Molecular Diagnosis & Therapy

, Volume 23, Issue 2, pp 291–299 | Cite as

Non-invasive Prenatal Testing Using Fetal DNA

  • Giulia Breveglieri
  • Elisabetta D’Aversa
  • Alessia Finotti
  • Monica BorgattiEmail author
Review Article


Non-invasive prenatal diagnosis (NIPD) is based on fetal DNA analysis starting from a simple peripheral blood sample, thus avoiding risks associated with conventional invasive techniques. During pregnancy, the fetal DNA increases to approximately 3–13% of the total circulating free DNA in maternal plasma. The very low amount of circulating cell-free fetal DNA (ccffDNA) in maternal plasma is a crucial issue, and requires specific and optimized techniques for ccffDNA purification from maternal plasma. In addition, highly sensitive detection approaches are required. In recent years, advanced ccffDNA investigation approaches have allowed the application of non-invasive prenatal testing (NIPT) to determine fetal sex, fetal rhesus D (RhD) genotyping, aneuploidies, micro-deletions and the detection of paternally inherited monogenic disorders. Finally, complex and innovative technologies such as digital polymerase chain reaction (dPCR) and next-generation sequencing (NGS) (exhibiting higher sensitivity and/or the capability to read the entire fetal genome from maternal plasma DNA) are expected to allow the detection, in the near future, of maternally inherited mutations that cause genetic diseases. The aim of this review is to introduce the principal ccffDNA characteristics and their applications as the basis of current and novel NIPT.


Compliance with ethical standards

Conflict of interest

Giulia Breveglieri, Elisabetta D’Aversa, Alessia Finotti, and Monica Borgatti have no conflicts of interest that are directly relevant to the content of this review.


This study was supported by the EU FP7 THALAMOSS Project (THALAssaemia MOdular Stratification System for personalized therapy of beta-thalassemia; grant number [306201]-FP7-Health-2012-INNOVATION-1). All funding bodies had no role in the design of the study, collection, analysis, and interpretation of data, or in writing the manuscript.


  1. 1.
    Lo YM, Corbetta N, Chamberlain PF, Rai V, Sargent IL, Redman CW, et al. Presence of fetal DNA in maternal plasma and serum. Lancet. 1997;350(9076):485–7.CrossRefPubMedGoogle Scholar
  2. 2.
    Mujezinovic F, Alfirevic Z. Procedure-related complications of amniocentesis and chorionic villous sampling: a systematic review. Obstet Gynecol. 2007;110(3):687–94.CrossRefPubMedGoogle Scholar
  3. 3.
    Perlado-Marina S, Bustamante-Aragones A, Horcajada L, Trujillo-Tiebas MJ, Lorda-Sanchez I, Ruiz Ramos M, et al. Overview of five-years of experience performing non-invasive fetal sex assessment in maternal blood. Diagnostics (Basel). 2013;3(2):283–90.CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Clausen FB, Damkjær MB, Dziegiel MH. Noninvasive fetal RhD genotyping. Transfus Apher Sci. 2014;50(2):154–62.CrossRefPubMedGoogle Scholar
  5. 5.
    Rolnik DL, da Silva Costa F, Lee TJ, Schmid M, McLennan AC. Association between fetal fraction on cell-free DNA testing and first trimester markers for pre-eclampsia. Ultrasound Obstet Gynecol. 2018;52(6):722–7. Scholar
  6. 6.
    van Boeckel SR, Davidson DJ, Norman JE, Stock SJ. Cell-free fetal DNA and spontaneous preterm birth. Reproduction. 2018;155(3):R137–45. Scholar
  7. 7.
    Skrzypek H, Hui L. Noninvasive prenatal testing for fetal aneuploidy and single gene disorders. Best Pract Res Clin Obstet Gynaecol. 2017;42:26–38.CrossRefPubMedGoogle Scholar
  8. 8.
    Bustamante-Aragonés A, et al. Non-invasive prenatal diagnosis of single-gene disorders from maternal blood. Gene. 2012;504(1):144–9.CrossRefPubMedGoogle Scholar
  9. 9.
