Separation of Hemoglobin Variants by Capillary Electrophoresis

Abstract

Quantitative defects in hemoglobin (Hb) are represented by Hb variants, where the amino acids sequence is modified as a consequence of a mutation in the α or β-globin genes. More than 1,100 variants have been described so far but only a few dozen are clinically significant; the most significant being Hb S, which in the homozygous state causes sickle cell disease. The majority of the methods used to detect Hb variants are based on the charge difference of the mutated globin chain. We have developed a micellar capillary electrophoresis (MEKC) method using highly acidic conditions and a high Triton(®) concentration. Separation times in the order of 20 min were able to resolve all normal and 29 abnormal globin chains including Hb E. This method was initially developed for Beckman P/ACE 5500 Instrument but has been modified for the more recent P/ACE MDQ and PA 800 instruments; however, the method can be adapted to any kind of CE analyzer.