SPECIFICITY OF HEMOGLOBIN A(1C) MEASUREMENT BY CATION-EXCHANGE LIQUID-CHROMATOGRAPHY - EVALUATION OF A MONO S COLUMN METHOD

A method was developed for evaluating the specificity of the ion excha nge chromatographic HbA(1c) method by Pharmacia which uses a Mono S co lumn. To investigate hemoglobin fractions potentially interfering in H bA(1c) analysis, the chromatographic resolution was enhanced and a pea k integration program was used which enables the quantitation of overl apping and even shoulder-like peaks. Hemoglobin was incubated with glu cose in vitro and the chromatograms were analyzed before and after inc ubation. Five minor peaks were detected, close to hemoglobin A(1c), wh ich could not be conclusively identified as hemoglobins previously des cribed. Foul of these peaks, the stable ones, were included in the fra ction of the chromatogram to be measured as HbA(1c) by the routine met hod. In practice, these peaks comprise 20%-35% of the routine HbA(1c) result in diabetic patients and 30%-45% in nondiabetic subjects. Three of these minor peaks were non-dependent on glucose. The chromatograph y and the peak integration method described can also be used to study hemoglobin adducts other than glycohemoglobin.