Cell‐free fetalDNAand fetal blood group genotyping: non‐invasive prenatal testing

Abstract

In transfusion medicine and clinical immunology, cell-free fetal DNA (cffDNA) is analysed from maternal plasma of pregnant women to predict fetal blood groups with the purpose of (1) assessing the risk of haemolytic disease of the fetus and newborn (HDFN) in immunized women and (2) guiding targeted Rh prophylaxis in non-immunized RhD-negative women. National programmes for guiding prophylaxis are now implemented in around 6-7 European countries; assay accuracy is very high, with sensitivities of 99Á9%. Sensitivity is challenged by low quantities of cffDNA, especially in early pregnancy. Specificity is challenged by the polymorphic Rh blood group system, where careful attention is needed to navigate among the many RHD variants that may complicate cffDNA analysis and interpretation of results, especially in populations with mixed ethnicities. However, fetal RHD testing is feasible when implemented with careful attention to these issues. The success of predicting fetal RhD and its successful clinical implementation should encourage widespread implementation. For blood groups that are determined by SNPs, such as KEL or Rhc, novel techniques such as next-generation sequencing and droplet digital PCR are now providing accurate non-invasive prediction of these fetal blood groups. Future work on non-invasive prenatal testing of fetal blood groups determined by SNPs may consolidate the application for cell-free DNA testing for such targets. At ISBT, the newly formed cfDNA subgroup of the Red Cell Immunogenetics and Blood Group Terminology Working Party will work to facilitate clinical applications, implementation and evaluation of cell-free DNA testing.

Diagnostic performance of the noninvasive prenatal FetoGnost RhD assay for the prediction of the fetal RhD blood group status

“…However, the specificity of an assay based on replicate testing of RHD exon 10 with 95.2% was lower than in the previous studies [32]. In our performance evaluation study, the sensitivity of 99.93% and specificity of 99.61% were as high as sensitivities and specificities determined in the second trimester with largescale screening studies [14,21,29,33]. Notably, our single false-negative result was observed at the beginning of the second trimester.…”

“…Since the feasibility of targeted antenatal RAADP based on the result from a NIPT for the prediction of the fetal RhD status has been proposed by Dennis Lo's group and a Dutch group in 1998 it took about 12 years until the first nationwide program for targeted RAADP was implemented in Denmark [11][12][13]. Until today many validation studies have been published and extensively reviewed which revealed an excellent diagnostic accuracy of NIPT for RhD [14][15][16][17][18][19][20][21]. It has to be stressed in the given context that only false-negative NIPT RhD results may have relevant consequences (i.e., increased risk for anti-D alloimmunisation).…”

Diagnostic performance of the noninvasive prenatal FetoGnost RhD assay for the prediction of the fetal RhD blood group status

“…However, the speci city of an assay based on replicate testing of RHD exon 10 with 95.2% was lower than in the previous studies [32]. In our performance evaluation study the sensitivity of 99.93% and speci city of 99.61% were as high as sensitivities and speci cities determined in the second trimester with large-scale screening studies [14,21,29,33]. Notably, our single false negative result was observed at the beginning of the second trimester.…”

“…Since the feasibility of targeted antenatal RAADP based on the result from a NIPT for the prediction of the fetal RhD status has been proposed by Dennis Lo's group and a Dutch group in 1998 it took about 12 years until the rst nationwide program for targeted RAADP was implemented in Denmark [11][12][13]. Until today many validation studies have been published and extensively reviewed which revealed an excellent diagnostic accuracy of NIPT for RhD [14][15][16][17][18][19][20][21]. It has to be stressed in the given context that only false negative NIPT RhD results may have relevant consequences (i.e.…”

Impact of long‐term storage of plasma and cell‐free DNA on measured DNA quantity and fetal fraction

“…Non‐invasive testing of cell‐free DNA (cfDNA) has become highly relevant in clinical medicine, with clinical applications in prenatal diagnosis [1,2], cancer [3] and transplantation [4]. In transfusion medicine and clinical immunology, non‐invasive testing of cell‐free fetal DNA (cffDNA) is used to predict fetal blood groups [5], including clinical applications for fetal RHD genotyping [6,7] and genotyping of fetal non‐RhD antigens [8‐10], to assist in pregnancy care where blood group incompatibility may have clinical consequences [11].…”