[Routine determination of HB A1 by micro-column technique (author's transl)]

For routine determination of glycosilated hemoglobin (HB A1) it is necessary to have a quick and easy assay which allows analysis of a greater number of samples. Taking into consideration known methods for hemoglobin chromatography a micro-column technique has been developed, which separates the glycosilated hemoglobins AIa+b+c from the main hemoglobin fraction HB AII. Micro-columns (7.0 x 1.3 cm) are filled up to a bed-height of 3 cm with Bio-Rex 70. Hemoglobin is given to these in hemolysate from (1,8 mg/200 microliter) and separated with two buffers of different ionic-strength and different pH-values into two fractions HB A1 and HB AII. With exact standardisation of the elution volumes (18 ml), the elution temperature (21.5 degrees C) and pH-value (6.74) of the first elution buffer a high reproducibility of the results is possible (Intraassay-VK: 2.38%; Interassay-VK: 3.68%). The optical density of the hemoglobin fractions is read for cyanhemoglobin at 413 nm or 415 nm or for cyanmethemoglobin at 419 nm. The HB A1-concentration is given as a percentage of the total hemoglobin. This method enables the determination of HB A1-value of 40 blood samples in triplicate within four hours. The normal value derived from 25 healthy and normal weight volunteers is 6.88 +/- 0.41% (mean +/- S.D.), from which an upper normal range of 7.7% has been calculated. In 103 patients with unimpaired glucose tolerance values were observed in similar range: 6.45 +/- 0.61% (mean +/- S.D.). In 121 diabetic patients with varying metabolic control HB A1-values up to 20.58% were noted.