Haemoglobin A₂′ and its significance in β thalassaemia diagnosis

Abstract

We have studied, haemoglobin A(2)' (A(2) prime), a delta chain variant haemoglobin occurring in a small percentage of individuals of African ancestry. In heterozygotes, the percentage of haemoglobin A(2)' was found to be slightly lower then the percentage of haemoglobin A(2), suggesting that the variant delta chain is synthesized at a reduced rate. When quantifying haemoglobin A(2) for the diagnosis of beta thalassaemia heterozygosity, it is essential to add together the A(2) and A(2)' to give 'total haemoglobin A(2)'. However, it not necessary to use a different reference range for total haemoglobin A(2) in A(2)' heterozygotes. When using microcolumn chromatography, A(2)' was found to be measured with A(2). On the high-performance liquid chromatography instrument studied, haemoglobin A(2)' fell in the haemoglobin S window but its retention time differed from that of haemoglobin S.

An insight into the suspected HbA2' cases detected by high performance liquid chromatography in Pakistan

“…Retention time of a hemoglobin variant can change with different lots of reagents (including the ion-exchange column)[15]. The levels of HbA2 were found to be higher than those of HbA2' in all cases similar to other published reports [2,6,14,16]. All cases with suspected HbA2' of >2% fell under the category of β-thalassaemia minor if S-window peak was taken into account, which was also consistent with previously published studies [10,14].…”

“…HPLC has been shown to be a reliable instrument for the detection of HbA2' in many parts of the world [6,13,14]. We found that the trends (retention time etc.)…”

“…Recent literature review raised the possibility of them representing HbA2' [6,14]. Although HbA2' has never been reported in Pakistan, the possibility of a rare silent mutation existing in the country without being reported was very likely owing to the fact that limited research occurs here.…”

“…Since then it has been reported with increasing frequency among blacks with prevalence being 2.16% in Mauritania (Northwestern Africa) [2], 2.7% in Dogon (Mali) [3] and 18% in Herero (South Africa) [4]. Recently, this hemoglobinopathy has been observed in two Brazilian Caucasian women [5], and in white British [6]. …”

“…Alkaline electrophoresis has limited resolution in this regard [12] and though detection by iso-electric focusing is easy, it lacks precise quantification [12]. On the other hand, microcolumn chromatography can accurately measure total HbA2 level without identifying HbA2'[6]. In contrast to these diagnostic tools, high performance liquid chromatography (HPLC) not only senses HbA2' as it elutes in the 'S' window (retention time of 4.59 ± 0.03 minutes) but also quantifies it which is usually in proportion of 1.2% of total hemoglobin [13].…”