A2 Test: A Simple Method for the Screening of Beta-Thalassemia Trait

Abstract

Rationale Hb A is normally found 3% in in human blood samples. Its propoportion increases in B-thalassemia trait and dramatically increases in the severe B-thalassemia trait cases. There are more than 30 types of B-globin gene mutation which cause B-thalassemia trait in Thailand. But the proportions of only few are necessary to be determined by an appropriate method in order to control and prevent the severe homozygous B-thalassemia in the Thai population. Method: The A2 test is a method developed from microcolumn DEAE sephadex A 50 chromatography using a 3.5 mL disposable syringe as a column and a 10 mL one as a reservoir for the measurement of Hb A. The CV and comelation coeficients (r) of the A2 test were compared to automated HPLC and LPLC methods. The method was applied for determination of Hb A, in blood samples of normal and severe B-thalassemia trait cases Results: The CV of A2 test method was 7.9% whereas automated HPLC and LPLC methods were 18% and 24%, respectively. The correlation coeficient (r) obtained from the determination of Hb A by A2 test and automated HPLC in 300 pregnant women and by A2 test and automated LPLC in 149 healthy volunteers was all found to be 0.99. The average HbA in 74 B-thalassemia trait cases determined by A2 test was 6.0810:76%. The lowest value was 4.3% and the highest was 7.5%. Four hundred forty nine blood samples determined by A2 test were classified into 3 groups ie., three hundred eighty five non B-thalassemia trait cases with average HbA of 2.2.010.4% (1.7-3.7%), 22 B-thalassemia trait cases with average Hb A of 5:4106% (4.5-6.7%) and 39 Hb cases with over 10% of Hb A, Finally, 6,448 pregnant women were screened by Hb A value of 4.1-8.0%, 21 couples were found to be at risk. Five cases in those couples were found to be homozygous B-thalassemia and the Hb A, of the parents were between 52-7.3%. Discussion and Summary: The A2 test is a simple method with a low CV for the determination of Hb A2. The results correspond with the automated HPLC and LPLC methods. It can be conducted in general laboratories at low cost. An Hb A2 value of between 4.3-8.0% indicated a severe B-thalassemia trait. It is therefore considered to be an alternative method for diagnosing severe B-thalassemia trait, which can help control and prevent of the homozygous B-thalassemia in Thailand.