Rapid method for eliminating labile glycosylated hemoglobin from the assay for hemoglobin A1

The irreversible formation of stable glycosylated hemoglobin proceeds through a labile intermediate that is indistinguishable, by most methods, from the stable glycosylated product. The inclusion of the labile intermediate, which changes acutely with acute blood glucose changes, detracts from the utility of the assay as an index of chronic glucose concentration. We have developed a rapid, reliable chemical method for eliminating the labile intermediate: a 30-min incubation of whole-blood samples with semi-carbazide (30 mmol/L) and aniline (12 mmol/L) at pH 5 and 38 degrees C. The semicarbazide serves as a glucose trap; the transfer of glucose from the labile glycosylated hemoglobin to the semicarbazide is catalyzed by the acidic pH and aniline. This treatment is effective in the three most commonly used assays: "high-performance" liquid chromatography, electrophoresis, and a minicolumn kit.