE heterozygotes with and without α0-thalassemia.
METHODS: HbE heterozygotes (N=650) presenting EA patterns on capillary electrophoresis (CE) were analyzed using the Premier Resolution (PR) HPLC system (Trinity Biotech). HbA2 and HbE levels were quantified, Pearson correlations with CE assessed, and diagnostic cutoffs determined. A prospective cohort of HbE heterozygotes (N=884) was used for validation. α0-Thalassemia genotyping was performed using gap-PCR.
RESULTS: PR-HPLC confirmed HbE heterozygosity and successfully separated HbA2 from HbE. The HbA2, HbE, and combined HbA2+HbE levels were consistently higher than but correlated with CE values (HbA2: bias=1.213, r=0.686; HbE: bias=0.286, r=0.960; combined: bias=1.502, r=0.952). HbE and combined levels were substantially lower in α0-thalassemia carriers (14.65%±2.2% vs. 19.38%±2.5%); HbA2 levels did not significantly differ. HbE cutoff <19% resulted in an area under the curve (AUC) of 0.975, with 100% sensitivity and 91.9% specificity. HbE+HbA2 <24.0% yielded an AUC of 0.970, with 100% sensitivity and 89.7% specificity. Validation experiments supported the robust performance for HbE detection alone (100% sensitivity, 93.6% specificity, and 94.5% accuracy).
CONCLUSIONS: PR-HPLC can quantify HbE precisely, potentially aiding in robust α0-thalassemia diagnosis and screening without HbA2 assessment.
METHODS: HbE heterozygotes (N=650) presenting EA patterns on capillary electrophoresis (CE) were analyzed using the Premier Resolution (PR) HPLC system (Trinity Biotech). HbA2 and HbE levels were quantified, Pearson correlations with CE assessed, and diagnostic cutoffs determined. A prospective cohort of HbE heterozygotes (N=884) was used for validation. α0-Thalassemia genotyping was performed using gap-PCR.
RESULTS: PR-HPLC confirmed HbE heterozygosity and successfully separated HbA2 from HbE. The HbA2, HbE, and combined HbA2+HbE levels were consistently higher than but correlated with CE values (HbA2: bias=1.213, r=0.686; HbE: bias=0.286, r=0.960; combined: bias=1.502, r=0.952). HbE and combined levels were substantially lower in α0-thalassemia carriers (14.65%±2.2% vs. 19.38%±2.5%); HbA2 levels did not significantly differ. HbE cutoff <19% resulted in an area under the curve (AUC) of 0.975, with 100% sensitivity and 91.9% specificity. HbE+HbA2 <24.0% yielded an AUC of 0.970, with 100% sensitivity and 89.7% specificity. Validation experiments supported the robust performance for HbE detection alone (100% sensitivity, 93.6% specificity, and 94.5% accuracy).
CONCLUSIONS: PR-HPLC can quantify HbE precisely, potentially aiding in robust α0-thalassemia diagnosis and screening without HbA2 assessment.