Rapid detection of α-thalassaemia alleles of -- SEA /, -α 3.7 / and -α 4.2 / using a dual labelling, self-quenching hybridization probe/melting curve analysis

Objectives The aim of the study was to set up an alternative automatic molecular diagnostic method for deletional α-thalassaemia mutations without gel electrophoresis. Methods Based on the sequence variation within the two Z boxes and melting curve analysis of dually labelled probes, a real-time PCR assay was developed and validated for the rapid detection of major α-genotypes (-- SEA /αα, -- SEA /-α 3.7 , -- SEA /-α 4.2 , -- SEA /-- SEA , -α 3.7 /-α 3.7 and -α 4.2 /-α 4.2 ). Results Samples with the -α 3.7 /-α 3.7 , -α 4.2 /-α 4.2 , -- SEA /αα, -- SEA /-α 3.7 , -- SEA /-α 4.2 , and -- SEA /-- SEA genotypes could be clearly distinguished. The accuracy of this technique for these samples was 100% sensitivity and specificity. Conclusion This technique is rapid and reliable, demonstrating feasibility for use in large-scale population screening and prenatal diagnosis of deletional Hb H disease and Hb Bart's hydrops fetalis.