We have investigated the use of anion exchange high performance liquid chromatography (HPLC) for hemoglobin analysis. Several gradient elution programs were developed for optimal separation of hemoglobins in hemolysates derived from newborns and from individuals with hemoglobin disorders. The high resolution achieved, coupled with the ability to carry out chromatographic analysis in an unattended mode including automatic quantitation of the separated hemoglobins indicate that this technique could be quite useful in meeting the need for efficient and accurate diagnosis of hemoglobin disorders.