    Hayward J, Chitty LS. Beyond screening for chromosomal abnormalities: Advances in non-invasive diagnosis of single gene disorders and fetal exome sequencing. Semin Fetal Neonatal Med. 2018;23(2):94–101. Scholar
  10. 10.
    Lee SY, Kim SJ, Han SH, Park JS, Choi HJ, Ahn JJ, et al. A new approach of digital PCR system for non-invasive prenatal screening of trisomy 21. Clin Chim Acta. 2018;476:75–80.CrossRefPubMedGoogle Scholar
  11. 11.
    Kagan KO, Sonek J, Wagner P, Hoopmann M. Principles of first trimester screening in the age of non-invasive prenatal diagnosis: screening for chromosomal abnormalities. Arch Gynecol Obstet. 2017;296(4):645–51.CrossRefPubMedGoogle Scholar
  12. 12.
    Hudecova I, Chiu RW. Non-invasive prenatal diagnosis of thalassemias using maternal plasma cell free DNA. Best Pract Res Clin Obstet Gynaecol. 2017;39:63–73.CrossRefPubMedGoogle Scholar
  13. 13.
    Perlado S, Bustamante-Aragonés A, Donas M, Lorda-Sánchez I, Plaza J, Rodríguez de Alba M. Fetal genotyping in maternal blood by digital PCR: towards NIPD of monogenic disorders independently of parental origin. PLoS One. 2016;11(4):e0153258.Google Scholar
  14. 14.
    Hui WW, Jiang P, Tong YK, Lee WS, Cheng YK, New MI, et al. Universal haplotype-based noninvasive prenatal testing for single gene diseases. Clin Chem. 2017;63(2):513–24.CrossRefPubMedGoogle Scholar
  15. 15.
    Lo YM, Tein MS, Lau TK, Haines CJ, Leung TN, Poon PM, et al. Quantitative analysis of fetal DNA in maternal plasma and serum: implications for noninvasive prenatal diagnosis. Am J Hum Genet. 1998;62(4):768–75.CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Bianchi DW, Flint AF, Pizzimenti MF, Knoll JH, Latt SA. Isolation of fetal DNA from nucleated erythrocytes in maternal blood. Proc Natl Acad Sci USA. 1990;87(9):3279–83.CrossRefPubMedGoogle Scholar
  17. 17.
    Lo YM. Fetal DNA in maternal plasma: biology and diagnostic applications. Clin Chem. 2000;46(12):1903–6.PubMedGoogle Scholar
  18. 18.
    Sekizawa A, Samura O, Zhen DK, Falco V, Farina A, Bianchi DW. Apoptosis in fetal nucleated erythrocytes circulating in maternal blood. Prenat Diagn. 2000;20(11):886–9.CrossRefPubMedGoogle Scholar
  19. 19.
    Sekizawa A, Yokokawa K, Sugito Y, Iwasaki M, Yukimoto Y, Ichizuka K, et al. Evaluation of bidirectional transfer of plasma DNA through placenta. Hum Genet. 2003;113(4):307–10.CrossRefPubMedGoogle Scholar
  20. 20.
    Jackson L. Fetal cells and DNA in maternal blood. Prenat Diagn. 2003;23(10):837–46.CrossRefPubMedGoogle Scholar
  21. 21.
    Alberry M, Maddocks D, Jones M, Abdel Hadi M, Abdel-Fattah S, Avent N, et al. Free fetal DNA in maternal plasma in anembryonic pregnancies: confirmation that the origin is the trophoblast. Prenat Diagn. 2007;27(5):415–8.CrossRefPubMedGoogle Scholar
  22. 22.
    Sandovici I, Hoelle K, Angiolini E, Constância M. Placental adaptations to the maternal-fetal environment: implications for fetal growth and developmental programming. Reprod Biomed Online. 2012;25(1):68–89.CrossRefPubMedGoogle Scholar
  23. 23.
    Lun FM, Chiu RW, Chan KC, Leung TY, Lau TK, Lo YM. Microfluidics digital PCR reveals a higher than expected fraction of fetal DNA in maternal plasma. Clin Chem. 2008;54(10):1664–72.CrossRefPubMedGoogle Scholar
  24. 24.
