Multiplex recombinase polymerase amplification (RPA) assay for carrier detection and prenatal diagnosis of α0-thalassemia (SEA and THAI deletions).

Abstract

Hemoglobin Bart's hydrops fetalis is the most severe form of α-thalassemia, caused by homozygous α⁰-thalassemia, which is highly prevalent in Southeast Asian countries. Simple and rapid identification of α⁰-thalassemia carrier and prenatal diagnosis of Hemoglobin Bart's hydrops is essential in the region. We have developed a multiplex RPA assay for simple detection of α⁰-thalassemia (SEA and THAI deletions) and tested it in carrier detection (n = 125) and prenatal diagnosis of Hb Bart's hydrops fetalis syndrome (n = 30). The sensitivity, specificity, positive predictive value, and negative predictive value for detecting α⁰-thalassemia carriers were 100% for both SEA and THAI deletions. The assay correctly identified 42 carriers of α⁰-thalassemia (SEA deletion), 4 carriers of α⁰-thalassemia (THAI deletion), and 79 non-carriers. For prenatal diagnosis, the results of RPA revealed a 100% concordance (30/30) with the conventional gap-PCR analysis. The multiplex RPA assay is a rapid, reliable, and cost-effective method for diagnosing α⁰-thalassemia-related disorders in routine clinical settings. This assay has the potential to significantly contribute to the prevention and control of Hb Bart's hydrops fetalis in the region.