Loop-mediated isothermal amplification (LAMP) colorimetric phenol red assay for rapid identification of α(0)-thalassemia: Application to population screening and prenatal diagnosis

BACKGROUND: Identification of α(0)-thalassemia (SEA and THAI deletions) is essential in preventing and controlling of severe thalassemia diseases. We have developed the LAMP colorimetric assays for the detection of these two thalassemia defects and validated them in population screening and prenatal diagnosis. METHODS: Three LAMP colorimetric assays specific for α(0)-thalassemia (SEA deletion), α(0)-thalassemia (THAI deletion) and normal DNA sequence were developed. These assays were validated on 341 subjects who had initial thalassemia screening positive and various thalassemia genotypes. Prenatal diagnosis of α(0)-thalassemia (SEA deletion) was done on 33 fetuses at risk of having Hb Bart’s hydrops fetalis syndrome. RESULTS: The LAMP colorimetric assays for α(0)-thalassemia (SEA and THAI deletions) could be clearly interpreted by naked eyes. The assay for α(0)-thalassemia (SEA deletion) showed a 100% (62/62 x 100) sensitivity and 98.2% (274/279 x 100) specificity whereas, that of the α(0)-thalassemia (THAI deletion) showed 100% (1/1 x 100) sensitivity and 99.7% (339/340 x 100) specificity. We obtained a 100% concordant prenatal diagnosis results using LAMP assays of α(0)-thalassemia (SEA deletion) in 33 fetuses as compared to the conventional PCR analysis. CONCLUSIONS: The LAMP colorimetric assays developed are simple, rapid, and do not require sophisticated equipment. Inclusion of the LAMP tests in the existing screening protocol significantly reduce the screening cost and the molecular analysis workload, which should prove useful in the prevention and control program of hemoglobinopathies in the region.