ACCURACY, PRECISION, AND STABILITY IN MEASUREMENT OF HEMOGLOBIN-A1C BY HIGH-PERFORMANCE CATION-EXCHANGE CHROMATOGRAPHY

Abstract

A high-performance liquid-chromatographic method for determination of HbA1c with use of a carboxylate cation-exchange column, ethanolic mobile phases and a hemolysate-storage reagent that stabilizes a sample for up to 5 wk at 4.degree. C. Temperature control and the addition of ethanol to the phosphate step-gradient increase precision by stabilizing peak shape and column activity. Lengthy equilibration of the column between samples is not necessary, because the pKa of the cation-exchange resin is decreased from 6.10 in aqueous media to 5.59 in ethanol/water (5/95 by vol) at 25.degree. C. Interfering heme compounds, distinct from HbA1d or HbA1e, are identified that elute with the glycosylated Hb and developed a method of correcting for their presence. The day-to-day coefficient of variation was 3.5%, and the reference interval for a mixed group of 20 healthy adults between the ages of 20 and 56 was 3.2-5.2% HbA1c. [Glycosylated Hb is of interest in the study of its association with diabetes mellitus.].