    Illanes S, Denbow M, Kailasam C, Finning K, Soothill PW. Early detection of cell-free fetal DNA in maternal plasma. Early Hum Dev. 2007;83(9):563–6.CrossRefPubMedGoogle Scholar
  25. 25.
    Zhou Y, Zhu Z, Gao Y, Yuan Y, Guo Y, Zhou L, et al. Effects of maternal and fetal characteristics on cell-free fetal DNA fraction in maternal plasma. Reprod Sci. 2015;22(11):1429–35.CrossRefPubMedGoogle Scholar
  26. 26.
    Vora NL, Johnson KL, Basu S, Catalano PM, Hauguel-De Mouzon S, Bianchi DW. A multifactorial relationship exists between total circulating cell-free DNA levels and maternal BMI. Prenat Diagn. 2012;32(9):912–4.PubMedPubMedCentralGoogle Scholar
  27. 27.
    Attilakos G, Maddocks DG, Davies T, Hunt LP, Avent ND, Soothill PW, et al. Quantification of free fetal DNA in multiple pregnancies and relationship with chorionicity. Prenat Diagn. 2011;31(10):967–72.CrossRefPubMedGoogle Scholar
  28. 28.
    Bischoff FZ, Lewis DE, Simpson JL. Cell-free fetal DNA in maternal blood: kinetics, source and structure. Hum Reprod Update. 2005;11(1):59–67.CrossRefPubMedGoogle Scholar
  29. 29.
    Chan KC, Zhang J, Hui AB, Wong N, Lau TK, Leung TN, et al. Size distributions of maternal and fetal DNA in maternal plasma. Clin Chem. 2004;50(1):88–92.CrossRefPubMedGoogle Scholar
  30. 30.
    Li Y, Zimmermann B, Rusterholz C, Kang A, Holzgreve W, Hahn S. Size separation of circulatory DNA in maternal plasma permits ready detection of fetal DNA polymorphisms. Clin Chem. 2004;50(6):1002–11.CrossRefPubMedGoogle Scholar
  31. 31.
    Angert RM, LeShane ES, Lo YM, Chan LY, Delli-Bovi LC, Bianchi DW. Fetal cell-free plasma DNA concentrations in maternal blood are stable 24 hours after collection: analysis of first- and third-trimester samples. Clin Chem. 2003;49(1):195–8.CrossRefPubMedGoogle Scholar
  32. 32.
    Lo YM, Zhang J, Leung TN, Lau TK, Chang AM, Hjelm NM. Rapid clearance of fetal DNA from maternal plasma. Am J Hum Genet. 1999;64(1):218–24.CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Hui L, Vaughan JI, Nelson M. Effect of labor on postpartum clearance of cell-free fetal DNA from the maternal circulation. Pren Diagn. 2008;28(4):304–8.CrossRefGoogle Scholar
  34. 34.
    Ordonez E, Rueda L, Cañadas MP, Fuster C, Cirigliano V. Evaluation of sample stability and automated DNA extraction for fetal sex determination using cell-free fetal DNA in maternal plasma. Biomed Res Int. 2013;2013:195363.CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Hidestrand M, Stokowski R, Song K, Oliphant A, Deavers J, Goetsch M, et al. Influence of temperature during transportation on cell-free DNA analysis. Fetal Diagn Ther. 2012;31(2):122–8.CrossRefPubMedGoogle Scholar
  36. 36.
    Zhang Y, Li Q, Hui N, Fei M, Hu Z, Sun S. Effect of formaldehyde treatment on the recovery of cell-free fetal DNA from maternal plasma at different processing times. Clin Chim Acta. 2008;397(1–2):60–4.CrossRefPubMedGoogle Scholar
  37. 37.
    Li Y, Di Naro E, Vitucci A, Zimmermann B, Holzgreve W, Hahn S. Detection of paternally inherited fetal point mutations for beta-thalassemia using size-fractionated cell-free DNA in maternal plasma. JAMA. 2005;293:843–9.CrossRefPubMedGoogle Scholar
  38. 38.
