Determination of glycosylated hemoglobin by affinity electrophoresis on agarose gel

Abstract

We describe a method for electrophoretic determination of glycosylated hemoglobin (GHb) on agarose gel membrane. Clear separation of GHb from non-GHb is achieved after 30 min electrophoresis at 60 V using a sodium citrate buffer solution (34 mmol/1, pH 6.5) containing 0.6 g/1 dextran sulfate. The results obtained by this method correlate well with those by agar gel electrophoresis ( r = 0.98) and column chromatography ( r = 0.96). However, unlike column Chromatographic methods, GHb values obtained on agarose gel are not affected by fluctuations in ambient temperature (18–28°C), changes in pH (6.2–6.6) and ionic strength of buffer solution (26–40 mmol/1), or variant hemoglobin S and C. Also, electrophoresis on agarose gel membrane eliminates the lengthy preparative steps required for cellulose acetate electrophoresis. We conclude that agarose gel electrophoresis is a simple method for quantitative determination of GHb.