Diagnosis of Hemoglobinopathies and thalassemias using both Haemoglobin capillary zone electrophoresis and High performance liquid chromatography - An experience from a tertiary care hospital in Northern India

Abstract Being an important cause of morbidity and mortality, thalassemias and hemoglobinopathies impose a heavy burden on families and the health sector in our country. It has has been estimated that in India, 0.37 per 1,000 fetuses have Hb disorder and more than 10,000 children are born annually with severe hemoglobinopathy and only around 10.0% are managed optimally, therefore screening and diagnosis of carriers is of paramount importance especially in an antenatal OPD. While Haemoglobin capillary zone electrophoresis (HbCZE) and High performance liquid chromatography (HPLC) are commonly used methods for diagnosis of these entities, there are far and few studies which attempt to compare these two techniques. Haemoglobin A2 (HbA2) constitutes less than 3% of the total hemoglobin (Hb) in adults and the determination of Hb A2 levels is important to diagnose the beta thalassemia trait (BTT). In some cases, the level of HbA2 is not typically elevated and some difficulties may arise in making the diagnosis and hence accurate assessment of HbA2 becomes of paramount importance. HPLC and HbCZE are considered acceptable methods to diagnose BTT but these vary in their accuracy and cut-offs. Complex elution patterns as well as co-elution of variants like HbA2, HbE, Hb Lepore, HbD Iran create difficulty in interpretation of HPLC plots.The aim of this study was to compare the results of HPLC and HbCZE technique for screening of clinically significant haemoglobin disorders. On comparing the HbA2 values the HPLC showed higher values for HbA2 with a median values as compared to HbCZE while when no variant was present, the Hb A2 concentrations showed excellent agreement. The Hb F, HbS measurement agreement was good between both methods. To conclude HPLC and HbCZE are complementary techniques and normal ranges and means normal values differ between different methods and different manufacturers and therefore it is important that each lab establish its own cut offs.