    Lun FMF, Tsui NB, Chan KC, Leung TY, Lau TK, Charoenkwan P, et al. Noninvasive prenatal diagnosis of monogenic diseases by digital size selection and relative mutation dosage on DNA in maternal plasma. Proc Natl Acad Sci USA. 2008;105:19920–5.CrossRefPubMedGoogle Scholar
  39. 39.
    Clausen FB, Krog GR, Rieneck K, Dziegiel MH. Improvement in fetal DNA extraction from maternal plasma. Evaluation of the NucliSens Magnetic Extraction system and the QIAamp DSP Virus Kit in comparison with the QIAamp DNA Blood Mini Kit. Prenat Diagn. 2007;27(1):6–10.Google Scholar
  40. 40.
    Stray J, Zimmermann B. Isolation of cell-free DNA from maternal plasma. Methods Mol Biol. 2019;1885:309–23. Scholar
  41. 41.
    Li Y, Holzgreve W, Page-Christiaens GC, Gille JJ, Hahn S. Improved prenatal detection of a fetal point mutation for achondroplasia by the use of size-fractionated circulatory DNA in maternal plasma–case report. Prenat Diagn. 2004;24(11):896–8.CrossRefPubMedGoogle Scholar
  42. 42.
    Li J, Makrigiorgos GM. COLD-PCR: a new platform for highly improved mutation detection in cancer and genetic testing. Biochem Soc Trans. 2009;37(Pt 2):427–32.CrossRefPubMedGoogle Scholar
  43. 43.
    Papageorgiou EA, Fiegler H, Rakyan V, Beck S, Hulten M, Lamnissou K, et al. Sites of differential DNA methylation between placenta and peripheral blood: molecular markers for noninvasive prenatal diagnosis of aneuploidies. Am J Pathol. 2009;174:1609–18.CrossRefPubMedPubMedCentralGoogle Scholar
  44. 44.
    Tong YK, Jin S, Chiu RW, Ding C, Chan KC, Leung TY, et al. Noninvasive prenatal detection of trisomy 21 by an epigenetic-genetic chromosome-dosage approach. Clin Chem. 2010;56(1):90–8.CrossRefPubMedGoogle Scholar
  45. 45.
    Ioannides M, Papageorgiou EA, Keravnou A, Tsaliki E, Spyrou C, Hadjidaniel M, et al. Inter-individual methylation variability in differentially methylated regions between maternal whole blood and first trimester CVS. Mol Cytogenet. 2014;7(1):73.CrossRefPubMedPubMedCentralGoogle Scholar
  46. 46.
    Pohl G, Shih IM. Principle and applications of digital PCR. Expert Rev Mol Diagn. 2004;4(1):41–7.CrossRefPubMedGoogle Scholar
  47. 47.
    Majumdar N, Banerjee S, Pallas M, Wessel T, Hegerich P. Poisson plus quantification for digital PCR systems. Sci Rep. 2017;7(1):9617. Scholar
  48. 48.
    Pinheiro LB, Coleman VA, Hindson CM, Herrmann J, Hindson BJ, Bhat S, et al. Evaluation of a droplet digital polymerase chain reaction format for DNA copy number quantification. Anal Chem. 2012;84:1003–11.CrossRefPubMedGoogle Scholar
  49. 49.
    Parkin B. Rare variant quantitation using droplet digital PCR. Methods Mol Biol. 2019;1881:239–51. Scholar
  50. 50.
    Hindson BJ, Ness KD, Masquelier DA, Belgrader P, Heredia NJ, Makarewicz AJ, et al. High-throughput droplet digital PCR system for absolute quantitation of DNA copy number. Anal Chem. 2011;83:8604–10.CrossRefPubMedPubMedCentralGoogle Scholar
  51. 51.
    D’Aversa E, Breveglieri G, Pellegatti P, Guerra G, Gambari R, Borgatti M. Non-invasive fetal sex diagnosis in plasma of early weeks pregnants using droplet digital PCR. Mol Med. 2018;24(1):14.CrossRefPubMedPubMedCentralGoogle Scholar
  52. 52.
