Sensitivity of Fetal RhD Screening for Safe Guidance of Targeted Anti-D Immunoglobulin Prophylaxis: Prospective Cohort Study of a Nationwide Programme in the Netherlands

The risk ofmaternal alloimmunization due to RhD incompatibility has decreased with use of antenatal and postnatal anti-D immunoglobulin prophylaxis. The discovery of cell-free fetal (cff) DNA in maternal plasma during pregnancy and the feasibility of fetal RhD testing using this source of DNA make it possible to determine fetal RhD type and to restrict the use of antenatal anti-D immunoglobulin to only those RhD-negative women carrying an RhD-positive fetus. Variable and low amount of fetal DNA present in maternal plasma and the genetic variation of RHD alleles in the fetus or mother complicate such noninvasive testing. This study was done to evaluate the efficacy of noninvasive fetal RHD testing in week 27 of pregnancy to restrict use of antenatal and postnatal anti-D immunoglobulin use to RhD-negative women carrying an RhD-positive fetus. Duplex real-time polymerase chain reaction analysis was carried out for RHD exon 5 and RHD exon 7 in triplicate on cellfreeDNA isolated frommaternal plasma. Cord blood serologywas used as the reference standard as it is the best test available for determination of neonatal RhD status. The sensitivity for detection of fetal RHD was 99.94% (95% confidence interval [CI], 99.89%-99.97%), and specificity was 97.74% (95% CI, 97.43%-98.02%). Cord blood serology showed 9 falsenegative fetal RHD test results (0.03%; 95% CI, 0.02%-0.07%) and 225 false-positive fetal RHD test results (0.87%; 95% CI, 0.76%-1.00%). The overall negative predictive value was 99.91%(95% CI, 99.82%-99.95%), and the positive predictive value was 98.60% (95% CI, 98.40%-98.77%). The results demonstrate that fetal RHD assay performed at week 27 of pregnancy as part of an antenatal screening program is reliable and effective and can be used to restrict the use of both antenatal and postnatal anti-D immunoglobulin in RhD-negative pregnant women