RESUMEN
Background & objectives: Non-invasive prenatal diagnosis (NIPD) of rhesus D (RHD) genotype using cell-free foetal DNA is extensively used in many developed countries. Studies on NIPD from India are scarce. The aim of the present study was to evaluate the performance of non-invasive foetal RHD genotyping by targeting exon 10 of the RHD gene using cell-free DNA. Methods: DNA was extracted from the maternal plasma of alloimmunized and non-alloimmunized women between 7 and 34 wk of gestation. RHD sequence was determined by quantitative real time polymerase chain reaction (PCR). Results were compared with RhD phenotype obtained from cord blood samples of neonates. Results: A total of 135 samples from RhD-negative pregnant women were collected. The foetal RHD status was conclusive in all 135 (100%) cases. The highest number of cases reported for RHD genotyping were from Punjab (38.5%) followed by Haryana (24.4%), Himachal Pradesh (17.0%) and Chandigarh Union Territory (13.3%). The non-invasive test correctly predicted the foetal RhD phenotype in 133 of 135 cases, making the accuracy of the test as 98.51 per cent [95% confidence interval (CI): 97.90-99.50%]. The overall sensitivity and specificity of the test were 99.18 per cent (95% CI: 95.52-99.98%) and 92.31 per cent (95% CI: 63.97-99.81%), respectively, with negative and positive predictive values of 99.80 per cent (95% CI: 94.85-99.87%) and 96.31 per cent (95% CI: 62.87-98.84%), respectively. Interpretation & conclusions: Non-invasive foetal RHD determination by single-exon quantitative PCR exhibited high accuracy and could be used in routine clinical practice after confirmatory studies are done.
RESUMEN
BACKGROUND: Genes for ABO and Rh phenotypes were recently identified. Although ABO genotyping don't find wide application in hospital transfusion services, it can play an important role in paternity and forensic investigation. In case of Rh system, however, DNA typing may find several practical applications such as prenatal determination of fetal Rho(D) genotype. METHODS: 64 blood samples for ABO genotyping were collected from blood donors (34 A, 30 B) and 18 samples for D genotyping (10 D+, 8 D-). To distinguish A, B and O alleles, we analyzed nucleotide positions 261 and 803 using polymerase chain reaction (PCR) and Restriction Fragment Length Polymorphism (RFLP). PCR products containing nucleotide position 261 were restricted with KpnI and BstEII. Rh genotyping was done by two sets of primers, one set for both RhD and RhCcEe gene amplification, and the other set for RhD only. RESULTS: The frequencies of ABO genotypes found in Korean blood donors were as follows: in the phenotype A group, AO=79% and AA= 21%; and in the phenotype B group, BO=93% and BB=7%. Of 18 blood samples for D genotytping, 10 were typed as RhD positive and 8 as RhD negative, showing full agreement with serological typing. CONCLUSION: ABO and D genotyping can be used when RBCs suitable for serological phenotyping are not available. Futhermore, these will be useful as a supplemental test to solve the problem of blood group typing caused by weak ABO and Rh phenotype.