    Lodrini M, Sprüssel A, Astrahantseff K, Tiburtius D, Konschak R, Lode HN, et al. Using droplet digital PCR to analyse MYCN and ALK copy number in plasma from patients with neuroblastoma. Oncotarget. 2017;8(49):85234–51.CrossRefPubMedPubMedCentralGoogle Scholar
  53. 53.
    Zhao G, Jiang T, Liu Y, Huai G, Lan C, Li G, et al. Droplet digital PCR-based circulating microRNA detection serve as a promising diagnostic method for gastric cancer. BMC Cancer. 2018;18:676.CrossRefPubMedPubMedCentralGoogle Scholar
  54. 54.
    Rutsaert S, Bosman K, Trypsteen W, Nijhuis M, Vandekerckhove L. Digital PCR as a tool to measure HIV persistence. Retrovirology. 2018;15:16.CrossRefPubMedPubMedCentralGoogle Scholar
  55. 55.
    Behjati S, Tarpey PS. What is next generation sequencing? Arch Dis Child Educ Pract Ed. 2013;98(6):236–8.CrossRefPubMedPubMedCentralGoogle Scholar
  56. 56.
    Mardis ER. Next-generation DNA sequencing methods. Annu Rev Genomics Hum Genet. 2008;9:387–402.CrossRefPubMedGoogle Scholar
  57. 57.
    McCarthy A. Third generation DNA sequencing: pacific biosciences’ single molecule real time technology. Chem Biol. 2010;17(7):675–6.CrossRefPubMedGoogle Scholar
  58. 58.
    Manegold-Brauer G, Hahn S, Lapaire O. What does next-generation sequencing mean for prenatal diagnosis? Biomark Med. 2014;8(4):499–508.CrossRefPubMedGoogle Scholar
  59. 59.
    Forest M, Morel Y, David M. Prenatal treatment of congenital adrenal hyperplasia. Trends Endocrinol Metab. 1998;9(7):284–9.CrossRefPubMedGoogle Scholar
  60. 60.
    Hyett JA, Gardener G, Stojilkovic-Mikic T, Finning KM, Martin PG, Rodeck CH, et al. Reduction in diagnostic and therapeutic interventions by non-invasive determination of fetal sex in early pregnancy. Pren Diagn. 2005;25:1111–6.CrossRefGoogle Scholar
  61. 61.
    Zhong XY, Holzgreve W, Hahn S. The levels of circulatory cell free fetal DNA in maternal plasma are elevated prior to the onset of preeclampsia. Hypertens Pregnancy. 2002;21(1):77–83.CrossRefPubMedGoogle Scholar
  62. 62.
    Shah VC, Smart V. Human chromosome Y and SRY. Cell Biol Int. 1996;20(1):3–6.CrossRefPubMedGoogle Scholar
  63. 63.
    Lo YMD, Patel P, Sampietro M, Gillmer MD, Fleming KA, Wainscoat JS. Detection of single-copy fetal DNA sequence from maternal blood. Lancet. 1990;335(8703):1463–4.CrossRefPubMedGoogle Scholar
  64. 64.
    Stanghellini I, Bertorelli R, Capone L, Mazza V, Neri C, Percesepe A, et al. Quantitation of fetal DNA in maternal serum during the first trimester of pregnancy by the use of a DAZ repetitive probe. Mol Hum Reprod. 2006;12(9):587–91.CrossRefPubMedGoogle Scholar
  65. 65.
    Devaney SA, Palomaki GE, Scott JA, Bianchi DW. Noninvasive fetal sex determination using cell-free fetal DNA: a systematic review and meta-analysis. JAMA. 2011;306(6):627–36.CrossRefPubMedPubMedCentralGoogle Scholar
  66. 66.
    Wright CF, Wei Y, Higgins JP, Sagoo GS. Non-invasive prenatal diagnostic test accuracy for fetal sex using cell-free DNA a review and meta-analysis. BMC Res Notes. 2012;5:476.CrossRefPubMedPubMedCentralGoogle Scholar
  67. 67.
