Capillary zone electrophoresis: An additional technique for the identification of hemoglobin variants

Abstract

Two capillary zone electrophoresis kits (Hb A₂ and Hb A_1c) were tested for confirmation and identification of hemoglobin variants. The capillary zone electrophoresis experiments were performed at pH 4.7 (Hb A_1c kit) and 8.7 (Hb A₂ kit) in a 24 cm uncoated fused silica capillary tube (25 μm I.D.). Normal hemoglobins and common hemoglobin variants, including Hbs S, D-Punjab, C, E, O-Arab, and G-Philadelphia, were successfully separated by both methods within a few minutes. Both systems provided completely different elution profiles of normal and abnormal hemoglobin fractions tested and were complementary. The inter-assay coefficient of variations of the migration times of hemoglobin variants were less than 1.0 and 1.3% by the Hb A₂ and Hb A_1c, respectively. This permits a higher resolution of some hemoglobin variants in low concentrations, like Hb S in newborns, compared with conventional electrophoresis methods. The present capillary zone electrophoresis methods are sensitive, rapid, not labor intensive, and highly selective for the separation of hemoglobin variants. Combination of both methods with some conventional methods, such as isoelectrofocusing, allows identification of Hbs C, E, O-Arab, S, and D-Punjab, as well as their quantification. We have demonstrated that the conventional electrophoresis methods (electrophoresis at pH 6.5 in citrate agar gel and electrophoresis at pH 8.6 on cellulose acetate) can be advantageously replaced by the present capillary zone electrophoresis methods in a clinical laboratory practice for the detection and quantification of hemoglobin variants.