Suwannakhon, NarutchalaHemvuthiphan, JittapholPangeson, TanapatMahingsa, KhwanruedeePingyod, AruneeBumrungpakdee, WanwipaSanguansermsri, Torpong2023-08-192023-08-192023-05Suwannakhon Narutchala, Hemvuthiphan Jittaphol, Pangeson Tanapat, Mahingsa Khwanruedee, Pingyod Arunee, Bumrungpakdee Wanwipa, Sanguansermsri Torpong. Non-invasive prenatal screening & diagnosis of ?-thalassaemia in an affected foetus. Indian Journal of Medical Research. 2023 May; 157(5): 447-4520971-5916http://imsear.searo.who.int/handle/123456789/223551Background & objectives: Non-invasive prenatal testing (NIPT) of maternally inherited alleles of ?-thalassaemia (MIB) remains to be a challenge. Furthermore, current techniques are not available for use as routine tests. NIPT for ?-thalassaemia disease was developed by using a specific droplet digital polymerase chain reaction (ddPCR) assay to analyze the cell-free foetal DNA (cffDNA) derived from maternal plasma. Methods: Pregnant women and their spouses who are at risk of bearing an offspring with ?-thalassaemia disease from common MIB mutations (CD 41/42-TCTT, CD17A>T, IVS1-1G>T and CD26G>A) were enrolled. The ddPCR assay sets were constructed for each of the four mutations. All cell-free DNA samples were first screened for the paternally inherited ?-thalassaemia (PIB) mutation. The PIB-negative samples were considered as non-disease and were not further analyzed. For PIB-positive samples, DNA fragments of 50-300 base pairs in size were isolated and purified, and further analyzed for MIB mutation. The allelic ratio between the mutant and the wild-type was used to determine the presence of MIB in cffDNA. All cases underwent a prenatal diagnosis by amniocentesis for a definite diagnosis. Results: Forty two couples at risk were enrolled. Twenty two samples were positive for PIBs. Among these 22 samples, there were 10 cases with allelic ratio >1.0 (MIB positive). All foetuses with over-represented mutant alleles were further diagnosed with ?-thalassaemia disease; eight with compound heterozygous and two with homozygous mutations. The 20 PIB-negative and 12 MIB-negative foetuses were non-affected. Interpretation & conclusions: The results of this study suggest that NIPT utilizing the ddPCR assay can be effectively used for the screening and diagnosis of foetal ?-thalassaemia in at risk pregnancies.Droplet digital polymerase chain reaction assaynon-invasiveprenatal diagnosis?-thalassaemiaJournal ArticleIndiaDepartment of Biology, School of Science, University of PhayaoDivision of Obstetrics & Gynecology, Phayao HospitalDepartment of Biochemistry, School of Medical Science, University of PhayaoThalassaemia Unit, University of Phayao Hospital, University of Phayao, Phayao, Thailand