    Breveglieri G, Bassi E, Carlassara S, Cosenza LC, Pellegatti P, Guerra G, et al. Y-chromosome identification in circulating cell-free fetal DNA using surface plasmon resonance. Prenat Diagn. 2016;36(4):353–61.CrossRefPubMedGoogle Scholar
  68. 68.
    Chim SSC, Tong YK, Chiu RW, Lau TK, Leung TN, Chan LY, et al. Detection of the placental epigenetic signature of the maspin gene in maternal plasma. Proc Natl Acad Sci USA. 2005;102(41):14753–8.CrossRefPubMedGoogle Scholar
  69. 69.
    Chan KC, Ding C, Gerovassili A, Yeung SW, Chiu RW, Leung TN, et al. Hypermethylated RASSF1A in maternal plasma: a universal fetal DNA marker that improves the reliability of noninvasive prenatal diagnosis. Clin Chem. 2006;52:2211–8.CrossRefPubMedGoogle Scholar
  70. 70.
    Bellido ML, Radpour R, Lapaire O, De Bie I, Hösli I, Bitzer J, et al. MALDI-TOF mass array analysis of RASSF1A and SERPINB5 methylation patterns in human placenta and plasma. Biol Reprod. 2010;82(4):745–50.CrossRefPubMedGoogle Scholar
  71. 71.
    Tang NL, Leung TN, Zhang J, Lau TK, Lo YM. Detection of fetal-derived paternally inherited X-chromosome polymorphisms in maternal plasma. Clin Chem. 1999;45(11):2033–5.PubMedGoogle Scholar
  72. 72.
    Tsui NB, Kadir RA, Chan KC, Chi C, Mellars G, Tuddenham EG, et al. Noninvasive prenatal diagnosis of hemophilia by microfluidics digital PCR analysis of maternal plasma DNA. Blood. 2011;117(13):3684–91.CrossRefPubMedGoogle Scholar
  73. 73.
    Lo YM, Hjelm NM, Fidler C, Sargent IL, Murphy MF, Chamberlain PF, et al. Prenatal diagnosis of fetal RhD status by molecular analysis of maternal plasma. N Engl J Med. 1998;339(24):1734–8.CrossRefPubMedGoogle Scholar
  74. 74.
    Scheffer PG, van der Schoot CE, Page-Christiaens GC, de Haas M. Noninvasive fetal blood group genotyping of rhesus D, c, E and of K in alloimmunised pregnant women: evaluation of a 7-year clinical experience. BJOG. 2011;118(11):1340–8.CrossRefPubMedGoogle Scholar
  75. 75.
    Kolialexi A, Tounta G, Mavrou A. Noninvasive fetal RhD genotyping from maternal blood. Expert Rev Mol Diagn. 2010;10(3):285–96.CrossRefPubMedGoogle Scholar
  76. 76.
    Tax MG, Van Der Schoot CE, Van Doorn R, Douglas-Berger L, Van Rhenen DJ, Maaskant-vanWijk PA. RHC and RHc genotyping in different ethnic groups. Transfusion. 2002;42(5):634–44.CrossRefPubMedGoogle Scholar
  77. 77.
    Clausen FB. Lessons learned from the implementation of non-invasive fetal RHD screening. Expert Rev Mol Diagn. 2018;18(5):423–31.CrossRefPubMedGoogle Scholar
  78. 78.
    Mennuti MT, Chandrasekaran S, Khalek N, Dugoff L. Cell-free DNA screening and sex chromosome aneuploidies. Prenat Diagn. 2015;35(10):980–5.CrossRefPubMedGoogle Scholar
  79. 79.
    Palomaki GE, Deciu C, Kloza EM, Lambert-Messerlian GM, Haddow JE, Neveux LM, et al. DNA sequencing of maternal plasma reliably identifies trisomy 18 and trisomy 13 as well as Down syndrome: an international collaborative study. Genet Med. 2012;14:296–305.CrossRefPubMedPubMedCentralGoogle Scholar
  80. 80.
    Swanson A, Sehnert AJ, Bhatt S. Non-invasive prenatal testing: technologies, clinical assays and implementation strategies for women’s healthcare practitioners. Curr Genet Med Rep. 2013;1(2):113–21.CrossRefPubMedPubMedCentralGoogle Scholar
  81. 81.
