title: Droplet digital PCR, a new approach to analyze fetal DNA from maternal blood: application to the determination of fetal RHD genotype.
abstract: The discovery of free fetal DNA in the maternal circulation has inaugurated the era of non-invasive prenatal diagnosis. The latter has the advantage of avoiding the use of conventional obstetric procedures, such as chorionic villus sampling or aspiration of amniotic fluid, thus limiting the risks of miscarriage they induce. However, as free fetal DNA accounts for about 10% of cell-free DNA in maternal plasma, the presence of ambient maternal DNA can make it difficult to detect fetal alleles of paternal origin. Digital Droplet PCR (ddPCR) is a very sensitive method derived from quantitative real-time PCR (qPCR) for the detection of rare alleles and their absolute quantification by removing the necessity of standards. Here we show that this new technology can be applied in routine prenatal fetal RHD genotyping from maternal blood. In conclusion, the use of quantitative properties of digital PCR, in terms of accuracy, sensitivity and specificity, allows one to consider extending the applications of this new technology in non-invasive prenatal diagnosis of many diseases such as autosomal monogenic diseases, either dominant or recessive.
authors.affiliation: Service de biochimie et génétique moléculaire, HUPC Hôpital Cochin, Paris, France.
authors.forename: Lucie
authors.initials: L
authors.lastname: Orhant
authors.affiliation: Service de biochimie et génétique moléculaire, HUPC Hôpital Cochin, Paris, France.
authors.forename: Sophie
authors.initials: S
authors.lastname: Rondeau
authors.affiliation: Service de biochimie et génétique moléculaire, HUPC Hôpital Cochin, Paris, France.
authors.forename: Aurélie
authors.initials: A
authors.lastname: Vasson
authors.affiliation: Maternité Cochin-Port Royal, HUPC Hôpital Cochin, Paris, France.
authors.forename: Olivia
authors.initials: O
authors.lastname: Anselem
authors.affiliation: Maternité Cochin-Port Royal, HUPC Hôpital Cochin, Paris, France.
authors.forename: François
authors.initials: F
authors.lastname: Goffinet
authors.affiliation: Laboratoire de cytogénétique, Hôpital Cochin-Maternité Port-Royal, AP-HP, Paris, France, Inserm U1016, Institut Cochin, Paris, France.
authors.forename: Laïla
authors.initials: L
authors.lastname: Allach El Khattabi
authors.affiliation: Service de biochimie et génétique moléculaire, HUPC Hôpital Cochin, Paris, France, Institut de myologie UPMC UM76 Inserm U974 CNRS UMR7215, GH Pitié-Salpêtrière, Paris, France.
authors.forename: France
authors.initials: F
authors.lastname: Leturcq
authors.affiliation: Service de biochimie et génétique moléculaire, HUPC Hôpital Cochin, Paris, France, EA7331, Faculté de pharmacie Paris, Université Paris-Descartes Sorbonne Paris-Cité, Paris, France.
authors.forename: Dominique
authors.initials: D
authors.lastname: Vidaud
authors.affiliation: Service de biochimie et génétique moléculaire, HUPC Hôpital Cochin, Paris, France, Inserm U1016, Institut Cochin, Paris, France.
authors.forename: Thierry
authors.initials: T
authors.lastname: Bienvenu
authors.affiliation: Maternité Cochin-Port Royal, HUPC Hôpital Cochin, Paris, France.
authors.forename: Vassilis
authors.initials: V
authors.lastname: Tsatsaris
authors.affiliation: Service de biochimie et génétique moléculaire, HUPC Hôpital Cochin, Paris, France, Inserm U1016, Institut Cochin, Paris, France.
authors.forename: Juliette
authors.initials: J
authors.lastname: Nectoux
mesh_terms: D001774:Blood Chemical Analysis / Q000379:methods; D004247:DNA / Q000032:analysis* / Q000378:metabolism; D005260:Female; D005333:Fetus / Q000737:chemistry* / Q000378:metabolism; D005602:France; D005838:Genotype; D060005:Genotyping Techniques / Q000379:methods; D006801:Humans; D062145:Maternal Serum Screening Tests / Q000379:methods*; D009035:Mothers; D011247:Pregnancy; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012204:Rh-Hr Blood-Group System / Q000097:blood / Q000235:genetics*; D012680:Sensitivity and Specificity
chemical_list: D012204:Rh-Hr Blood-Group System; D004247:DNA
keywords: RHD genotyping; droplet digital PCR; non-invasive prenatal diagnosis