A comprehensive analysis of hemoglobin variants by high‐performance liquid chromatography (HPLC)

Summary Introduction High‐performance liquid chromatography ( HPLC ) is a method commonly used for the detection of hemoglobin ( H b) variants. In addition to providing precise quantitation of H b A 2 and H b F , the reported retention time and peak shape of a high number of hemoglobin ( H b) variants are very helpful for presumptive identification. However, there is a scarcity of summarized data in the literature of the mobility of H b variants on this method. Methods A total of 383 Hb variants were studied on the Bio‐Rad Variant ™ Classic HPLC instrument. Hb variant identification used a number of methods, including confirmation by DNA sequencing in at least one case for all alpha and beta chain H b variants. Results Retention time data and the number of occurrences of each H b variant were obtained. This showed that rare H b variants can have similar retention times to the five most common alpha or beta chain H b variants. Conclusion HPLC is a very powerful tool in the evaluation of H b variants, particularly when combined with other methods. However, it should not be used as a stand‐alone method for definitive identification of H b variants.