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Indian J Med Res. 2019 Jul;150(1):62-66. doi: 10.4103/ijmr.IJMR_1787_17.

Non-invasive prenatal rhesus D genotyping using cell-free foetal DNA.

Author information

Department of Obstetrics & Gynaecology, Postgraduate Institute of Medical Education & Research, Chandigarh, India.
Department of Experimental Medicine & Biotechnology, Postgraduate Institute of Medical Education & Research, Chandigarh, India.


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.


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.


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.


Alloimmunized; cell-free foetal DNA; exon; maternal plasma; non-invasive prenatal diagnosis; rhesus D genotyping

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