A Rare Hemoglobin Variant Detected for the First Time in Türkiye (Hb Iraq-Halabja): Evaluation of the Effect of Variant Hemoglobins on HbA1c Methods.

Abstract

The commonly used methods for HbA1c measurement are cation-exchange high performance chromatography (CE-HPLC), immunologic method, capillary electrophoresis and boronate affinity HPLC. Hb-variants can reduce the reliability of HbA1c measurements. We aimed to emphasize the importance of step-by-step solutions to the difficulties encountered in HbA1c measurement methods due to Hb-variants. We also aimed to evaluate the advantages and disadvantages of different methods used in HbA1c analysis with the example of the Hb-Iraq-Halabja variant detected for the first time in Türkiye. HbA1c level could not be measured by CE-HPLC (Adams HA-8180V analyzer, Arkray, Japan) and a peak-tailing signal was detected indicating an abnormality between stable HbA1c and HbA0 peaks (concurrent glucose level was 8.55mmol/L). In the second step, HbA1c was able to be measured by immunologic method and boronate affinity method and were found to be 5.87% and 5.90%, respectively (estimated average glucose equivalent 6.66mmol/L). In the last step, genetic analysis was performed due to suspicion of Hb variant and the very rare Hb Iraq-Halabja variant was detected. After genetic verification, HbA1c test was repeated with a different CE-HPLC (HLC-723 G11, T osoh, Tokyo, Japan) and a peak indicating variant Hb was detected between stable HbA1c and HbA0. A remarkable finding was that, unlike the previous CE-HPLC (Arkray) result, HbA1c could be measured as 3.20%. In the concurrent measurement performed on the boronate affinity HPLC (Premier Hb9210, Trinity Biotech, County Wicklow, Ireland), HbA1c result was found to be 5.40% (concurrent glucose 5.22 mmol/L). In addition, concurrent fructosamine serum value was found to be 210 μmol/L (estimated mean glucose equivalent 4.88 mmol/L). The patient's laboratory tests were generally within normal limits, and iron deficiency, hemolytic anemia, and B12-folate deficiency were excluded. The glucose bounding area of hemoglobin is generally preserved and is not affected by common Hb-variants. Boronate affinity and immunologic method (these two methods target glucose bounding areas) that give HbA1c results consistent with the patient's fasting blood glucose and fructosamine results. However, the CE-HPLC method has been observed to either fail to measure HbA1c or to measure falsely low HbA1c due to overlapping peaks of Hb variants.