Advertisement

Multiplexed analysis of circulating cell-free fetal nucleic acids for noninvasive prenatal diagnostic RHD testing

Published:November 19, 2010DOI:https://doi.org/10.1016/j.ajog.2010.09.028

      Objective

      The objective of the study was the evaluation of a novel multiplex assay to detect fetal Rh blood group D-antigen gene (RHD) loci in maternal plasma from RhD-negative, pregnant women.

      Study Design

      An RHD genotyping assay was designed to detect exons 4, 5, 7, and 10 and RHDΨ (pseudogene) of the RHD gene along with a Y chromosome–specific assay and a generic polymerase chain reaction amplification control. Plasma samples from 150 RhD-negative pregnant women were assayed for fetal RHD genotype using the MassARRAY system.

      Results

      The fetal RHD status of 148 of 150 samples (98.7%) was correctly classified; 86 (57.3%) and 62 (41.3%) were positive and negative, respectively.

      Conclusion

      This study demonstrates that noninvasive prenatal diagnostics with a single-reaction multiplexed assay is a viable path toward routine characterization of fetal RHD genotypes using circulating cell-free fetal DNA in maternal plasma on the MassARRAY system and is perhaps preferable to serologic testing as currently used clinically.

      Key words

      To read this article in full you will need to make a payment

      Purchase one-time access:

      Academic & Personal: 24 hour online accessCorporate R&D Professionals: 24 hour online access
      One-time access price info
      • For academic or personal research use, select 'Academic and Personal'
      • For corporate R&D use, select 'Corporate R&D Professionals'

      Subscribe:

      Subscribe to American Journal of Obstetrics & Gynecology
      Already a print subscriber? Claim online access
      Already an online subscriber? Sign in
      Institutional Access: Sign in to ScienceDirect

      References

        • Lo Y.M.
        • Corbetta N.
        • Chamberlain P.F.
        • et al.
        Presence of fetal DNA in maternal plasma and serum.
        Lancet (British edition). 1997; 350: 485-487
        • Lo Y.M.D.
        • Chiu R.W.K.
        Prenatal diagnosis: progress through plasma nucleic acids.
        Nat Rev Genet. 2007; 8: 71-77
        • Lo Y.M.
        • Lun F.M.
        • Chan K.C.
        • et al.
        Digital PCR for the molecular detection of fetal chromosomal aneuploidy.
        Proc Natl Acad Sci USA. 2007; 104: 13116-13121
        • Lo Y.M.
        • Tsui N.B.
        • Chiu R.W.
        • et al.
        Plasma placental RNA allelic ratio permits noninvasive prenatal chromosomal aneuploidy detection.
        Nat Med. 2007; 13: 218-223
        • Lo Y.M.
        • Chiu R.W.K.
        Noninvasive prenatal diagnosis of fetal chromosomal aneuploidies by maternal plasma nucleic acid analysis.
        Clin Chem. 2008; 54: 461-466
        • Lo Y.M.
        Noninvasive prenatal detection of fetal chromosomal aneuploidies by maternal plasma nucleic acid analysis: a review of the current state of the art.
        BJOG. 2009; 116: 152-157
        • Chiu R.W.
        • Chan K.C.
        • Gao Y.
        • et al.
        Noninvasive prenatal diagnosis of fetal chromosomal aneuploidy by massively parallel genomic sequencing of DNA in maternal plasma.
        Proc Natl Acad Sci USA. 2008; 105: 20458-20463
        • Fan H.C.
        • Blumenfeld Y.J.
        • Chitkara U.
        • Hudgins L.
        • Quake S.R.
        Noninvasive diagnosis of fetal aneuploidy by shotgun sequencing DNA from maternal blood.
        Proc Natl Acad Sci USA. 2008; 105: 16266-16271
        • Bianchi D.W.
        • Avent N.D.
        • Costa J.M.
        • van der Schoot C.E.
        Noninvasive prenatal diagnosis of fetal rhesus D: ready for Prime(r) time.
        Obstet Gynecol. 2005; 106: 841-844
        • Finning K.
        • Martin P.
        • Daniels G.
        A clinical service in the UK to predict fetal Rh (Rhesus) D blood group using free fetal DNA in maternal plasma.
        Ann N Y Acad Sci. 2004; 1022: 119-123
        • Van der Schoot C.E.
        • Soussan A.A.
        • Koelewijn J.
        • Bonsel G.
        • Paget-Christiaens L.G.
        • de Haas M.
        Non-invasive antenatal RHD typing.
        Transfus Clin Biol. 2006; 13: 53-57
        • Singleton B.K.
        • Green C.A.
        • Avent N.D.
        The presence of an RHD pseudogene containing a 37 base pair duplication and a nonsense mutation in Africans with the Rh D-negative blood group phenotype.
        Blood. 2000; 95: 12
        • Grill S.
        • Banzola I.
        • Li Y.
        • et al.
        High throughput non-invasive determination of foetal Rhesus D status using automated extraction of cell-free foetal DNA in maternal plasma and mass spectrometry.
        Arch Gynecol Obstet. 2009; 279: 533-537
        • Muller S.P.
        • Bartels I.
        • Stein W.
        • et al.
        The determination of the fetal D status from maternal plasma for decision making on Rh prophylaxis is feasible.
        Transfusion. 2008; 48: 2292-2301
        • Legler T.J.
        • Heermann K.H.
        • Liu Z.
        • Soussan A.A.
        • van der Schoot C.E.
        Fetal DNA: strategies for optimal recovery.
        Methods Mol Biol. 2008; 444: 209-218
        • Legler T.J.
        • Liu Z.
        • Mavrou A.
        • et al.
        Workshop report on the extraction of foetal DNA from maternal plasma.
        Prenat Diagn. 2007; 27: 824-829
        • Oeth P.
        • del Mistro G.
        • Marnellos G.
        • Shi T.
        • van den Boom D.
        Qualitative and quantitative genotyping using single base primer extension coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MassARRAY).
        Methods Mol Biol. 2009; 578: 307-343
        • Ding C.
        Maldi-TOF mass spectrometry for analyzing cell-free fetal DNA in maternal plasma.
        Methods Mol Biol. 2008; 444: 253-267
        • Li Y.
        • Finning K.
        • Daniels G.
        • Hahn S.
        • Zhong X.
        • Holzgreve W.
        Noninvasive genotyping fetal Kell blood group (KEL1) using cell-free fetal DNA in maternal plasma by MALDI-TOF mass spectrometry.
        Prenat Diagn. 2008; 28: 203-208
        • Li Y.
        • Wenzel F.
        • Holzgreve W.
        • Hahn S.
        Genotyping fetal paternally inherited SNPs by MALDI-TOF MS using cell-free fetal DNA in maternal plasma: influence of size fractionation.
        Electrophoresis. 2006; 27: 3889-3896
        • Scheffer P.G.
        • van der Schoot C.E.
        • Page-Christiaens G.C.
        • Bossers B.
        • van Erp F.
        • de Haas M.
        Reliability of fetal sex determination using maternal plasma.
        Obstet Gynecol. 2010; 115: 117-126