Loop-mediated isothermal amplification (LAMP) colorimetric phenol red assay for rapid identification of α0-thalassemia: Application to population screening and prenatal diagnosis

• Published in: PloS one Vol. 17; no. 4; p. e0267832
• Main Authors: Jomoui, Wittaya, Srivorakun, Hataichanok, Chansai, Siriyakorn, Fucharoen, Supan
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 2022; Public Library of Science (PLoS)
• Subjects: alpha-Thalassemia - diagnosis; alpha-Thalassemia - genetics; Assaying; Biology and Life Sciences; Blood diseases; Colorimetry; Cost analysis; Deletion; Deoxyribonucleic acid; Diagnosis; DNA; Edema; Female; Fetuses; Gene amplification; Genetic counseling; Genotypes; Humans; Hydrops fetalis; Laboratories; Medical diagnosis; Medicine and Health Sciences; Molecular Diagnostic Techniques; Nucleic Acid Amplification Techniques; Nucleotide sequence; Phenols; Phenolsulfonphthalein; Pregnancy; Prenatal diagnosis; Prenatal Diagnosis - methods; R&D; Research & development; Research and Analysis Methods; Screening; Sensitivity; Thalassemia; Thailand
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0267832

Identification of α0-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. Three LAMP colorimetric assays specific for α0-thalassemia (SEA deletion), α0-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 α0-thalassemia (SEA deletion) was done on 33 fetuses at risk of having Hb Bart's hydrops fetalis syndrome. The LAMP colorimetric assays for α0-thalassemia (SEA and THAI deletions) could be clearly interpreted by naked eyes. The assay for α0-thalassemia (SEA deletion) showed a 100% (62/62 x 100) sensitivity and 98.2% (274/279 x 100) specificity whereas, that of the α0-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 α0-thalassemia (SEA deletion) in 33 fetuses as compared to the conventional PCR analysis. 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.