    Nicolaides KH, Syngelaki A, Gil M, Atanasova V, Markova D. Validation of targeted sequencing of single-nucleotide polymorphisms for non-invasive prenatal detection of aneuploidy of chromosomes 13, 18, 21, X, and Y. Prenat Diagn. 2013;33(6):575–9.CrossRefPubMedGoogle Scholar
  82. 82.
    Ryan A, Hunkapiller N, Banjevic M, Vankayalapati N, Fong N, Jinnett KN, et al. Validation of an enhanced version of a single-nucleotide polymorphism-based noninvasive prenatal test for detection of fetal aneuploidies. Fetal Diagn Ther. 2016;40(3):219–23.CrossRefPubMedGoogle Scholar
  83. 83.
    Bianchi DW, Parsa S, Bhatt S, Halks-Miller M, Kurtzman K, Sehnert AJ, et al. Fetal sex chromosome testing by maternal plasma DNA sequencing: clinical laboratory experience and biology. Obstet Gynecol. 2015;125(2):375–82.CrossRefPubMedGoogle Scholar
  84. 84.
    Taneja PA, Snyder HL, de Feo E, Kruglyak KM, Halks-Miller M, Curnow KJ, et al. Noninvasive prenatal testing in the general obstetric population: clinical performance and counseling considerations in over 85,000 cases. Prenat Diagn. 2016;36(3):237–43.CrossRefPubMedPubMedCentralGoogle Scholar
  85. 85.
    Stokowski R, Wang E, White K, Batey A, Jacobsson B, Brar H, et al. Clinical performance of non-invasive prenatal testing (NIPT) using targeted cell-free DNA analysis in maternal plasma with microarrays or next generation sequencing (NGS) is consistent across multiple controlled clinical studies. Prenat Diagn. 2015;35(12):1243–6.CrossRefPubMedPubMedCentralGoogle Scholar
  86. 86.
    Juneau K, Bogard PE, Huang S, Mohseni M, Wang ET, Ryvkin P, et al. Microarray-based cell-free DNA analysis improves non-invasive prenatal testing. Fetal Diagn Ther. 2014;36(4):282–6.CrossRefPubMedGoogle Scholar
  87. 87.
    Bianchi DW. From prenatal genomic diagnosis to fetal personalized medicine: progress and challenges. Nat Med. 2012;18(7):1041–51.CrossRefPubMedPubMedCentralGoogle Scholar
  88. 88.
    Ferrari M, Carrera P, Lampasona V, Galbiati S. New trend in non-invasive prenatal diagnosis. Clin Chim Acta. 2015;451(Pt A):9–13.CrossRefPubMedGoogle Scholar
  89. 89.
    Evans MI, Wright DA, Pergament E, Cuckle HS, Nicolaides KH. Digital PCR for non-invasive detection of aneuploidy: power analysis equations for feasibility. Fetal Diagn Ther. 2012;31(4):244–7.CrossRefPubMedGoogle Scholar
  90. 90.
    Traeger-Synodinos J. Real-time PCR for prenatal and preimplantation genetic diagnosis of monogenic diseases. Mol Aspects Med. 2006;27(2–3):176–91.CrossRefPubMedGoogle Scholar
  91. 91.
    Galbiati S, Monguzzi A, Damin F, Soriani N, Passiu M, Castellani C, et al. COLD-PCR and microarray: two independent highly sensitive approaches allowing the identification of fetal paternally inherited mutations in maternal plasma. J Med Genet. 2016;53(7):481–7.CrossRefPubMedGoogle Scholar
  92. 92.
    Mauger F, How-Kit A, Tost J. COLD-PCR technologies in the area of personalized medicine: methodology and applications. Mol Diagn Ther. 2017;21(3):269–83.CrossRefPubMedGoogle Scholar
  93. 93.
    Guissart C, Debant V, Desgeorges M, Bareil C, Raynal C, Toga C, et al. Non-invasive prenatal diagnosis of monogenic disorders: an optimized protocol using MEMO qPCR with miniSTR as internal control. Clin Chem Lab Med. 2015;53(2):205–15.CrossRefPubMedGoogle Scholar
  94. 94.
