Levels of glycated haemoglobin A – HbA1c in nondiabetic carriers of haemoglobin O‐Arab

Sir, the glycated haemoglobin (GHb) test is a valid index of mean glycaemia during the preceding 2–3 months, valuable for monitoring glycaemic control in diabetic patients. Abnormal haemoglobins influence the GHb analysis (Bry, Ghen & Sacks, 2001). The effect of haemoglobinopathies on GHb measurements is highly method-dependent (Schnedl et al., 2000). The presence of a haemoglobin (Hb) variant may erroneously lead to underestimated GHb measurements by high-performance liquid chromatography (HPLC) (Camargo & Gross, 2004). In these cases, the interpretation of GHb varies according to the variant which is present and to the assay method (Schnedl et al., 2001; Sacks et al., 2002). The presence of Hb variants should be suspected when HbA1c results fail to correlate with clinical data (Florkowski et al., 2003). Clinical laboratories should be aware of limitations of their HbA1c assay methods (Schnedl et al., 2004) and have suitable policies in place to ensure patients to receive appropriate diagnostic tests (Reynolds et al., 2004). Sacks et al. (2002) recommend that the interference of a Hb variant with HbA1c may be subtle in the reference range but may increase steadily with an increase in the HbA1c. Most ion-exchange HPLC methods allow detection of most Hb variants (Lahousen et al., 2002). The analysis in the ARKRAY , HA-8160 analyzer (ARKRAY Europe, B.V., Amstelveen, The Netherlands) on b-thalassaemia screening mode, is certified by the National Glycohemoglobin Standardization Program, but the determination of HbA1c is impossible, when early migrating Hb variants like Hb Graz and Hb Sherwood Forest are present, causing an additional peak that overlaps HbA1c (Schnedl et al., 2005). In the presence of a late migrating Hb variants like HbO Padova and HbD, the obtained HbA1c results are clinically too low (Schnedl et al., 2005). Schnedl et al. (2000) propose that every laboratory must establish and secure its own assay method for HbA1c based on the ethnic origin of the population and underline the importance of visual inspection of ion-exchange chromatograms to detect abnormalities caused by the Hb variants (Schnedl et al., 2004). Haemoglobin O-Arab (alpha2beta2121GlufiLys), is characterized by a more positive net charge (Papadopoulos et al., 2003) and is very frequent in our region (5% in Greek Pomaks) (Papadopoulos et al., 2006). Thus, we studied the HbA1c levels determined by HPLC in nondiabetic HbO-Arab carriers to estimate the reference range for this population and thus enable accurate monitoring of diabetic HbO-Arab carriers. For that purpose, a cohort of 26 nondiabetic (oral glucose tolerance test, World Health Organization Criteria) adult individuals with HbO-Arab trait (11 men, 15 women, aged 18–52 years, reticulocyte count (RET#) and high fluorescence reticulocyte proportion (HFR) within our laboratory reference range, assigned as group A) were selected to enter the study. Additionally, a cohort of 61 nondiabetic adult individuals without haemoglobinopathy (28 men, 33 women, aged 18–58 years, presenting normal RET# and HFR, assigned as group B) were selected as a normal–control group. Additionally, one diabetic HbO-Arab carrier, one nondiabetic HbS carrier and one diabetic HbO-Arab homozygote were analysed on ADAMS A1c HA-8160 (ARKRAY Europe, B.V., Amstelveen, The Netherlands) for HbA1c measurement to study the chromatogram. Specimens were analysed on a Sysmex XT-2000i (Sysmex Corporation, Kobe, Japan) instrument in the reticulocyte channel for complete blood cell count within the first hour from collection. The separation of haemoglobin fractions was performed by HPLC (G7 b-Thalassemia Mode, HLC 723G7, TOSOH); Figure 1a. The determinations of HbA1c were performed within 5 days from collection (samples were stored at 4 C) by automated reversed-phase cation-exchange HPLC on an ADAMS A1c HA-8160, ARKRAY, Inc. analyzer, in the whole blood measurement mode. Measurements made with HA-8160 are accurate because labile HbA1c, carbamilated Hb and acetylated Hb are eluted separately from stable HbA1c peak. Descriptive statistics refer to mean and their SD values. Group comparisons were performed by Mann–Whitney U-test and v-test. The reference range is calculated between the 2.5 and 97.5 percentiles of the values. Values of P < 0.05 were considered to indicate statistical significance. LETTER TO THE EDITOR INTERNATIONAL JOURNAL OF LABORATORY HEMATOLOGY