    Phylipsen M, Yamsri S, Treffers EE, Jansen DT, Kanhai WA, Boon EM, et al. Non-invasive prenatal diagnosis of beta-thalassemia and sickle-cell disease using pyrophosphorolysis-activated polymerization and melting curve analysis. Prenat Diagn. 2012;32(6):578–87.CrossRefPubMedGoogle Scholar
  95. 95.
    Lo YM, Chan KC, Sun H, Chen EZ, Jiang P, Lun FM, et al. Maternal plasma DNA sequencing reveals the genome-wide genetic and mutational profile of the fetus. Sci Transl Med. 2010;2(61):61ra91.Google Scholar
  96. 96.
    Breveglieri G, Travan A, D’Aversa E, Cosenza LC, Pellegatti P, Guerra G, et al. Postnatal and non-invasive prenatal detection of β-thalassemia mutations based on Taqman genotyping assays. PLoS One. 2017;12(2):e0172756.CrossRefPubMedPubMedCentralGoogle Scholar
  97. 97.
    Li Y, Di Naro E, Vitucci A, Grill S, Zhong XY, Holzgreve W, et al. Size fractionation of cell-free DNA in maternal plasma improves the detection of a paternally inherited beta-thalassemia point mutation by MALDI–TOF mass spectrometry. Fetal Diagn Ther. 2009;25(2):246–9.CrossRefPubMedGoogle Scholar
  98. 98.
    Zhong XY, Holzgreve W. MALDI–TOF MS in prenatal genomics. Transfus Med Hemother. 2009;36(4):263–72.CrossRefPubMedPubMedCentralGoogle Scholar
  99. 99.
    Papasawa T, Kalikas I, Kyrri A, Kleanthous M. Arrayed primer extension for the noninvasive prenatal diagnosis of beta-thalassemia based on detection of single nucleotide polymorphisms. Ann N Y Acad Sci. 2008;1137:302–8.CrossRefGoogle Scholar
  100. 100.
    Yi P, Chen Z, Zhao Y, Guo J, Fu H, Zhou Y, et al. PCR/LDR/capillary electrophoresis for detection of single-nucleotide differences between fetal and maternal DNA in maternal plasma. Prenat Diagn. 2009;29(3):217–22.CrossRefPubMedGoogle Scholar
  101. 101.
    Camunas-Soler J, Lee H, Hudgins L, Hintz SR, Blumenfeld YJ, El-Sayed YY, et al. Noninvasive prenatal diagnosis of single-gene disorders by use of droplet digital PCR. Clin Chem. 2018;64(2):336–45. Scholar
  102. 102.
    Nectoux J. Current, emerging, and future applications of digital PCR in non-invasive prenatal diagnosis. Mol Diagn Ther. 2018;22(2):139–48. Scholar
  103. 103.
    Hahn S, Hösli I, Lapaire O. Non-invasive prenatal diagnostics using next generation sequencing: technical, legal and social challenges. Expert Opin Med Diagn. 2012;6(6):517–28.CrossRefPubMedGoogle Scholar
  104. 104.
    Chiu RW, Chan KC, Gao Y, Lau VY, Zheng W, Leung TY, 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(51):20458–63.CrossRefPubMedGoogle Scholar
  105. 105.
    Wong FC, Lo YM. Prenatal diagnosis innovation: genome sequencing of maternal plasma. Annu Rev Med. 2016;67:419–32.CrossRefPubMedGoogle Scholar
  106. 106.
    Strom CM, Anderson B, Tsao D, Zhang K, Liu Y, Livingston K, et al. Improving the positive predictive value of non-invasive prenatal screening (NIPS). PLoS One. 2017;12(3):e0167130. Scholar

Copyright information

© Springer Nature Switzerland AG 2019

Authors and Affiliations

  1. 1.Department of Life Sciences and BiotechnologyUniversity of FerraraFerraraItaly
  2. 2.Interuniversity Consortium for Biotechnologies (CIB)TriesteItaly
  3. 3.Biotechnology CenterUniversity of FerraraFerraraItaly

Personalised recommendations